Clsi TDM6 A

T/DM6-A
Vol.17 No.14
Replaces T/DM6-P
September 1997 Vol. 8 No. 10
Blood Alcohol Testing in the Clinical Laboratory;

Approved Guideline
This guideline provides technical and administrative guidance on laboratory procedures related to blood
alcohol testing, including specimen collection, methods of analysis, quality assurance, and reporting of
results.
ABC
NCCLS...
Serving the World's Medical Science Community Through Voluntary Consensus
NCCLS is an international, interdisciplinary, nonprofit, Proposed An NCCLS consensus document undergoes the first
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September 1997 T/DM6-A

Approved Guideline
Abstract
T/DM6-A, Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline, is designed to aid the
clinical laboratory in producing timely and accurate blood alcohol results. Its key objective is to address, as
comprehensively as possible, recommendations to assure the integrity of the laboratory report on blood
alcohol. The document conforms to the objective by addressing specimen collection, methods of analysis,
quality assurance, and reporting and significance of results as separate sections. Statutory provisions are
included as additional resource information.
The subcommittee recognizes the possible medicolegal impact of blood alcohol testing. The section
devoted to the chain-of-custody strives to define the laboratory's responsibility regarding the specimen by
outlining specific procedures for the handling and storage of the specimen and subsequent documentation.
[NCCLS. Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline. NCCLS Document T/DM6-A
(ISBN 1-56238-333-7). NCCLS, 940 West Valley Road, Suite 1400, Wayne, PA 19087-1898 USA,
1997.]
THE NCCLS consensus process, which is the mechanism for moving a document through two or
more levels of review by the healthcare community, is an ongoing process. Users should expect
revised editions of any given document. Because rapid changes in technology may affect the
procedures, methods, and protocols in a standard or guideline, users should replace outdated
editions with the current editions of NCCLS documents. Current editions are listed in the NCCLS
Catalog, which is distributed to member organizations, and to nonmembers on request. If your
organization is not a member and would like to become one, and to request a copy of the NCCLS
Catalog, contact the NCCLS Executive Offices. Telephone: 610.688.0100; Fax: 610.688.0700;
E-Mail: exoffice@nccls.org.
NCCLS VOL.17 NO.14 i

NCCLS VOL.17 NO.14 ii

T/DM6-A
ISBN 1-56238-333-7
September 1997 ISSN 0273-3099

Approved Guideline
Volume 17 Number 14
Kurt M. Dubowski, Ph.D.

Patricia H. Field, Ph.D.
Walter Frajola, Ph.D.
Vidmantas A. Raisys, Ph.D.
Robert H. Reeder, J.D.
M. Jeffery Shoemaker, Ph.D.
Vana L. Smith, Ph.D.
ABC
This publication is protected by copyright. No part of it may be reproduced, stored in a retrieval system, or
transmitted in any form or by any means (electronic, mechanical, photocopying, recording, or otherwise)
without written permission from NCCLS, except as stated below.
NCCLS hereby grants permission to reproduce limited portions of this publication for use in laboratory
procedure manuals at a single site, for interlibrary loan, or for use in educational programs provided that
multiple copies of such reproduction shall include the following notice, be distributed without charge, and,
in no event, contain more than 20% of the document's text.
Reproduced with permission, from NCCLS publication T/DM6-A, Blood Alcohol Testing in the
Clinical Laboratory; Approved Guideline. Copies of the current edition may be obtained from
NCCLS, 940 West Valley Road, Suite 1400, Wayne, Pennsylvania 19087-1898 USA.
Permission to reproduce or otherwise use the text of this document to an extent that exceeds the
exemptions granted here or under the Copyright Law must be obtained from NCCLS by written request. To
request such permission, address inquiries to the Executive Director, NCCLS, 940 West Valley Road, Suite
1400, Wayne, Pennsylvania 19087-1898 USA.
Copyright ©1997. The National Committee for Clinical Laboratory Standards.
Suggested Citation
NCCLS. Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline. NCCLS Document T/DM6-A
(ISBN 1-56238-333-7). NCCLS, 940 West Valley Road, Suite 1400, Wayne, PA 19087-1898 USA, 1997.
Proposed Guideline
December 1988
Approved Guideline
September 1997
ISBN 1-56238-333-7
ISSN 0273-3099
NCCLS VOL.17 NO.14 iv

Committee Membership
Area Committee on Clinical Chemistry and Toxicology
Basil T. Doumas, Ph.D. Medical College of Wisconsin

Chairholder Milwaukee, Wisconsin
W. Gregory Miller, Ph.D. Medical College of Virginia Hospital

Vice Chairholder Richmond, Virginia
Subcommittee on Blood Alcohol Testing
Kurt M. Dubowski, Ph.D. The University of Oklahoma

Chairholder Oklahoma City, Oklahoma
Patricia H. Field, Ph.D. WI State Laboratory of Hygiene

Madison, Wisconsin
Walter Frajola, Ph.D. Columbus, Ohio
Vidmantas A. Raisys, Ph.D. University of Washington

Seattle, Washington
Robert H. Reeder, J.D. Northwestern University

Evanston, Illinois
M. Jeffery Shoemaker, Ph.D. Pennsylvania Dept. of Health

Lionville, Pennsylvania
Vana L. Smith, Ph.D. Abbott Laboratories

Abbott Park, Illinois
Sharon S. Ehrmeyer, Ph.D. University of Wisconsin

Board Liaison Madison, Wisconsin
Beth Ann Wise, M.T.(ASCP), M.S.Ed. NCCLS

Staff Liaison Wayne, Pennsylvania
Patrice E. Polgar NCCLS

Editor Wayne, Pennsylvania
NCCLS VOL.17 NO.14 v

ACTIVE MEMBERSHIP (as of 1 July 1997)
Sustaining Members College of Physicians and Government of Malta

Surgeons of Saskatchewan Department of Health
American Association for Commission on Office Health Care Financing
Clinical Chemistry Laboratory Accreditation Administration
Bayer Corporation Institut für Stand. und Dok. im INMETRO
Beckman Instruments, Inc. Med. Lab. (INSTAND) Instituto de Salud Publica de
Becton Dickinson and Company International Council for Chile
Boehringer Mannheim Standardization in Instituto Scientifico HS. Raffaele
Diagnostics, Inc. Haematology (Italy)
College of American International Federation of Iowa State Hygienic Laboratory
Pathologists Clinical Chemistry Manitoba Health
Coulter Corporation International Society for Massachusetts Department of
Dade International Inc. Analytical Cytology Public Health Laboratories
Johnson & Johnson Clinical Italian Society of Clinical Michigan Department of Public
Diagnostics Biochemistry Health
Ortho Diagnostic Systems Inc. Japan Assn. Of Medical National Health Laboratory
Technologists (Osaka) (Luxembourg)
Professional Members Japanese Assn. Of Medical National Institute of Standards
Technologists (Tokyo) and Technology
American Academy of Family Japanese Committee for Clinical Ohio Department of Health
Physicians Laboratory Standards Oklahoma State Department of
American Association of Joint Commission on Health
Bioanalysts Accreditation of Healthcare Ontario Ministry of Health
American Association of Blood Organizations Ordre professionnel des
Banks National Academy of Clinical technologistes médicaux du
American Association for Biochemistry Québec
Clinical Chemistry National Society for Saskatchewan Health-
American Association for Histotechnology, Inc. Government of Saskatchewan
Respiratory Care Ontario Medical Association South African Institute for
American Chemical Society Laboratory Proficiency Testing Medical Research
American Medical Technologists Program Swedish Institute for Infectious
American Public Health Sociedade Brasileira de Analises Disease Control
Association Clinicas
American Society for Clinical Sociedad Espanola de Quimica Industry Members
Laboratory Science Clinica
American Society of VKCN (The Netherlands) AB Biodisk
Hematology Abbott Laboratories
American Society for Government Members ABC Consulting Group, Ltd.
Microbiology AccuMed International, Inc.
American Society of Armed Forces Institute of Aejes
Parasitologists, Inc. Pathology Ammirati Regulatory Consulting
American Type Culture Association of State and Atlantis Laboratory Systems
Collection, Inc. Territorial Public Health Avecor Cardiovascular, Inc.
ASQC Food, Drug and Cosmetic Laboratory Directors Bayer Corporation - Elkhart, IN
Division BC Centre for Disease Control Bayer Corporation - Middletown,
Asociacion Espanola Primera de Centers for Disease Control and VA
Socorros Prevention Bayer Corporation - Tarrytown,
Association Microbiology Clinici China National Centre for the NY
Italiani-A.M.C.L.I. Clinical Laboratory Bayer Corporation - West
Australasian Association of Commonwealth of Pennsylvania Haven, CT
Clinical Biochemists Bureau of Laboratories Bayer-Sankyo Co., Ltd.
Canadian Society of Laboratory Department of Veterans Affairs Beckman Instruments, Inc.
Technologists Deutsches Institut für Normung Becton Dickinson and Company
Clinical Laboratory Management (DIN) Becton Dickinson Consumer
Association FDA Center for Devices and Products
College of American Radiological Health Becton Dickinson
Pathologists FDA Division of Anti-Infective Immunocytometry Systems
College of Medical Laboratory Drug Products Becton Dickinson Italia S.P.A.
Technologists of Ontario
NCCLS VOL.17 NO.14 vi

Becton Dickinson Microbiology Hoechst Marion Roussel, Inc. Roche Diagnostic Systems
Systems Hybritech, Incorporated (Div. Hoffmann-La Roche
Becton Dickinson VACUTAINER Hycor Biomedical Inc. Inc.)
Systems I-STAT Corporation Roche Laboratories (Div.
Behring Diagnostics Inc. Integ, Inc. Hoffmann-La Roche Inc.)
Behring Diagnostics Inc. - San International Biomedical ROSCO Diagnostica
Jose, CA Consultants The R.W. Johnson
bioMérieux Vitek, Inc. International Remote Imaging Pharmaceutical Research
Biometrology Consultants Systems (IRIS) Institute (Div. Ortho Diagnostic
Bio-Rad Laboratories, Inc. International Technidyne Systems Inc.)
Bio-Reg Assoicates, Inc. Corporation Sarstedt, Inc.
Biosite Diagnostics Johnson & Johnson Clinical Schering Corporation
Biotest AG Diagnostics Schleicher & Schuell, Inc.
Boehringer Mannheim Johnson & Johnson Health Care Second Opinion
Diagnostics, Inc. Systems, Inc. SenDx Medical, Inc.
Boehringer Mannheim GmbH Kimble/Kontes Sherwood - Davis & Geck
Bristol-Myers Squibb Company Labtest Sistemas Diagnosticos Shionogi & Company, Ltd.
Canadian Reference Laboratory Ltda. Showa Yakuhin Kako Company,
Ltd. LifeScan, Inc. (Sub. Ortho Ltd.
CASCO Standards Diagnostic Systems Inc.) Sienna Biotech
Checkpoint Development, Inc. Lilly Research Laboratories SmithKline Beecham
ChemTrak Luminex Corporation Corporation
Chiron Diagnostics Corporation Mallinckrodt Sensor Systems SmithKline Beecham, S.A.
Chiron Diagnostics Corporation - MBG Industries, Inc. SmithKline Diagnostics, Inc.
International Operations Medical Device Consultants, (Sub. Beckman Instruments,
Chiron Diagnostics Corporation - Inc. Inc.)
Reagent Systems Medical Laboratory Automation SRL, Inc.
Cholestech Inc. Streck Laboratories, Inc.
Clinical Lab Engineering MediSense, Inc. Sumitomo Metal Bioscience Inc.
COBE Laboratories, Inc. Merck & Company, Inc. Sysmex Corporation
Cosmetic Ingredient Review Metra Biosystems TOA Medical Electronics
Coulter Corporation Neometrics, Inc. Unipath Co (Oxoid Division)
Cytometrics, Inc. Nissui Pharmaceutical Co., Ltd. Vetoquinol S.A.
CYTYC Corporation Norfolk Associates, Inc. Vysis, Inc.
Dade International - Deerfield, IL North American Biologicals, Inc. Wallac Oy
Dade International - Glasgow, Olympus Corporation Warner-Lambert Company
DE Optical Sensors, Inc. Wyeth-Ayerst
Dade International - Miami, FL Organon Teknika Corporation Xyletech Systems, Inc.
Dade International - Orion Diagnostica, Inc. Yeongdong Pharmaceutical
Sacramento, CA Ortho Diagnostic Systems Inc. Corp.
DAKO A/S Otsuka America Pharmaceutical, Zeneca
Diagnostic Products Corporation Inc.
Diametrics Medical, Inc. Pfizer Canada, Inc. Trade Associations
Difco Laboratories, Inc. Pfizer Inc
Direct Access Diagnostics Pharmacia & Upjohn - Michigan Association of Medical
Eiken Chemical Company, Ltd. Pharmacia & Upjohn - Sweden Diagnostic Manufacturers
Enterprise Analysis Corporation Procter & Gamble Health Industry Manufacturers
Donna M. Falcone Consultants Pharmaceuticals, Inc. Association
Fujisawa Pharmaceutical Co. The Product Development Group Japan Association of Clinical
Ltd. Radiometer America, Inc. Reagents Industries (Tokyo)
Gen-Probe Radiometer Medical A/S Medical Industry Association
Glaxo, Inc. Research Inc. of Australia
H&S Consultants David G. Rhoads Associates, National Association of Testing
Health Systems Concepts, Inc. Inc. Authorities - Australia
Helena Laboratories Rhône-Poulenc Rorer
Higman Healthcare
NCCLS VOL.17 NO.14 vii

Associate Active Members Ellis Fischel Cancer Center (MO) New Jersey Department of
Elyria Memorial Hospital (OH) Health
Affinity Health System (WI) Evanston Hospital (IL) The New York Blood Center
Allegheny University of the Federal Medical Center (MN) New York State Department of
Health Sciences (PA) Fort Leonard Wood Army Health
Allergy Testing Laboratory (TX) Community Hospital (MO) New York State Library
Alton Ochsner Medical Frye Regional Medical Center New York University Medical
Foundation (LA) (NC) Center
American Oncologic Hospital Grady Memorial Hospital (GA) North Carolina Laboratory of
(PA) Great Smokies Diagnostic Public Health
Anzac House (Australia) Laboratory (NC) North Central Bronx Hospital
Associated Regional & Hartford Hospital (CT) (NY)
University Pathologists (UT) Heritage Hospital (MI) Northwestern Memorial Hospital
Baptist Medical Center of Hopital Saint Pierre (Belgium) (IL)
Oklahoma Hunter Area Pathology Service Olin E. Teague Medical Center
Baptist Memorial Healthcare (Australia) (TX)
System (TX) Incstar Corporation (MN) Omni Laboratory (MI)
BC Children’s Hospital (Canada) International Health Our Lady of Lourdes Hospital
Bethesda Hospital (OH) Management Associates, Inc. (NJ)
Beth Israel Medical Center (NY) (IL) Our Lady of the Resurrection
Bristol Regional Medical Center Jewish Hospital of Cincinnati Medical Center (IL)
(TN) (OH) Palo Alto Medical Foundation
Brooke Army Medical Center Kaiser Permanente (CA) (CA)
(TX) Kangnam St. Mary’s Hospital PAPP Clinic P.A. (GA)
Brooks Air Force Base (TX) (Korea) Pathogenesis Corp. (WA)
Broward General Medical Center Kenora-Rainy River Regional Pathology Associates
(FL) Laboratory Program (Dryden, Laboratories (CA)
Canterbury Health Laboratories ON, Canada) Permanente Medical Group (CA)
(New Zealand) Klinisches Institute für PLIVA d.d. Research Institute
Central Peninsula General Medizinische (Austria) (Croatia)
Hospital (AK) Laboratoire de Santé Publique Polly Ryon Memorial Hospital
CENTREX Clinical Laboratories du Quebec (Canada) (TX)
(NY) Laboratory Corporation of Providence Medical Center (WA)
Childrens Hospital Los Angeles America (NC) Puckett Laboratories (MS)
(CA) Lancaster General Hospital (PA) Queens Hospital Center (NY)
Children's Hospital Medical Los Angeles County & USC Quest Diagnostics (MI)
Center (Akron, OH) Medical Center (CA) Ravenswood Hospital Medical
Children's Hospital Medical Louisiana State University Center (IL)
Center (Cincinnati, OH) Medical Center Reid Hospital & Health Care
Children's Hospital - New Maine Medical Center Services (IN)
Orleans (LA) Malcolm Grow USAF Medical Riverside Clinical Laboratories
City Hospital (WV) Center (MD) (VA)
The Cleveland Clinic Foundation The Medical Center of Ocean Royal Brisbane Hospital
(OH) County (NJ) (Australia)
Columbia Medical Center (FL) Melbourne Pathology (Australia) St. Boniface General Hospital
Commonwealth of Kentucky Memorial Medical Center (IL) (Winnipeg, Canada)
CompuNet Clinical Laboratories Memorial Medical Center (LA) St. Francis Medical Center (CA)
(OH) Mercy Hospital (MN) St. John Regional Hospital (St.
Consultants Laboratory (WI) Methodist Hospital (TX) John, NB, Canada)
Dean Medical Center (WI) Methodist Hospitals of Memphis St. John’s Regional Health
Detroit Health Department (MI) (TN) Center (MO)
Dhahran Health Center (Saudi Montreal Children’s Hospital St. Luke’s Hospital (PA)
Arabia) (Canada) St. Luke’s Regional Medical
Diagnostic Systems Mount Sinai Hospital (NY) Center (IA)
Laboratories, Inc. (TX) Mount Sinai Hospital (Toronto, St. Luke’s-Roosevelt Hospital
Duke University Medical Center ON, Canada) Center (NY)
(NC) National Genetics Institute (CA) St. Mary of the Plains Hospital
Dunn Memorial Hospital (IN) Naval Hospital Cherry Point (NC) (TX)
Dwight David Eisenhower Army New Britain General Hospital St. Paul Ramsey Medical Center
Medical Center (Ft. Gordon, (CT) (MN)
GA) New Hampshire Medical St. Vincent Medical Center (CA)
Easton Hospital (PA) Laboratories
NCCLS VOL.17 NO.14 viii

San Francisco General Hospital University of Cincinnati Medical UZ-KUL Medical Center
(CA) Center (OH) (Belgium)
Seoul National University University of Florida VA (Albuquerque) Medical
Hospital (Korea) University Hospital (Gent) Center (NM)
Shanghai Center for the Clinical (Belgium) VA (Denver) Medical Center
Laboratory (China) University Hospital (Linkoping, (CO)
Shore Memorial Hospital (NJ) Sweden) VA (Long Beach) Medical Center
SmithKline Beecham Clinical University Hospital (London, (CA)
Laboratories (GA) ON, Canada) VA (Miami) Medical Center (FL)
SmithKline Beecham Clinical University Hospital (IN) Venice Hospital (FL)
Laboratories (TX) University Hospital of Veterans General Hospital
South Bend Medical Foundation Cleveland (OH) (Republic of China)
(IN) The University Hospitals (OK) Virginia Baptist Hospital
Southeastern Regional Medical University of Medicine & Warde Medical Laboratory (MI)
Center (NC) Dentistry, NJ University William Beaumont Hospital (MI)
Southern California Permanente Hospital Winn Army Community Hospital
Medical Group University of Nebraska Medical (GA)
SUNY @ Stony Brook (NY) Center Wisconsin State Laboratory of
Travis Air Force Base (CA) University of the Ryukyus Hygiene
UNC Hospitals (NC) (Japan) Yonsei University College of
University of Alberta Hospitals The University of Texas Medical Medicine (Korea)
(Canada) Branch York Hospital (PA)
University of California, San University of Virginia Medical Zale Lipshy University Hospital
Francisco Center (TX)
U.S. Army Hospital, Heidelberg
OFFICERS BOARD OF DIRECTORS
A. Samuel Koenig, III, M.D., Carl A. Burtis, Ph.D. Robert F. Moran, Ph.D.,
President Oak Ridge National Laboratory FCCM, FAIC
Family Medical Care mvi Sciences
Sharon S. Ehrmeyer, Ph.D.
William F. Koch, Ph.D., University of Wisconsin David E. Nevalainen, Ph.D.
President Elect Abbott Laboratories
National Institute of Standards Elizabeth D. Jacobson, Ph.D.
and Technology FDA Center for Devices and Donald M. Powers, Ph.D.
Radiological Health Johnson & Johnson Clinical
F. Alan Andersen, Ph.D., Diagnostics
Secretary Hartmut Jung, Ph.D.
Cosmetic Ingredient Review Boehringer Mannaheim GmbH Eric J. Sampson, Ph.D.
Centers for Disease Control
Donna M. Meyer, Ph.D., Tadashi Kawai, M.D., Ph.D. and Prevention
Treasurer International Clinical Pathology
Sisters of Charity Health Care Center Marianne C. Watters,
System M.T.(ASCP)
Kenneth D. McClatchey, M.D., Parkland Memorial Hospital
Charles F. Galanaugh, Past D.D.S.
President Loyola University Medical Ann M. Willey, Ph.D.
Becton Dickinson and Center New York State Department of
Company (Retired) Health
John V. Bergen, Ph.D.,

Executive Director
NCCLS VOL.17 NO.14 ix

Contents
Page
Abstract . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .... . . .I
Committee Membership . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .... . . v
Active Membership . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .... . . vi
Foreword . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .... . . xi
1 Scope and Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.1 Scope of the Problem . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

1.2 Medical Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1
1.3 Industrial-Medical Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
1.4 Medicolegal Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2
2 Specimen Collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.1 Responsibility for Specimen Collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

2.2 Types of Specimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3
2.3 Specimen Collection, Handling, and Preservation . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5
2.4 Specimen Handling . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
2.5 Replicate Blood Specimens . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
3 Chain-of-Custody Procedures . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
3.1 Purpose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6
3.2 Documentation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7
4 Methods of Analysis . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8
4.1 Gas Chromatography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

4.2 Enzymatic Oxidation with Alcohol Dehydrogenase . . . . . . . . . . . . . . . . . . . . . . . . . . . 9
5 Quality Assurance . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11
5.1 Calibrators (Standards) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

5.2 Controls . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
6 Reporting and Significance of Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12
6.1 Blood Alcohol Records . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

6.2 Conversions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
7 Terminology and Statutory Provisions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
7.1 Terminology . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
7.2 Statutory Provisions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15
Appendix A: Stages of Acute Alcoholic Influence/Intoxication . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 17
Appendix B: Guide to Serum-Alcohol Test Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18
Appendix C: Special Specimen Handling Considerations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
Summary of Comments and Subcommittee Responses . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 20
Related NCCLS Publications . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28
NCCLS VOL.17 NO.14 x

Foreword
This guideline, Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline, was developed in
response to the frequently expressed need for a readily available information source which addresses the
increasingly more frequent involvement of the hospital or independent clinical laboratory in collecting and
analyzing blood (and other biological specimens) for ethyl alcohol. The presence of alcohol is frequently
associated with trauma, and with a great variety of acute illnesses and chronic diseases. Further, alcohol
presence often adversely affects both morbidity and mortality. Therefore, appropriate trauma care and
diagnosis and treatment of many medical syndromes and diseases in adults require information about the
patient's alcohol status. Demand for blood alcohol testing of patients can, therefore, be expected to
continue to increase.
Years of experience have borne out the expectation that the absence, or presence and concentration of
alcohol in blood will often have later medicolegal or forensic implications and significance, in addition to its
immediate clinical relevance. Some simple and practical measures taken at the outset can greatly reduce
the impact of such subsequent legal developments on the clinical laboratory and its personnel. This
guideline addresses that issue and the resultant responsibilities of clinical laboratories which are not limited
to, but include the collection of blood specimens, quality assurance, records, and reports.
The Subcommittee on Blood Alcohol Testing has endeavored to produce a brief but adequate set of criteria
to assist clinical laboratories in meeting the demand for timely and reliable blood alcohol testing for clinical
purposes, while minimizing the impact of later medicolegal developments on the laboratory.
Universal Precautions
Because it is often impossible to know which might be infectious, all patient blood specimens are to be
treated with "universal precautions." Guidelines for specimen handling are available from the U.S. Centers
for Disease Control and Prevention. NCCLS document M29, Protection of Laboratory Workers from
Infectious Disease Transmitted by Blood, Body Fluids, and Tissue, deals specifically with all aspects of this
issue.
Key Words
Alcohol, blood alcohol, analysis, blood alcohol concentration, chain-of-custody, intoxication.
NCCLS NO.17 VOL.14 xi

Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline

1 Scope and Requirements Acute Alcoholic Influence/Intoxication3 (Appendix A
and the Guide to Serum-Alcohol Test Results
1.1 Scope of the Problem (Appendix B) become important in the evaluation
of the patient, for they define possible
The cost of alcohol abuse in our society is extra- correlations of blood alcohol concentration with
ordinarily high in terms of the loss of human life, its clinical signs and symptoms.
its detrimental contribution to the causes of
illness and injury, productivity losses in the Because alcoholism can masquerade as many
workplace, and the stress these in turn put on our other diseases, vital signs become very important
medical resources and our judicial system. In a during the physical examination, and the
series of special workshops on alcoholism and possibility of closed head injury or neurological
alcohol abuse, the American Society of Clinical disorder should be considered. Associated
Pathologists Task Force on Drug Abuse and disease diagnoses include neurological disorders;
Toxicology1 recognized the magnitude of the alcohol-induced or nonalcohol-associated
problem of alcohol in our society. The title of cardiovascular disorders; arrhythmias,
these seminars was "Alcohol - The Second Great tachycardias, electrocardiographic alterations;
Imitator" because of the medical challenges in the liver disease; fatty liver, alcoholic hepatitis, portal
diagnosis of this problem. fibrosis, cirrhosis and possible liver carcinoma;
blood and blood clotting disorders: anemia,
1.2 Medical Requirements prothrombin elevations and thrombocytopenia;
alcoholic pancreatitis; infections; alcoholic
Alcohol abuse should be considered one of the myopathies; digestive tract disorders: ulcers,
most important contributory causes of injury and gastritis, esophagitis, esophageal varices and
disease today. The diagnosis of alcohol-use cancer; endocrine disorders; skeletal system
disorders, such as alcohol dependence and disorders: ischemic necrosis of the femoral heads
alcohol abuse, is a clinical procedure and subject and fractures; skin disorders; and toxic
to the problem of inexactness.2 Diagnosis of the psychoses.
disease may be based not only on the features of
alcoholism, with all its signs and symptoms, but Indicated laboratory studies include analysis for
on an accurate determination of blood alcohol blood alcohol concentration and other relevant
concentration at the time of examination of a clinical laboratory tests.
patient.
The evaluation of the comatose patient may be
The use of alcohol acutely affects the central more difficult due to the lack of patient history.
nervous system. Many of the signs and The physical examination and results of
symptoms manifested by the patient are related laboratory studies often reveal the diagnosis.
to the degree of intoxication, as reflected by the These should be combined with radiological
concentration of alcohol in the patient's blood. studies, particularly of the skull and chest. A
Since many patients with traumatic injuries who proper evaluation of these studies will be valuable
are admitted to the emergency department are for decisions regarding admission to the hospital,
noncomatose, patient history or initial interview proper treatment, and to minimize possible
becomes most important. Patients showing medical and legal complications.
direct effects of alcohol—breath odor, released
inhibitions, alcoholic facies, toxic amblyopia, This document necessarily emphasizes certain
possible tachycardia and cardiac arrhythmias, legal and other nonclinical aspects of blood
nystagmus, and traumatic injuries of varying alcohol testing. It is, therefore, important to
degrees—warrant further studies. These studies recognize at all times that the clinical laboratory’s
should include a complete physical examination, first and primary responsibility is to the patient
clinical laboratory analyses, and determination of and to the physician.
the blood or serum alcohol concentration. Other
drugs of misuse or abuse should also be
considered in the testing procedures. The
information in Dubowski's table on the Stages of
NCCLS NO.17 VOL.14 1

1.3 Industrial-Medical Requirements exclusively legal purposes such as traffic law

enforcement or for potential adversarial
Alcohol is the most commonly used drug in the proceedings such as accident-related workplace
United States. It accounts for industrial losses in alcohol testing6 is a regular occurrence. In those
billions of dollars. For example, productivity circumstances, the laboratory can establish a
losses due to alcohol abuse were estimated at two-option system, in which the full legal
$27 billion in 1985, or 39 percent of the total requirements such as chain-of-custody procedures
economic cost of alcohol abuse for that year.4 and specimen seals and secure specimen storage
Because of the recognition of the economic cost are reserved for legal-category tests. The
due to alcoholism and alcohol-related injuries, investigation of a medico-legal case and the
blood alcohol determinations may be required in interpretation of the results of analysis in the
connection with the following: legal context require good judgment and the
assurance that the specimen has been properly
! Employment-related (on-the-job) injury collected and processed. If the following factors
! Workers compensation (federal, state, are appropriately handled, the laboratorian will
local) proceedings have far fewer problems with testimony or legal
! Employee insurance programs proceedings in a court of law.
! Pre-employment screening and evalua-
tions 1.4.1 Consent
! Employee drug screening
! Alcoholism treatment programs. This problem can be dealt with through statutory
provisions such as implied consent laws in each
1.4 Medicolegal Requirements jurisdiction, or through legal advice regarding
individual consent to obtaining a sample.
Most blood-alcohol analyses in hospitals and
other clinical settings are performed solely for 1.4.2 Collection Techniques
medical diagnostic, treatment-related, or other
clinical purposes. In such clinical laboratories Knowledge of the statutes of the jurisdiction
there is no requirement for chain-of-custody concerned and implementing national or regional
procedures (see Section 3). In many jurisdictions regulations are necessary, for these authorities
the results of alcohol analyses may ultimately may designate who may draw the sample, the
become evidence in civil or criminal legal specimen container to be used, and how it is
proceedings, regardless of their original purpose. preserved.
There is, however, no rational basis for the
mystery and trepidation with which alcohol 1.4.3 Identification Procedures
analyses are often regarded in clinical
laboratories.5 Some simple practices can For both medical and subsequent legal purposes,
minimize the extent of involvement of clinical if any, a foundational requirement is to establish
laboratory personnel in subsequent legal from whom the blood specimen was collected, by
proceedings, with respect to the collection and whom, at what date and time, etc. These data
analysis of blood for alcohol for clinical purposes. provide for “traceability” of the specimen—an
A frequently updated, comprehensive, written aspect as important as specimen integrity
(and/or computerized) protocol which is adhered (unaltered state).
to as necessary—minimizing the number of
people involved in collection, transport, analysis 1.4.4 Chain-of-Custody
for alcohol, and storage of a given blood
specimen; as well as a clear statement on all This is the documentation discussed in Section 3
records and reports that the specimen analyzed that accompanies the specimen if chain-of-
was whole blood, serum, or plasma, etc., can custody procedures are used in a given instance.
significantly reduce the involvement of clinical It certifies that: the specimen was obtained from
laboratory personnel in subsequent legal the individual named as the source of the
proceedings. specimen, the specified laboratory was
responsible for the analysis, all individuals who
In some institutions, the analysis (or collection had possession of the specimen prior to analysis
only) of blood specimens for alcohol for are listed, as well as the name of the technician
who performed the analysis.

1.4.5 Security System 2.1 Responsibility for Specimen Collection
This must be utilized, when appropriate, to

maintain the chain of custody and to exclude the Several important considerations are involved in
possibility of tampering. decisions made by medical personnel to collect
blood specimens from traffic accident victims for
1.4.6 Method of Analysis alcohol analysis. From a medical treatment
viewpoint, it may be desirable, or necessary, to
This must be a recognized method having the know the patient's blood alcohol concentration
requisite reliability, and it must be accompanied before administering anesthetics or medications.
by adequate quality assurance procedures. See Such test results may also be utilized in later
Section 4. legal proceedings to determine whether the
person was intoxicated or under the influence of
1.4.7 Interpretations alcohol.
There are many requirements for interpretation of 2.2 Types of Specimens

blood alcohol concentrations and they are both
medical and legal in nature. Many of the Plasma and serum are physiologically and
problems presented in this overview will be ad- pharmacologically more appropriate specimens
dressed in other parts of this document. Certain than whole blood. Intravascular alcohol trans-
aspects, however, such as extrapolation of port involves both the cellular and noncellular
alcohol test results to other times, are beyond its components of blood, but alcohol distribution
scope. occurs primarily between the circulating plasma
and other body tissues and fluids.
1.4.8 Checklist of Issues
Motor Vehicle Codes or other laws may state that
A checklist of issues that routinely surface during breath, blood, or urine may be analyzed for
legal proceedings involving the analysis and alcohol content. Of these fluids, blood, serum, or
reported results of a blood alcohol specimen plasma are usually analyzed in clinical
appears as Table 1. laboratories, since breath-alcohol testing equip-
ment is generally not available in the clinical
2 Specimen Collection laboratory, and since urine alcohol concen-
trations are not well correlated with blood alcohol
Recent revisions of motor vehicle codes in many concentrations.5,7,8
states, in order to combat the problem of driving
under the influence of alcohol (DUI) or while When the term "blood" is used in motor vehicle
intoxicated (DWI) have placed additional statutes, whole blood is the universal meaning.
responsibilities on hospitals and clinical Most regional laws define the alcohol element of
laboratories. These revisions have created a need drinking/driving offenses wholly or partly in terms
for guidance concerning the collection and of blood-alcohol concentrations; and may specify
processing of specimens for blood alcohol whole blood as the required specimen when
analysis. The discussion which follows is “blood” is analyzed. Hence, if blood rather than
intended to assist phlebotomists and laboratory breath is to be analyzed for alcohol, either
personnel involved with the collection and exclusively or primarily in connection with traffic
processing of specimens for alcohol deter- law enforcement, it is best to analyze whole
mination. blood.

Table 1. Checklist of Issues Commonly Arising in Legal Proceedings Involving Blood-Alcohol Analysis
Results
Laboratory and Personnel
! Identity, status, and qualifications of the person giving testimony

! Name, status, and location of laboratory
! Identity, status, and qualifications of the analyst(s) and phlebotomist
! Identity, status and qualifications of the laboratory director
! Licensure and accreditation of laboratory and personnel
Specimens
! Information and documentation on identity of the specimen donor; and on the nature, integrity, and security of
specimens
! Authority for ordering the test and for procurement of the specimens
! Appropriateness and validity of specimen selection
! Details of specimen collection, handling, storage
! Time, date, location of specimen collection; and bodily sampling site
! Chain-of-custody of specimens, and evidence to establish absence of irregularities or tampering; seals, labels, etc.
! Details of specimen containers, anticoagulants, preservatives, etc.
! Present location and condition of specimens or residues
Analysis
! Analysis protocols and standard operating procedures; literature references

! Details of the analysis actually performed; including apparatus, equipment, devices, and procedures involved
! Analytical “raw” data and final results and findings, and their derivation
! Details of instrument or system calibration
! Method characteristics, especially with respect to accuracy, precision, linearity, sensitivity, specificity, interferences
! Validity and reliability of the analysis scheme
Quality Assurance
! Overall quality assurance/quality control schemes

! Control specimens; nominal concentrations, actual results
! Standards, and their origin and validation
! Replicate analyses of the unknown specimens
! Laboratory performance in external alcohol analysis proficiency testing programs and surveys
Interpretation of Results
! Scientific validity of the test results

! Pharmacological, toxicological, or clinical significance and meaning of the results
! Relevance or significance of the results to the legal issues
! Relevant patient history, clinical care and treatment details; effects on results of fluid(s) administration, medications,
shock, trauma or other details of patient status or medical conditions
! Compliance with applicable statute law, case law decisions, and rules and administrative procedures
Documentation
! Records, laboratory requisitions and requests, reports, manufacturers’ literature and other source documents
pertaining to any of the foregoing matters
__________________________________________________________________________________________________________________
Dubowski KM © 1994. Reproduced with permission. All rights reserved.

Alcohol, at equilibrium, is generally distributed be expected to significantly affect the alcohol

throughout the body in proportion to the water concentration of the blood, it is better to select a
content of various fluids, tissues, and organs. venipuncture site remote from the location of
In particular, the alcohol concentration of whole fluid administration in order to ensure a specimen
blood is not identical to that of plasma or of representative of the true alcohol concentration of
serum. However, the alcohol concentration of the specimen. If possible, it is best to collect the
either serum or plasma is, in practice, the same. specimen before any treatment is begun. Ideally,
Both theoretical calculations, based on water venipuncture should be performed in accordance
content, and experimental data yield typical mean with applicable procedures described in the
ratios of 1.12/1 to 1.18/1 in normal subjects for NCCLS document H3, Procedures for the
serum/whole blood alcohol concentrations, with Collection of Diagnostic Blood Specimens by
typical experimental ranges of 1.05/1 to Venipuncture.12
1.25/1.9,10
2.3.3 Disinfectant
The specimen type analyzed should be identified.
Results of alcohol analysis on serum or plasma The disinfectant used for cleansing the
specimens should not be converted to whole venipuncture site should not contain alcohol or
blood concentrations. If courts require the other volatile organic substances. The most
interpretation of serum alcohol concentrations or frequently employed disinfectants for this purpose
the conversion of serum concentrations to whole are aqueous benzalkonium chloride or aqueous
blood concentrations, experts can be retained to povidone-iodine.13 Studies by Dubowski and
perform these functions. It is a complex issue.11 Essary13 have revealed that blood specimens can
be significantly contaminated if alcohol containing
2.3 Specimen Collection, Handling, and sponges are used to cover the venipuncture site
Preservation at the time when the needle is withdrawn from
the vein while attached to the vacuum tube.
The blood collection procedure for forensic Therefore, to avoid the possibility of
alcohol determinations must be conducted so that contamination and legal challenges to the
no doubt exists as to the authenticity and validity acceptability of the specimen collection
of the specimen. In this regard, several points procedure, only nonalcoholic disinfectants should
should be emphasized. be employed, and sterile dry sponges should be
used to cover the venipuncture site. Further, if
2.3.1 Time of Collection evacuated collection tubes are used, the tube
should be removed from the multisample
The time of collection is critical information collection needle and holder before withdrawing
which must be recorded and should appear on the the needle from the puncture site.
report of results.
2.3.4 Specimen Container
2.3.2 Site of Venipuncture
The specimen container is important and will vary
The site of the venipuncture is usually the median depending on whether serum, plasma, or whole
cubital or one of the other superficial veins of the blood is to be analyzed. If serum is required, the
forearm. Veins in the lower extremities can also blood should be collected in a container without a
be used if the forearms are not accessible preservative or anticoagulant and allowed to clot.
because of injuries or for other reasons. During The serum can be sent directly to the laboratory
the early phases of alcohol absorption, peripheral without further processing if the specimen is to
venous blood concentrations lag behind arterial be analyzed for alcohol content within four hours.
blood concentrations, particularly in the lower If the analysis will be delayed, the serum should
extremities. be transferred to another container and treated
with sufficient sodium fluoride to produce a
Blood should not be removed from veins into minimum concentration of 10 mg/mL
which intravenous fluids or other medications are (0.24mmol/ml).
being administered at the time. The dilution effect
can lower the alcohol concentration. Even when For whole blood or plasma specimens, the type
the presence of such parenteral fluids would not and amount of anticoagulant present is not
important if the specimen is analyzed within four

hours of collection. It is only necessary that the 2.3.5 Size of Sample

anticoagulant not interfere with the alcohol
determination and that a sufficient quantity is The size of the sample should be sufficient to
present in the specimen to prevent clotting. If permit retesting, if necessary.
the analysis is to be delayed, additional
safeguards must be instituted to prevent changes 2.4 Specimen Handling
in the alcohol content of the blood. For this
purpose, potassium oxalate monohydrate (5 To ensure complete dissolution of the fluoride in
mg/mL of blood; 2.7Fmol/mL) and sodium the blood, the closed container of blood should be
fluoride (1.5 mg/mL of blood; 3.6Fmol/ml) are an gently inverted several times immediately
appropriate anticoagulant and preservative following specimen collection.
combination for storage at 5 oC of initially sterile
blood specimens for up to 48 hours.14 Blood The laboratory request form for alcohol analysis
alcohol specimens stored at -20 EC or below are should be completed legibly and should contain
stable indefinitely. the following information:
Specimens that are to be transported or mailed in ! Patient's full name

an unrefrigerated condition, or stored for more ! Identification number
than 48 hours should be preserved with higher ! Time and date of specimen collection
concentrations of sodium fluoride (10 mg/mL of ! Site of venipuncture
blood; 0.24mmol/mL).8 However, it has been ! Phlebotomist's name
documented that changes produced by contamin- ! Name and address of facility where
ating microorganisms can affect alcohol specimens were collected.
concentrations in blood specimens even in the
presence of preservatives. Blume and Lakatua15 Collection kits designed to facilitate the sampling
reported that various organisms isolated from process are available from commercial sources.
contaminated blood specimens were capable of Potential purchasers should determine that the
producing ethanol when inoculated into bank kits meet their needs and comply with local laws
blood. Candida albicans was particularly active in concerning blood alcohol analysis.
this regard, producing significant quantities of
alcohol even in the presence of sodium fluoride. Additional special specimen handling consid-
These investigators recommended that fluoride erations are addressed in Appendix C.
(10 mg/mL; 0.24mmol/ml) be used as a
preservative and that care should be taken to 2.5 Replicate Blood Specimens
assure that microbial organisms are not
introduced into the specimens. When it is known at the outset that alcohol
analysis results will be required for legal purposes
Winek and Paul16 reported that alcohol analyses as well as for immediate clinical patient care, it
of blood obtained under sterile conditions from may be practical and appropriate to collect
living humans can be delayed as long as 14 days replicate blood specimens in parallel and with
without a significant change in alcohol content. consideration for the required kind of specimen,
They state that this holds true whether the blood e.g., unpreserved serum for immediate analysis
sample is refrigerated or not, or whether a for clinical purposes, and a preserved
preservative is added to the sample. anticoagulated whole-blood specimen for separate
Nevertheless, the question can still arise as to medicolegal analysis. Different documentation
how the phlebotomist could know with certainty, and handling may be required in such instances.
even if aseptic collection techniques were
employed, that no micro-organisms entered the 3 Chain-of-Custody
specimen and produced changes in the alcohol
concentration. For this reason, it is advisable to
Procedures17,18
employ preservatives and to refrigerate
specimens as additional safeguards against 3.1 Purpose
changes in alcohol content.
The established relationships between alcohol,
trauma, and litigation ensure that the results of
many medically-indicated blood-alcohol analyses

performed in hospital and other clinical dedicated record. The concept of the chain-of-
laboratories will later be sought as evidence in custody and the practices associated with it
civil or criminal litigation or other adversarial originated long ago in the collection and
proceedings, such as formal arbitration. preservation of physical evidence and its
(Introduction of such evidence demands a subsequent admission in litigation. These
requisite degree of proof; an important element of practices have become wide-spread and familiar
that proof is the chain-of-custody.) In addition, to clinical laboratories especially in connection
alcohol analysis is frequently sought exclusively with the recent large scale drug-use testing of
for legal reasons, such as in drinking-driving biological specimens.
investigations. The goal is to provide adequate
and acceptable proof of the identity of the 3.2 Documentation
specimen and specimen donor, to assure the
integrity of the specimen throughout its Laboratories which routinely undertake blood
existence, and to eliminate significant changes in alcohol testing should anticipate being required to
the specimen composition before analysis. provide chain-of-custody information and
documentation, sometimes months or years after
The laboratory's responsibility for initiating and the testing date. The obvious choices are to
maintaining the chain-of-custody begins when the establish, maintain, and document the chain-of-
specimen is collected (if by that laboratory's custody for all blood-alcohol specimens/ tests, or
personnel) or when the specimen first reaches the to do so only in selected predetermined instances
laboratory. The laboratory's responsibility ends of the kind most likely to be involved in later
when the specimen is entirely consumed in its litigation, such as accidents, assaults, trauma,
analysis, or when custody of the specimen is workplace-related events, etc. Clearly, blood
transferred to any party outside of the laboratory, alcohol testing originally conducted under legal
or if the specimen is destroyed on proper mandate, e.g., police-directed or court-ordered
authority. In the first event (specimen tests of arrested drivers, should always involve
consumption), the laboratory's chain-of-custody the full chain-of-custody scheme.
responsibilities continue with respect to the
original specimen container. Several principles apply to chain-of-custody
procedures and their documentation with special
Chain-of-custody, in the context of this purpose forms. Both the procedures and their
document, is a term-of-art pertaining to the documentation should be simple, limited to
procedures and documentation necessary for legal essentials, easy to use, and readily traceable,
purposes to establish the identity and origin of with minimal reliance on testimony of the people
the blood specimen; to enable it to be traced to involved with a given blood specimen. Access to
the person from whom it was collected; to specimens, at any stage, should be restricted as
reasonably assure absence of specimen mix-up; much as possible. Adequate, unique labeling
deliberate adulteration; or other tampering; to should establish the identity of the specimen.
exclude unauthorized access at any stage of Physical security measures, such as proper seals,
specimen collection, storage, analysis and tamper-evident containers, and secure (locked)
retention; as well as to prevent loss of the storage should be employed for all affected
specimen. Such proof is required in part because specimens.
blood is fungible—meaning that blood specimens
typically cannot be visually distinguished from The chain-of-custody record should contain
each other, and because alteration or substitution necessary information on the following elements
could readily occur in the absence of proper in logical sequential form:
safeguards. The basic concept of the chain-of-
custody is to trace a specimen, step-by-step, and (1) Data on the origin of the specimen, as
person-by-person, from its origin to the listed in Section 2.4.
completion of the analysis (and beyond if further
retained). The chain-of-custody must document (2) Date, time, location, when relevant,
the location and care of the specimen at all times and full identity of all those involved in
by identifying sequentially every person having every transfer of custody or physical
responsibility for and control of the specimen for possession of the specimen, together
any purpose. Every change in custody must be
documented by appropriate entries into a

with the purpose of, or reason for, location is up to the laboratory. In most
each transfer. situations, keeping the specimen and a dedicated
chain-of-custody document together at all times
(3) Details of any irregularity, such as is preferable.
container breakage, spillage, or
other loss which can affect the 4 Methods of Analysis
specimen.
Of the multitude of methods available for the
(4) Any pertinent remarks bearing on determination of ethyl alcohol (or ethanol) in
the original identity, integrity, blood specimens, the two summarized in Table 2
composition, condition, location, or are appropriate for use in clinical laboratories and
custody of the specimen, (or any are listed in the order of preference.14 A
change not reflected in other comprehensive review of major developments in
entries). alcohol analysis has been published.19
The entries should be brief and to the point. For 4.1 Gas Chromatography
example, a temporary transfer of custody may
occur when the analyst is absent during a break Gas chromatographic (GC) determination of ethyl
and the unsealed specimen remains on a alcohol (or ethanol) analysis can be performed
laboratory workbench. The required entries in the with simple and relatively inexpensive
chain-of-custody document include the date(s) instruments. It yields information on both the
and time(s) and the people involved (from whom- identity and concentration of ethanol and other
to whom-to whom) in the initial transfer of volatile constituents of the sample. Currently, GC
custody from the analyst to a qualified coworker. is the method of choice. Its desirable features
The transfer returning custody to the analyst and include rapidity, sensitivity, minimal sample
a statement of reason for each change, such as preparation and manipulation, and specificity for
"transfer due to absence during a break" and ethanol. Essentially complete specificity for
"resumption of analysis" must be documented. ethanol can be achieved with gas
chromatography. This is accomplished by using
Clearly, it is desirable to minimize the number of multiple columns, by varying analysis conditions,
custody transfers, especially temporary ones, or by performingGC analysis after treating one of
and, hence, the number of chain-of-custody two identical sample aliquots with approximately
entries. The need for temporary transfers can be 250 units of alcohol dehydrogenase (EC* 1.1.1.1)
largely obviated by arranging for storage of per mL of blood or serum. This procedure is
specimens during brief absences in a locked followed by gas chromatography; thereby
refrigerator compartment, lockbox, locker, abolishing any ethanol GC “peak” in the treated
container, or secure location accessible only to sample in comparison with the untreated sample,
the custodian. and greatly enhancing the certainty of
identification of ethanol. Analysis of “headspace”
Chain-of-custody documents can be designed for vapor above a blood or serum sample saturated
individual or multiple specimens. In most clinical with sodium chloride, ammonium sulfate, or
laboratories, forms designed for use with a single certain other salts and then equilibrated at 50 EC
specimen are most useful. The essential or other controlled temperature is a simple and
information, in addition to the identity and origin desirable gas chromatographic technique.20
data under item 1 above, can be recorded in Headspace analysis is usually preferable to direct
separate rows for each change of custody or injection of diluted whole blood or serum
other transaction, in four columns: the date and specimens into the chromatograph inlet because
time; typed/printed/stamped name of the person it eliminates problems of inlet and column
releasing the specimen; the identical information contamination and syringe plugging. Direct
for the person receiving the specimen; and the injection of a diluted liquid sample may be
reason for the change. Legibility of all entries,
including the signature, is indispensable for such
a record. All other handwritten entries should be *
EC = Enzyme Commission. In the listing of an
legibly printed in ink. Whether or not the chain-
enzyme, the abbreviation is followed by four numbers indicating
of-custody document physically accompanies the the classification of the enzyme according to main division,
specimen at all times or is kept in a secure subclass, sub-subclass, and serial number of the sub-subclass.

Table 2. Summary of Methods for Blood-Alcohol Analysis
Required
Sample
Treatment/ Specificity for Apparatus Final
Method Separation Ethanol Requirements Measurement Limitations
Gas Dilution or Selective for Gas Electrical Requires

chromatography headspace ethanol; multiple chromatograph, voltage calibration at
equilibrium columns/ water bath (for or current, via time of
conditions headspace), strip chart analysis
greatly strip-chart recorder or
increase recorder or integrator
selectivity integrator
Enzymatic Dilution (for Selective for Ultraviolet Photometric/ Potential

oxidation with plasma or ethanol; some spectro- spectrophoto- interference
alcohol serum) interference by photometer, metric reading by
dehydrogenase or isopropanol and visible higher
deproteinization methanol in high photometer, or alcohols,
concentrations some automatic some enzyme
analyzers inhibitors,
etc.
Modified with permission from Dubowski KM. Alcohol determination in the clinical laboratory. Am J Clin Path. 1980:74:747–750
preferable for emergency tests because it It is also practical for occasional or infrequent
eliminates the 15–30 minute equilibration period use, because elaborate preparation is not
required for headspace analysis. When many necessary, and reliable single-assay reagents are
blood specimens are to be analyzed, automated commercially available. For the same reasons, it
gas chromatographic headspace analysis21,22 or serves well as a backup method for gas
use of automatic liquid sampling (injection) chromatography. The principal variants are use
attachments is indicated. The principal variants of of alcohol oxidase or alcohol dehydrogenase,
gas chromatographic methods for blood-alcohol measurement of the change in ultraviolet
analysis are listed in Table 3. Use of internal absorbance of the reaction mixture at 260 or 340
standards has become the accepted norm.20 nm, or visual photometry of a secondary indicator
reaction. A variety of commercial reagents and
4.2 Enzymatic Oxidation with kits are available for ADH methods. The
Alcohol Dehydrogenase characteristics and performance of three ADH
methods have been reported.23,24
Enzymatic oxidation with alcohol dehydrogenase
(ADH) or alcohol oxidase (AO) is a sensitive and The principal features of ADH methods are given
simple method for alcohol measurement. A in Table 4.
variety of both automated and manual methods
based on this principle is available for routine use.

Table 3. Principal Variants of Gas Chromatographic Methods for Blood-Alcohol Analysis

Detector
Flame-ionization detector
Appropriate packed columns
Solid phase (Porapak and Chromosorb, are examples of commonly used solid phase
columns)*
Liquid phase (Carbowax and Hallcomid are examples of commonly used liquid phase
columns)*
*Other suitable columns may be substituted.
Appropriate capillary columns
Large bore
Liquid-coated walls
Analysis techniques
Headspace sampling after equilibration
Direct injection of diluted blood or serum
Protein precipitation and direct injection of the supernatant liquid
Internal standard added to sample
Quantitation
Electronic integration/printout of peak areas
Dedicated electronic controllers/data systems
Measurement of strip-chart recording of detector response: Peak heights or peak areas
Modified with permission from Dubowski KM. Alcohol determination in the clinical laboratory. Am J Clin Path. 1980;74:747–750
Table 4. Principal Features of Enzymatic (ADH) Oxidation Methods for Blood-Alcohol Analysis
Reactions
Catalyst
NAD oxidoreductase (alcohol dehydrogenase: EC 1.1.1.1)
Preparation of plasma, serum, or blood
Dilution with saline solution or deproteinization with perchloric acid or trichloracetic acid or Ba(OH)2
+ ZnSO4
Final measurement
Ultraviolet spectrophotometry
ª Absorbance at 260 nm (NAD+ absorbs)
ª Absorbance at 340 nm (NADH absorbs)
Visible photometry: measurement of stable red formazan color at 500 nm
Amperometric measurement with O2 electrode
Fluorometry
Modified with permission from Dubowski KM. Alcohol determination in the clinical laboratory. Am J Clin Path. 1980;74:747–750.

There is considerable variation in the types of (1) NCCLS Document C24: Internal
instruments employed in these procedures. Some Quality Control Testing: Principles and
procedures use rapid centrifugal analyzers, while Definitions.
others employ rapid electrochemical measurement
using immobilized ADH and AO enzymes and (2) NCCLS Document EP10: Preliminary
oxygen-sensing electrodes. Other methods have Evaluation of Quantitative Clinical
been adapted to automated analyzers, or use Laboratory Methods.
discrete sample analysis employing radiative
energy attenuation with ADH, based on the 5.1 Calibrators (Standards)
principle of fluorescence quenching. ADH-based
enzymatic oxidation methods are subject to The calibrators should be selected to represent
potential interference by elevated concentrations critical concentrations, which span the clinically
of isopropanol and higher alcohols to varying and forensically relevant alcohol concentrations
extents, while AO-based methods cannot and include the upper limit of linearity of the
distinguish between ethanol and methanol. At analysis. These calibrators will bracket the
commonly encountered concentrations of majority of positive results and can be used to
isopropanol and methanol, ADH-based methods demonstrate linearity, in conjunction with a
are not significantly affected. specimen established to be free of alcohol.
Details of a well-documented enzymatic oxidation Calibrators can be prepared from a freshly opened
procedure are given in Gadsden and Taylor in container of pure ethanol, from a tightly sealed
“Ethanol in “Biological Fluids by Enzymic container of stock solution prepared by diluting an
Analysis.” (Selected Methods of Emergency aliquot from a freshly opened container of pure
Toxicology. Washington, DC, AACC Press, ethanol, or from analytical grade 95% ethanol.
1986;63–65.) Calibrators at various concentrations are available
from commercial sources. Aqueous Standard
5 Quality Assurance Reference Materials containing ethanol are
available from the National Institute of Standards
The general recommendations for quality and Technology (SRM 1828a). Aqueous standard
assurance which follow exemplify good labora- solutions are also available from the College of
tory practice standards as applied to blood American Pathologists, and other sources.
alcohol testing. Because clinical laboratory pro-
cedures are continually evolving technologically, Calibrators contain a designated mass (weight) of
it is important to integrate into the laboratory’s ethanol per volume of solution. Technicians
good laboratory practices, as a minimum, preparing standards should take into account that
directions for test performance, establishment the density of ethanol is less than 1 and varies
and validation of calibrations, checks on linearity, with temperature.
and other analysis instructions provided by the
applicable manufacturer(s) of the instrument(s) Gas Chromatography: Every alcohol analysis or
and commercial reagents utilized. Further, the batch of analyses performed by GC methods
ongoing mandates on use of controls and other should begin with the analysis of at least one,
good laboratory practices promulgated by and preferably two or more different calibrators
regulatory and other applicable authority should together with an alcohol-free “blank,” because
be recognized and complied with when the operating parameters and calibration of GC
applicable. instruments vary with each startup and can also
drift during prolonged operation.
In the discussion on calibrators and controls
which follows, separate consideration is given to Enzymatic Oxidation: The instrument manu-
certain aspects applicable to GC or ADH facturer’s recommendations for calibration should
methods, respectively, to the extent that they be followed. Most current generation analyzers
differ. have been shown to have stable calibrations over
time, often for months. Well-characterized, stable
The laboratory should also follow, to the extent controls should be included in each analysis or
feasible, the principles and practices outlined in run to verify the validity of such historical
these NCCLS documents: calibration. For integrated analysis systems, it is

important to use the calibrators provided by the

instrument manufacturer, and to avoid using 6 Reporting and Significance of
unvalidated calibrators from other sources which Results
may introduce analysis bias as the result of
matrix effects.
6.1 Blood Alcohol Records
5.2 Controls Information concerning the specimen for alcohol
analysis can be recorded in a bound ledger-type
Every analysis or batch of analyses should be
record book with numbered pages which is
accompanied by the analysis of negative and
maintained in the laboratory. Alternatively,
positive controls. The controls should consist of
recordkeeping can be computerized and often is
known, appropriate concentrations of ethanol in
in the modern clinical laboratory. The same
the same biological matrix as the specimens to be
principles apply when computer laboratory
analyzed. Each control should be processed
information systems are used. Specimen records
through all steps of the procedure, exactly as
should include:
each specimen is processed. Concentrations of
multiconcentration controls should be targeted at
! Specimen number
or near decision points such as “cut-offs”
! Patient's name
separating positive from negative results and near
! Date and time of specimen receipt
the limit of linearity.
! Condition of seals on container and
specimen tubes
Controls can be prepared by adding known
! Condition of specimen
quantities of ethanol to homogeneous preserved
! Other pertinent information
biological specimens, mixing thoroughly, and
! Date and time of analysis
freezing aliquots in tightly sealed containers.
! Name of analyst
Prepared controls for serum and whole blood
! Blood (or serum if applicable) alcohol
ethanol (alone or with other alcohols) are
concentration
commercially available.
To shorten the chain-of-custody, if used, it is best
Target values and limits for laboratory-prepared
if one person processes the specimen from
controls should be established as described in
receipt in the laboratory through conducting the
NCCLS publication C24, Internal Quality Control
alcohol analysis and reporting results.
Testing: Principles and Definitions.
A file should also be established in which all
If only one specimen is to be analyzed, the
information relating to the specimen and its
standard(s) (if needed) should precede and the
analysis is maintained. These files may be
control(s) should follow or bracket the specimen.
organized either alphabetically using the patient's
last name, or numerically, using the identification
Each day's quality assurance results should be
number on the report form. If a numerical system
recorded and compared to previous results to
is selected, an alphabetical cross reference index
facilitate early detection of changes. The principal
should be established. In either event, it should
purpose of including controls in an analytical run
be possible to access the necessary information
is to determine whether the analytical method at
chronologically, by the patient's name, and by
the time is yielding acceptable results. Out-of-
the specimen identification number. Any
control results require immediate action to
information which cannot be conveniently placed
investigate and correct the analysis performance.
directly into this file, should be photocopied, if
Pending such action, blood-alcohol testing should
possible, and included with the other records.
be suspended.
This is frequently necessary when information is
recorded directly onto the shipping container.
Gas Chromatography: Because of the variability
Items which cannot be copied should be recorded
of instrument parameters and calibration with
in the section provided for other information in
each startup, and the tendency of these factors
the bound record book. The source of the items
to drift during prolonged instrument operation, at
which are copied or transcribed should be clearly
least every tenth specimen should be a control
specified so that upon subsequent examination,
when multiple, sequential analyses are
the material can be easily identified.
conducted. Multilevel controls are preferred.

All records of forensic blood alcohol analyses determinations that were performed two to five
should be stored in a secure location, with access years previously and occasionally, even earlier.
limited to authorized personnel, for a defined For this reason, it is important to preserve such
period of time (e.g., ten years). Since civil suits records if the laboratory changes ownership or
resulting from traffic accidents often take place discontinues operations.
long after the alcohol analysis, it is not
uncommon to have to testify about
6.2 Conversions
The following conversions are frequently used in blood alcohol testing.

g/L g/dL % w/v mg/dL mmol/L
1.00 0.10 0.10 100 21.71
Conversion factors: mg/dL X 0.2171 = mmol/L

mmol/L X 4.61 = mg/dL
7 Terminology and Statutory

Provisions 7.1.3 Intoxication
The American Medical Association's Manual25

7.1 Terminology
lists eleven signs and symptoms which are
usually accepted as supporting evidence of
7.1.1 Alcohol
alcoholic intoxication:
Alcohol refers to ethanol which may carry
! odor of the breath
synonyms of ethyl alcohol, EtOH. If another
! flushing of skin
alcohol is present, it must be specifically
! loss of muscular coordination
identified and uniquely analyzed.
! speech difficulties
! disorderly or unusual conduct
7.1.2 Blood Alcohol Concentration (BAC)
! mental disturbance
! visual disorders
The concentration of ethyl alcohol in whole blood
! sleepiness
is expressed most commonly as percent by
! muscular tremors
weight/volume (% w/v) or as grams per deciliter
! dizziness
(g/dL).
! nausea.
Many regional laws prescribe blood alcohol
Many of these conditions can result from
concentrations constituting impairment or
nonalcohol related pathological conditions. Only
intoxication in terms of percent alcohol "by
careful differential diagnosis, including a
weight." This means grams of alcohol in 100 mL
determination of alcohol, can distinguish alcoholic
of blood. The clinical laboratory most commonly
intoxication from disease or illness, or show their
determines this by using a measured volume of
simultaneous presence.
blood for assay and reporting the alcohol
concentration as milligrams of alcohol per 100
mL or dL of blood. The laboratory value may be 7.2 Statutory Provisions
converted to BAC expressed as percent w/v by
moving the decimal point three places to the left. States and provinces have varying definitions of
Example: 125 mg/dL converts to 0.125% w/v. intoxication, impairment, alcoholic influence, and
However, only two figures are used in reporting the blood alcohol concentration at or above which
percent. The value is truncated to produce the it is illegal to operate a motor vehicle. For
two digit results, i.e., the third digit is omitted. example, in the United States the laws in general
Example: 139 mg/dL is reported as 0.13% w/v or follow the alcohol-related provisions of the
0.13 g/dL. Uniform Vehicle Code which have been endorsed

by the National Safety Council's Committee on The result of a chemical determination of the
Alcohol and Other Drugs and by the House of concentration of alcohol in a person's blood
Delegates of the American Medical Association. should not be confused with the effect of that
alcohol upon the person’s brain. The former is
The following provisions of the UVC26 are widely measurable with a high degree of accuracy. The
followed: latter is not. At each extreme of the BAC (very
low or none and very high, 0.40 % w/v or more)
1. Section11-903(a)5: Alcohol concentration there is little doubt about whether the alcohol has
shall mean either grams of alcohol per 100 had an effect. To set a single concentration, i.e.,
milliliters of blood or grams of alcohol per 210 0.08 % w/v, as a cut-off point above which each
liters of breath (FORMERLY § 11-902(b)4; and every person is supposedly affected by
REVISED, 1979.) alcohol to an equal degree of impairment for all
purposes is inappropriate. Some individuals are
2. Section 11-903(b)1: If there was at that impaired at less than 0.08 % w/v; others are not
time an alcohol concentration less than 0.08, obviously impaired at much higher
such fact shall not give rise to any presumption concentrations.
that the person was or was not under the
influence of alcohol, but such fact may be "Under the influence" is a phrase with very wide
considered with other competent evidence in implications because its significance is not
determining whether the person was under the universally agreed upon. One drink may very
influence of alcohol. (REVISED, 1979, 1984; well produce demonstrable and measurable
REVISED AND RENUMBERED, 1992.) impairment of judgment and response time. On
the other hand, to some people, under the
3. Section 11-903(b)2: If there was at that influence means drunk and disorderly and nothing
time an alcohol concentration of 0.08 or more, it less. Therefore, it is advisable to use other,
shall be presumed that the person was under the specific terms to describe impairment for a given
influence of alcohol. (REVISED, 1979, 1984; task.
RENUMBERED, 1992.)
Finally, a source of difficulty in equating a blood-
It should be noted that these provisions apply to alcohol concentration with its effect upon the
motor vehicle operation. There are additional brain is that the identical blood alcohol
factors that must be taken into account should concentration(s) can occur in a single individual at
interpretations be required for other purposes. two or more different times in a drinking episode.
There is a wide variety of sources for author- During the absorption of alcohol the patient will
itative information regarding the pharmacology pass through various blood alcohol concentrations
and physiological effects27 of alcohol on the and again later during elimination of the alcohol
human body. Any interpretation of the alcohol will experience them all. To say the patient was
concentration is best left to those qualified by equally affected at the two times is unrealistic
experience and training. because of the acute tolerance phenomenon
(Mellanby effect) and other factors.28 In many
instances, there is greater impairment at a given
blood alcohol concentration during the rapid
absorption of alcohol than during the elimination
phase.

References 11. Rainey PM. Relation between serum

and whole-blood ethanol concentrations. Clin
1. Froede RC. The pathologist's role in the Chem. 39:2288–2292, 1993.
recognition and management of alcoholism. Am
J Clin Pathol. 74: 718, 1980. 12. National Committee for Clinical Lab-
oratory Standards. Procedures for the Collection
2. Froede RC, Gordon JD. Alcoholism -the of Diagnostic Blood Specimens by
second great imitator. An introduction to the Venipuncture–Second Edition; Approved
problem of alcoholism. Am J Clin Pathol. 74: Standard. NCCLS document H3-A2. Wayne, Pa.:
719–720, 1980. NCCLS;1984.
3. Poklis A. Analytical/Forensic Toxicology. In: 13. Dubowski KM, Essary NA. Contam-
Casarett and Dorell’s Toxicology. The Basic ination of blood specimens for alcohol analysis
Science of Poisons, 5th ed., ed by C.D. Klaussen. during collection. Abstracts & Reviews in
McGraw Hill, New York, 1995, p. 963. Alcohol & Driving 4 (No. 2): 3–7, 1983.
4. Economic issues in alcohol use and abuse. 14. Dubowski KM. Alcohol determination
In: Eighth Special Report to the U.S. Congress on in the clinical laboratory. Am J Clin Pathol. 74:
Alcohol and Health from the Secretary of Health 747–750, 1980.
and Human Services, September 1993. NIH
Publications No. 94-3699, Rockville, MD: DHHS, 15. Blume P, Lakatua DJ. The effect of
National Institute on Alcohol Abuse and microbial contamination of the blood sample on
Alcoholism, 1994, p. 256. the determination of ethanol levels in serum. Am
J Clin Pathol. 60: 700–702, 1973.
5. Dubowski KM. Alcohol analysis: Clinical
laboratory aspects, Part 1. Laboratory 16. Winek CL, Paul LJ. Effect of short-
Management. 20 (No. 3):43–54, 1982. term storage conditions on alcohol concentrations
in blood from living human subjects. Clin Chem.
6. Dubowski KM, Caplan YH. Alcohol testing 29: 1959–1960, 1983.
in the workplace. In: Medicolegal Aspects of
Alcohol, 3rd ed. Edited by Garriott JC. Lawyers 17. Giannelli PC, Imwinkelried EJ. Scien-
and Judges Publishing Co., Tucson, Arizona, tific Evidence. 2nd ed. Charlottesville, VA: The
1996, pp. 439–475. Mitchie Co. 1993; (1):198–214.
7. Dubowski KM. Manual for Analysis of 18. Moenssens AA, Starrs JE, Henderson
Ethanol in Biological Liquids. Report No. DOT- CE, and Inbau. Scientific Evidence in Civil and
TSC-NHTSA-76-4 (HS 802 208). U.S. Criminal Cases. 4th ed.,Westbury, NY: The
Department of Transportation, National Highway Foundation Press, Inc., p. 79-80, 1995.
Traffic Safety Administration, p. 94, 1977.
19. Jones AW. Measuring alcohol in blood
8. Kaye S. The collection and handling of and breath for forensic purposes - a historical
blood alcohol specimen. Am J Clin Pathol. 74: review. Forensic Science Review 8(No. 1): 13-
743–746, 1980. 43, 1996.
9. Brettel HF. Blutalkohol und Wassergehalt. 20. Dubowski KM. Ethanol - Type C pro-
Lubeck, Germany,Verlag Max Schmidt- cedure. In Sunshine, ed. Methodology for
Romhild,1972,pp. 28–57. Analytical Toxicology. Cleveland: CRC Press,
Inc. 1975;149–154.
10. Winek CL, Carfagna M. Comparison of
plasma, serum, and whole blood ethanol 21. Gadsden RH, Terry CS, Thompson BC.
concentrations. J Analyt Toxic. 11:267–268, Alcohols in biological fluids by gas
1987. chromatography (automated head-space method).
In: C.S. Frings and W.R. Faulkner, eds. Selected
Methods of Emergency Toxicology. Washington:
AACC Press. 1986;40–43.

22. Shaw RF. Methods for fluid analysis. In 25. Alcohol and the Impaired Driver.
J.C. Garriott. ed. Medicolegal Aspects of Alcohol, Chicago, American Medical Association, pp.
3rd ed. Tucson, AZ: Lawyers and Judges 143–144, 1970 (Reprinted 1976 by National
Publishing Co. 1996; 219–237. Safety Council).
23. Nine JS, etal. Serum-ethanol determination: 26. Uniform Vehicle Code and Model
Comparison of lactate and lactate dehydrogenase Traffic Ordinance, Revised 1987, Evanston, IL,
interference in three enzymatic assays. J. Analyt. National Committee on Uniform Traffic Laws and
Toxicol. 19: 192-196 (1995). Ordinances, pp. 65–66, 1987.
24. Winek CL, Wahba WW. A response to 27. Hardman JG, Limbird LE, Molinoff PB,
“Serum-ethanol determination. Comparison of Ruddon RW. Goodman and Gilman's The
lactate and lactate dehydrogenase interference in Pharmacological Basis of Therapeutics. 9th ed.
three enzymatic assays.” J. Analyt. Toxicol. New York: McGraw-Hill. 1996:386–392
20:211-212(1996).
28. Mitchell A. Alcohol-induced
impairment of central nervous system function:
behavioral skills involved in driving. J Studies on
Alcohol. 1985; (suppl 10): 106–116, 1985.

Appendix A. Stages of Acute Alcoholic Influence/Intoxication (Whole Blood Alcohol)
BLOOD-ALCOHOL STAGE OF
CONCENTRATION ALCOHOLIC CLINICAL SIGNS/SYMPTOMS
grams/100 mL. INFLUENCE
Influence/effects not apparent or obvious
Behavior nearly normal by ordinary observation
0.01-0.05 Subclinical
Impairment detectable by special tests
Mild euphoria, sociability, talkativeness

Increased self-confidence; decreased inhibitions
Diminution of attention, judgment and control
0.03-0.12 Euphoria Some sensory-motor impairment
Slowed information processing
Loss of efficiency in critical performance tests
Emotional instability; loss of critical judgment

Impairment of perception, memory and comprehension
Decreased sensitory response; increased reaction time
Reduced visual acuity, peripheral vision and glare recovery
0.09-0.25 Excitement Sensory-motor incoordination; impaired balance
Drowsiness
Disorientation, mental confusion; dizziness

Exaggerated emotional states (fear, rage, grief, etc.)
Disturbances of vision (diplopia, etc.) and of perception of
color, form, motion, dimensions
Confusion Increased pain threshold
0.18-0.30
Increased muscular incoordination; staggering gait; slurred
speech
Apathy, lethargy
General inertia; approaching loss of motor functions

Markedly decreased response to stimuli
0.25-0.40 Stupor Marked muscular incoordination; inability to stand or walk
Vomiting; incontinence of urine and feces
Impaired consciousness; sleep or stupor
Complete unconsciousness; coma; anesthesia

Depressed or abolished reflexes
0.35-0.50 Coma Subnormal temperature
Impairment of circulation and respiration
Possible death
0.45+ Death Death from respiratory arrest
Dubowski KM © 1997. Reproduced with permission. All rights reserved.

Appendix B. Guide to Serum-Alcohol Test Results
SERUM-ALCOHOL
CONCENTRATION STAGE OF
grams/100 mL. ALCOHOLIC CLINICAL SIGNS/SYMPTOMS
INFLUENCE
0.01–0.06 Subclinical No apparent influence

Behavior nearly normal by ordinary observation
Slight changes detectable by special tests
0.03–0.14 Euphoria Mild euphoria, sociability, talkativeness

Increased self-confidence; decreased inhibitions
Diminution of attention, judgment and control
Beginning sensory-motor impairment
Slowed information processing
Loss of efficiency in finer performance tests
0.11–0.29 Excitement Emotional instability; loss of critical judgment

Impairment of perception, memory and comprehension
Decreased sensitory response; increased reaction time
Reduced visual acuity, peripheral vision and glare
recovery
Sensory-motor incoordination; impaired balance
Drowsiness
0.21–0.35 Confusion Disorientation, mental confusion; dizziness

Exaggerated emotional states (fear, rage, sorrow, etc.)
Disturbances of vision(diplopia, etc.) and of perception
of
color, form, motion, dimensions
Increased pain threshold
Increased muscular incoordination; staggering gait;
slurred speech
Apathy, lethargy
0.29–0.46 Stupor General inertia; approaching loss of motor functions

Markedly decreased response to stimuli
Marked muscular incoordination; inability to stand or
walk
Vomiting; incontinence of urine and feces
Impaired consciousness; sleep or stupor
0.40–0.58 Coma Complete unconsciousness; coma; anesthesia

Depressed or abolished reflexes
Subnormal temperature
Incontinence of urine and feces
Impairment of circulation and respiration
Possible death
0.50+ Death Death from respiratory arrest
Dubowsk KM © 1991. Reproduced with permission. All rights reserved.

Appendix C. Special Specimen Handling Considerations
Each container of blood should be sealed immediately after the specimen has been drawn and mixed with
the preservative. To accomplish this, the phlebotomist should initial the seal, record the date and time on
it, and affix it to the stopper and side of the specimen tube in such a manner that the stopper cannot be
removed or punctured without disrupting the seal. Paper labels approximately 2 x 2 inches with
nonpeelable adhesive on one side are suitable for sealing containers. The specimen container should also
be clearly labeled. The label should specify the patient's name and any identification or accession numbers
that are needed to relate the specimen to the patient or to the laboratory request form.
If the specimen is to be analyzed in the same facility where it was collected, it should be placed together
with the request form and patient consent form in a sealed container and delivered directly to the laboratory
where it should be stored in a locked refrigerator until it can be further processed. When the analysis is
performed, the person conducting the determination should break the seal on the container and open it to
gain access to the forms and specimen.
Specimens to be sent to another facility for analysis should be sealed as described above and either
delivered by a messenger or forwarded by First Class Mail (see Related NCCLS Publication H5-A3).
Sending specimens by certified or registered mail is recommended to reduce problems with the chain of
custody. In a 1951 Nebraska case, the court held that there is a presumption that articles transported by
regular U.S. Mail and delivered in the ordinary course of the mails are received in substantially the same
condition in which they are sent.1 The same presumption of regularity and hence lack of a requirement that
air shipment couriers or transport agency employees make entries in a chain-of-custody document for a
urine sample in transit for workplace drug testing was affiirmed in a 1994 decision of the US Court of
Appeals for the Eleventh Circuit.2 Other court decisions exist to the same effect. If the sample was hand
carried and several different people handled it along the way, then technically each person could be
subpoenaed to establish a complete chain-of-custody. If the specimen is adequately sealed and forwarded
by mail, generally only the person who collected, sealed and mailed the specimen, and the person who
broke the seals and analyzed the specimen need testify.
References to Appendix C
1. Schacht vs. State, 50 N.W. 2d 78, 1951.
2. Interstate Brands Corp. vs. Local 441, 10 IER Cases 146–150, 1994.

Summary of Comments and Subcommittee Responses
T/DM6-P: Blood Alcohol Testing in the Clinical Laboratory; Proposed Guideline
General Comments
1. Further information should be included on the clinical laboratory's obligation pertaining to clinical
specimens which become legal issues after the fact. Specimens drawn under clinical orders only are
often presented in court cases.
! The subcommittee does not believe it feasible to provide details beyond the information in Section 1.4
and interspersed elsewhere, because such matters vary substantially among different jurisdictions and
with changing case law.
2. The document should focus more on the analytical aspects of testing. Gas chromatography (GC) is
the method of choice and only 1-1/2 pages discusses GC.
! There is ample current information on alcohol analysis readily available in standard sources. These
include J Forensic Sci, J. Analyt Toxicol, Clin Chem, Garriott’s Medicolegal Aspects of Alcohol
Determination, 3rd ed., 1996, and many others. This document is not intended to serve as a
substitute for these analytical methods resources.
3. Consider adding a section that lists items that clinical laboratory personnel are often called upon to
address during alcohol-related court proceedings.
! A checklist of issues which commonly arise in legal proceedings which involve blood-alcohol analysis
results appears as Table 1 in Section 1.4.8.
4. The document indicates plasma, serum, or whole blood may be used. It indicates urine alcohol
concentrations are not well correlated with blood alcohol concentrations. It does not mention saliva.
I believe the document should say something about the use of saliva (i.e., some literature indicates
saliva may be used but this requires further investigation).
! While the subcommittee agrees it would be useful information, both breath and saliva testing are
beyond the scope of this document. These issues will be referred to the Area Committee on Clinical
Chemistry and Toxicology for consideration as the subject of a future NCCLS project.
5. In our approach to the regulation of alcohol devices, we make a distinction between alcohol analyses
obtained for medical purposes and alcohol analyses obtained for legal purposes. We regulate the
former but not the latter. Pages 9-10 of the proposed draft indicates this concept is invalid. It states
that results obtained for purely medical purposes may ultimately be used for legal purposes. This
implies we should regulate all uses and that all uses should have uniform labeling.
! Section 1.4 has been reworded to clarify this issue.
6. The document should be expanded to include concerns of a clinical laboratory as a reference

laboratory serving industrial-medical clients (especially in sections 2 and 3) versus off-site specimen
collection.
! Most matters related to blood alcohol testing in connection with industrial accidents/injuries are
covered by the text. Workplace testing for alcohol is a separate subject which is beyond the scope of
this document, and one in which blood-alcohol analysis plays a strictly limited role.

7. We do not agree that a blood alcohol test must have legal credibility. Our primary responsibility is to
the medical community of our region and to the health and well being of the citizens of our
community. Therefore, we provide the blood alcohol determination as a service to the physician in
order to determine a disease state without regard to how it may affect the patient's employment
and/or his standing with it. We would prefer to maintain a two level system so that a blood alcohol
can be measured without regard for the consent by the patient or for the establishment of criminal or
civil liability.
! A “two-level” system is quite acceptable, as long as there is awareness of potential future legal
aspects. Section 1.4 has been reworded to clarify this matter.
8. I object to the implicit assertion that a clinical laboratory performing blood alcohol testing for medical
purposes should institute chain-of-custody procedures. This would greatly increase the cost of
providing blood alcohol determinations. This in turn would contribute to the ever increasing spiral of
medical costs. Costs would be incurred for all tests although very few results may be subpoenaed.
Given the limited effectiveness of increased enforcement in reducing the incidence of driving under
the influence, the cost effectiveness of these increased expenditures would be extremely low.
! Sections 1.4.4 and 3 have been revised to clarify that chain-of-custody procedures apply chiefly to
known medicolegal situations. Moreover, Section 3.2 outlines the choice of omitting chain-of-custody
procedures for routine “clinical” situations.
9. In reviewing the proposed guidelines, I am appreciative that you have taken the time and interest to
investigate this area. I believe it is an area that has needed attention in the past. I am concerned,
however, that we do not go so far in structuring our testing for the judicial system, that we lose sight
of the fact that the physician and his patient are our primary responsibility and purpose.
! The subcommittee agrees and the Foreword and Scope of T/DM6-A have been revised to emphasize
that the primary responsibility is to the patient and the physician.
10. Many of the items discussed in this proposed guideline are impractical for a busy clinical laboratory.
Two examples of this are the requirement for controlled access to all specimens (Section 3.2) and the
suggestion that records should be maintained for each case in an individual file (Section 6.1). These
are not practical for a clinical laboratory serving an emergency room for instance, and will actually
slow down the speed of testing.
! The subcommittee agrees and Section 6.1 has been revised to include the use of computers in record-
keeping.
11. To be practical, it is necessary to distinguish between those specimens drawn to make a clinical
diagnosis, and those drawn as evidence in a legal proceeding. The two situations are not compatible;
a legal process is deliberately slow, while a clinical diagnosis on an emergency patient is deliberately
fast. It is unacceptable to slow down the process of arriving at a clinical diagnosis on the basis of
laboratory data based on the possibility that the results might be used in some legal process in the
future. If such a legal process is anticipated, then two specimens can be drawn at the same time,
one to be sent quickly for a medical diagnosis, and the other sent through the slow, deliberate process
to be used as evidence. What I think should be addressed by this proposed guideline are criteria for
what constitutes reasonable sample handling for the medical process.
! The subcommittee agrees that drawing replicate specimens may streamline the process. See Section
2.5.

12. Nowhere in this proposed guideline is the problem of calculating blood alcohol concentration at some
previous time based on a specimen collected later discussed. I'm sure that the authors did this
deliberately, feeling that such calculations are really the providence of an expert witness. If this is the
case, they should state so explicitly.
! The subcommittee considers the issue of extrapolation beyond the scope of the document. Section
1.4.7 has been revised to so indicate.
Section 1.0
13. In Section 1.2, page 186, the last sentence refers only to clinical chemistry tests in a laboratory
evaluation of a suspected alcohol-related patient condition. A laboratory workup also might include
hematologic and histopathologic tests.
! The document has been revised to include all relevant laboratory tests.
14. I concur that gas chromatography (GC) methods are more specific and are the methods of choice.
However, there is no discussion of whether confirmation is appropriate, particularly if methods other
than GC are used. Some discussion of the need for confirmation is appropriate in light of the
statement in Section 1.4 that medical purposes cannot be separated from legal purposes.
! Section 1.4.6 has been revised to address this comment.
15. I would like to take issue with the following statement: "The concept of obtaining a blood specimen
for medical purposes only is invalid because in most legal jurisdictions the laboratory results can be
subpoenaed in some circumstances." Obtaining a blood alcohol with the intent that it may later be
used in a criminal proceeding without obtaining a patient's consent for that use would in many states
be a violation not only of the patient's legal rights, but also of the physician-patient privilege. The
setting up of chain-of-custody procedures for alcohol determinations in a laboratory which does not do
forensic testing could be considered prima facie evidence that there was intent to use this result for
legal purposes.
! The relevant language in Section 1.4 has been revised to clarify the issue. The subcommittee,
however, does not agree with the commentor’s last two sentences.
16. I disagree with statements in Section 1.4. We run specimens for medical purposes only.
! The relevant language in Section 1.4 and elsewhere has been revised to clarify the issue.
Section 2.0
17. In Section 2.2, page 190, the first paragraph states that plasma and serum are more appropriate
specimens than whole blood for analysis, but then fails to state why whole blood is generally used.
! Section 2.2 has been reworded to clarify this matter.
18. Some states have passed an "implied consent law." What do you think the laboratory's responsibility
should be as far as obtaining consent from the patient. If another department (such as the Emergency
Room) takes care of having the consent forms signed, what do you feel to be the laboratory's
responsibility for ensuring that the forms have actually been signed prior to specimen collection?
! Because requirements vary significantly from state to state, it is necessary to obtain and follow your
state's regulation regarding informed consent. Recent case law has uniformly held that separate and
additional hospital (or laboratory) consent may not be demanded from a person who has consented to
an implied-consent blood-alcohol test.

19. Section 2.2, page 191, paragraph 3 recommends that laboratories not convert serum or plasma
values for alcohol to the whole blood equivalent, but states that courts should retain an expert to
make this conversion. At least a reference should be given explaining the problems with making such
conversions that would require an expert.
! A reference has been added as suggested.
20. If serum/plasma as a specimen is allowed there should also be help in interpretation.
! Appendix B has been added to assist in the interpretation of serum-alcohol concentrations.
21. Although povidone-iodine may be acceptable for cleansing the venipuncture site when only a blood
alcohol is to be obtained, it has been reported to interfere with other clinical assays and should not be
used when a specimen is being obtained for multiple tests which include a blood alcohol level.
! The recommendations in this document focus on specimen collection for alcohol testing. For further
information consult NCCLS document H3: Procedures for the Collection of Diagnostic Blood
Specimens by Venipuncture.
22. Sending samples out through the mail seems to be a procedure that would lead to legal problems.
! If accompanied by full identification and laboratory test report and mailed via registered mail or
certified mail, no problems should be encountered.
23. Section 2.3.4, page 195, should the collection of two specimens be considered instead of one when
the specimen is almost certain to be involved in a medical legal question?
! There is nothing to preclude taking two specimens if it is not clinically contraindicated and it meets
local legal rules. See response to Comment 11 above. A new Section 2.5 also discusses this matter.
24. In Section 2.3.4, I recommend that a specific time period be used in "If the specimen is to be
analyzed for alcohol content within a period of a few hours." Also use specific time period in the
second paragraph.
! The document has been revised to include a specific time period (i.e., within four hours).
25. In Section 2.3.4, the statement "If the analysis will be delayed. . .sufficient sodium fluoride to
produce a minimum concentration of 10 mg/mL" is inconsistent with a statement on page 194 which
indicates a 1.5 mg/mL NaF for short-term storage at 5 degrees C. Also a specific time period should
be specified in place of "short-term."
! See response to Comment 24 above. The statements are both correct. The higher NaF concentration
is needed if the specimen is not refrigerated or is transported.
26. Please define "extended periods of time" (page 194, second paragraph.)
! Section 2.3.4 has been reworded.
27. We don't usually identify the venipuncture site as indicated in Section 2.4. Is it necessary?
! Yes, because of i.v. sites. Blood collection, preparation of the puncture site, and related issues are
frequently raised in relation to blood-alcohol test results.

Section 3.0
28. We have a policy that a single technologist or technician draws the specimen, handles it exclusively,
and performs the analysis for ethanol. We have done this mainly to avoid having more than one
employee subpoenaed to testify in court. Therefore, we do not have a chain of custody form,
tamper-evident seals, or locked storage for our alcohols performed in house. Do you see a problem
with this?
! The subcommittee considers that this system, with proper documentation by the technologist, can
obviate the need for more complex procedures.
Section 4.0
29. The discussion of each analytical method should ideally address expected CVs.
! The subcommittee considers this beyond the scope of the document. Expected CVs are specific to
the laboratory and methodology; therefore, each laboratory should establish its own.
30. Three methods listed in the order of preference are stated as appropriate for clinical laboratories. They
are: gas chromatography (GC), enzymatic oxidation with alcohol dehydrogenase, and osmometry.
There is no mention of alcohol test strips employing alcohol dehydrogenase. I believe the draft could
make some kind of statement about such devices (i.e., they have been reported in the literature but
require further evaluation).
! Osmometry has been eliminated from the document. The other subject is beyond the scope of the
document.
31. It should be stated that osmometry, while helpful in clinical situations, is not suitable for legal
proceedings.
! Section 4.3 has been deleted in T/DM6-A.
32. In Section 4.2, you also say, "All ADH-based enzymatic oxidation methods are subject to potential
interference by isopropanol and higher alcohols to varying extents..." Published studies indicate that
the "ADH" enzyme currently in use is very specific for ethanol and that isopropanol and similar
organic substances (methanol, acetone, ethylene glycol) do not affect results.
! High concentrations of isopropanol and higher alcohols do affect these results. The extent of the
potential interference is a function of the individual test kit. The section has been reworded to clarify
that commonly-encountered concentrations of methanol and isopropanol do not significantly interfere.
33. In Section 4.3, page 206, an example of an osmolality calculation might be useful to include in this
section. The reference cited in the text might not be readily available in many laboratories, especially
in those laboratories not performing toxicology measurements on a regular basis.
! This section has been deleted in T/DM6-A.
34. The enzymatic method and method involving freezing point depression ought to be discouraged since
they are susceptible to interference from other alcohols.
! Freezing point osmometry has been deleted. Enzymatic methods are the most widely used test
methods for clinical applications. It is impractical to discourage their use at this time.
35. I would like to see a more detailed discussion of enzymatic methods, i.e., greater distinction between
alcohol dehydrogenase (ADH) and alcohol oxidase (AO), especially since most hospitals use ADH.

! Because of the length of such additional information, and changing commercial kits, it is not feasible
to incorporate that information into the document.
36. Maximum information on analytical interferences for all generic and/or specific methods considered
acceptable should be included.
! The subcommittee considers this beyond the scope of the document.
37. The listing of methods in Table 1, Summary of Methods for Blood Alcohol Analysis is incomplete.
Two methods used by some states are not listed; namely, Chemical Oxidation Following Distillation to
Separate Alcohol from Blood, and Chemical Oxidation Following Diffusion to Separate Alcohol from
Blood. It may be that these methods were omitted because of the limitation set forth in Paragraph
4.0 of the document that these are methods "appropriate for use in clinical laboratories."
Nevertheless, the focus of the document is the forensic application of the blood alcohol test,
especially as related to drunk driver arrests. Therefore, it appears reasonable to me that the
document should strive for completeness by listing and describing these two versions of the chemical,
oxidation-reductions tests for alcohol in blood.
! Chemical oxidation has become nearly extinct, and the subcommittee considers a discussion of it
unnecessary.
38. The methods of analysis section was short and did not provide as much information concerning
analysis and interferences that are possible in the analysis.
! The subcommittee considers this beyond the scope of the document. For further information consult
Medicolegal Aspects of Alcohol. J. Garriott (Ed.), 3rd ed., Tucson, AZ, Lawyers & Judges Publishing
Co., 1996.
39. Are you sure that sodium fluoride doesn't inhibit enzymatic methods?
! If used in the concentrations given, sodium fluoride doesn't inhibit enzymatic methods for ethanol
analysis, as evidenced by the CAP Survey results for Surveys AL1 and AL2.
40. Would you comment on the interference by hemolysis in the ACA method, as hemolysis is a
problem which commonly results from the use of potassium oxalate/NaF in the blood collection tube?
! Interference in spectrophotometric methods for chemical analysis using enzymatic oxidation can be
avoided by preparing a 1:3 or 1:4 protein-free filtrate and analyzing it. For details, consult Gadsden
RH and Taylor EH. Ethanol in biological fluids by enzymatic analysis. In: Selected Methods of
Emergency Toxicology. Washington DC, AACC Press, 1986, pp. 63–65.
41. Please add references for interference by isopropanol, higher alcohols (these should be named), and
methanol.
! Such information should be obtained from the manufacturer of a given reagent kit, such as package
inserts and analysis manuals.
Section 5.0
42. I recommend that Section 5.0, Quality Assurance, begin with a new section 5.1 entitled “Blanks.”
The value and usefulness of blank samples in quantitative procedures is well established. Their
particular value and usefulness in the forensic application of blood alcohol tests is clear.
! The use of blank specimens is required but their incorporation in the test procedure is method
dependent.

43. Under Section 5.1 “Calibrators (Standards),” the density of ethanol should be stated. This would
assist people who are preparing standards on a weight basis using volumetric equipment.
! That information is readily available in standard reference manuals, e.g., The Merck Index, which
should be available in all laboratories.
44. The requirement to run a calibrator with every analysis and every batch, discussed in Section 5.1, is
unreasonable and excessive for many analytical systems. The requirement should specify that this be
done for GC methods or on batch or random access analyzers that require standards with each
analytical run for any test.
! Sections 5 and 5.1 have been rewritten to clarify this point. However, it is good practice to run at
least one positive and one negative control with each patient sample for a determination that has such
high probability of legal consequences as alcohol; documentation is important.
45. Under Section 5.2, “Controls,” the frequency that controls should be run does not correspond to the
requirements for some instruments. This enzymatic method is in frequent use in the clinical
laboratories. Generally controls are run on this instrument once a day. If the authors feel that this is
insufficient for legal proceedings, then this problem should be addressed specifically. This is true of
the standards also.
! See response to Comment 44.
46. I question the need to initiate every alcohol test with calibrators.
! See response to Comment 44.
47. I recommend the addition of two concepts to Section 5.2, “Controls.” First, it should be stated that
when a laboratory is setting the mean and standard deviation for its controls, the 10 or 20 replicate
analyses should be done at the rate of no more than 2 per day. Secondly, the goal of a quality
control program would be better stated if, at the end of Section 5.2, there were language to the
effect that: "Whenever analysis of controls is outside the acceptable limits, the method shall be
regarded to be in error, and the laboratory must take remedial action to investigate and correct the
source of error. Until such time that the error has been corrected, as shown by return of analysis of
controls to values within the acceptable limits, no samples will be analyzed for blood alcohol."
! A brief comment has been added to Section 5.2 to confirm the need for action when out-of-control
results occur; and for assignment of control values and limits, reference has been made to NCCLS
document C24 to clarify these matters.
48. You made reference to control in this guideline, but we have not been able to locate a whole blood
control. Can you specify the type of controls you make reference to?
! There are commercial preparations listed in laboratory supply catalogs and available through
manufacturers of control materials.
Section 6.0
49. We have a notebook which lists patients by name and number as well as their ethanol concentration
and the value of the control that was run. It also includes the date, time, and initials of the person
performing the test. It is a chronological file and we have no alphabetical or numerical cross-
reference. It is stored on a shelf in our chemistry lab and it is not locked. Are you suggesting a
computer cross-reference system? Do you feel our present filing system is adequate? You suggested
a ten year storage time. The statute of limitations has been shortened, is this still your recommended
time?

! Any system is subject to challenge, but one can minimize the likelihood of successful challenge. The
notebook mentioned in this comment should be kept under lock and key when not in actual use in the
laboratory.
Section 7.0
50. On page 211 Section 7.1.2, referring to Blood Alcohol Concentration (BAC) with respect to rounding
off from 3 digits to 2 digits, you state 139 mg/dl, 0.139% (g/dl) would round off to 0.13%. Are you
saying that the third digit is always dropped rather than traditional rounding off. For example, would
0.099 round off to 0.09 or to 0.10?
! Yes, the universal practice is to truncate, i.e., drop the third decimal entirely, not round upward. This
section has been revised to clarify when and how you round to a two-digit value.
51. I would like to see a reference for the statement "some individuals are impaired at less than 0.10%
w/v; others are not measurably impaired at much higher concentrations." I am not aware of any
studies which show that there are individuals whose performance is the same at blood alcohol levels
of 0.0 and 0.1% when measured quantitatively (e.g., reaction time). If there are no such studies,
perhaps it would be better to state that "others are not obviously impaired at much higher
concentrations" or that some individuals may appear to be unimpaired at much higher concentrations.
! The document has been changed as suggested (see Section 7.2).
52. I think the expression of concentration as percent should be eliminated. If % (w/v) = g/dl, then let's
say so. Then there would be no assumption as to whether we mean % (w/v) or % (w/w). This
sometimes is a major problem in court.
! The percent weight/volume notation is ingrained in many regional laws and state regulations, and
some federal laws. The subcommittee cannot affect these practices.

Related NCCLS Publications†
C24-A Internal Quality Control Testing: Principles and Definitions; Approved Guideline (1991).
Discusses the purpose of internal quality control; defines various analytical intervals, such as
"analytical run"; and addresses the use of quality control material and control data, including
the use of data in quality assurance and interpretation.
EP10-T2 Preliminary Evaluation of Quantitative Clinical Laboratory Methods - Second Edition; Tentative
Guideline (1993). Discusses experimental design and data analysis for preliminary evaluation
of the performance
GP2-A3 Clinical Laboratory Technical Procedure Manuals–Third Edition; Approved Guideline (1996).
Offers guidelines that address the design, preparation, maintenance, and use of technical
procedure manuals in the clinical laboratory.
H3-A3 Procedures for the Collection of Diagnostic Blood Specimens by Venipuncture–Third Edition;
Approved Standard (1991). Discusses methods for the collection of blood specimens by
venipuncture and an appropriate training program aimed at increasing analyte integrity and
minimizing laboratory error. Includes a 24-step protocol for specimen collection,
recommendations for "order of draw," and considerations for performing venipuncture on
children.
H5-A3 Procedures for the Handling and Transport of Diagnostic Specimens and Etiologic Agents-Third
Edition; Approved Standard (1994). American National Standard. Gives proper packaging,
handling, and transport requirements for medical specimens. Includes federal regulations.
H18-A Procedures for the Handling and Processing of Blood Specimens; Approved Guideline (1990).
Addresses the multiple factors associated with handling and processing specimens, factors
that can introduce imprecision or systematic bias into test results.
H31-P Collection Containers for Specimens for Toxicological Analysis; Proposed Guideline (1986).
Discusses recommended toxicology/drug monitoring requirements for blood collection
containers.
M29-T2 Protection of Laboratory Workers from Infectious Disease Transmitted by Blood, Body
Fluids, and Tissue—Second Edition; Tentative Guideline (1991). Guidance on the risk
of transmission of hepatitis B virus and human immunodeficiency viruses in the
laboratory; specific precautions for preventing transmission of bloodborne infection
during clinical and anatomical laboratory procedures.
†
Proposed- and tentative-level documents are being advanced through the NCCLS consensus process; therefore, readers
should refer to the most recent editions.

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T/DM6-A

Blood Alcohol Testing in the Clinical Laboratory;

! The revision of documents in response to comments by VOLUNTEER PARTICIPATION

Blood Alcohol Testing in the Clinical Laboratory;

NCCLS VOL.17 NO.14 i

NCCLS VOL.17 NO.14 ii

Blood Alcohol Testing in the Clinical Laboratory;

Kurt M. Dubowski, Ph.D.

Copyright ©1997. The National Committee for Clinical Laboratory Standards.

NCCLS VOL.17 NO.14 iv

Area Committee on Clinical Chemistry and Toxicology

Basil T. Doumas, Ph.D. Medical College of Wisconsin

W. Gregory Miller, Ph.D. Medical College of Virginia Hospital

Subcommittee on Blood Alcohol Testing

Kurt M. Dubowski, Ph.D. The University of Oklahoma

Patricia H. Field, Ph.D. WI State Laboratory of Hygiene

Walter Frajola, Ph.D. Columbus, Ohio

Vidmantas A. Raisys, Ph.D. University of Washington

Robert H. Reeder, J.D. Northwestern University

M. Jeffery Shoemaker, Ph.D. Pennsylvania Dept. of Health

Vana L. Smith, Ph.D. Abbott Laboratories

Sharon S. Ehrmeyer, Ph.D. University of Wisconsin

Beth Ann Wise, M.T.(ASCP), M.S.Ed. NCCLS

Patrice E. Polgar NCCLS

NCCLS VOL.17 NO.14 v

ACTIVE MEMBERSHIP (as of 1 July 1997)

Sustaining Members College of Physicians and Government of Malta

NCCLS VOL.17 NO.14 vi

NCCLS VOL.17 NO.14 vii

NCCLS VOL.17 NO.14 viii

OFFICERS BOARD OF DIRECTORS

John V. Bergen, Ph.D.,

NCCLS VOL.17 NO.14 ix

1 Scope and Requirements . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

1.1 Scope of the Problem . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

2.1 Responsibility for Specimen Collection . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 3

4.1 Gas Chromatography . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 8

5.1 Calibrators (Standards) . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 11

6 Reporting and Significance of Results . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

6.1 Blood Alcohol Records . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

7 Terminology and Statutory Provisions . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

NCCLS VOL.17 NO.14 x

Alcohol, blood alcohol, analysis, blood alcohol concentration, chain-of-custody, intoxication.

NCCLS NO.17 VOL.14 xi

Blood Alcohol Testing in the Clinical Laboratory; Approved Guideline

NCCLS NO.17 VOL.14 1

1.3 Industrial-Medical Requirements exclusively legal purposes such as traffic law

NCCLS NO.17 VOL.14 2

1.4.5 Security System 2.1 Responsibility for Specimen Collection

This must be utilized, when appropriate, to

There are many requirements for interpretation of 2.2 Types of Specimens

NCCLS NO.17 VOL.14 3

Laboratory and Personnel

! Identity, status, and qualifications of the person giving testimony

! Analysis protocols and standard operating procedures; literature references

! Overall quality assurance/quality control schemes

! Scientific validity of the test results

NCCLS NO.17 VOL.14 4

Alcohol, at equilibrium, is generally distributed be expected to significantly affect the alcohol