Histopathology Specimen

collection and handling

Objectives:
• Identify histopathology specimens.
• Processing and handling of histopathology
specimen.
• Gross examination
Histopathology
Specimens for histopathology laboratory
• Biopsy specimens
• Surgical specimens
• Cytology specimens
• Autopsy specimens
• Frozen sections
• Special stains
• Immunohistochemistry (IHC) specimens
• Electron microscopy specimens
• Biopsy specimens:

• Biopsy specimens are small tissue samples

collected from a specific area of the body for
diagnostic purposes by the clinicians. They are
typically handled by fixing them in formalin to
preserve the tissue structure and prevent
degradation.
• Biopsies can be obtained:
• - Needle biopsy
• - Endoscopic biopsy
• - Incisional biopsy
Surgical specimens:
• Surgical specimens are larger tissue samples
removed during surgical procedures. These
specimens are usually handled by immediately
placing them in formalin to preserve the tissue
integrity and prevent contamination.
• Example :Organ resections,Lymph node
dissections,Tumor resections,Gastrointestinal
resections,Bone and joint surgeries,Reproductive
organ surgeries. Etc.
Cytology specimens:

• Cytology specimens consist of individual cells or

small groups of cells collected from various body
fluids or tissues. These specimens are often fixed in
alcohol or other fixatives to preserve cell
morphology for examination.

• Example :Fine needle aspirations (FNA), pap

smears,Body fluid cytology, urine cytology, sputum
cytology, effusion cytology. Etc..
Autopsy specimens:
• Autopsy specimens: Autopsy specimens are tissue
samples collected during postmortem examinations
to determine the cause of death. These specimens
are handled with care to prevent contamination
and degradation, and they may be fixed in formalin
for preservation.
Conti....
• Frozen sections: Frozen sections are fresh tissue
samples that are rapidly frozen, sliced, and stained for
immediate examination during surgery. These specimens
require special handling techniques to maintain tissue
integrity and prevent artifacts.
• Special stains: Special stains are used to highlight
specific components of tissues, such as proteins,
carbohydrates, or microorganisms. These stains require
precise handling and processing to ensure accurate
results and interpretation.
Conti...
• Immunohistochemistry (IHC) specimens: IHC
specimens are tissue samples processed for
immunohistochemical staining to detect specific
proteins or antigens within the tissue. These
specimens require careful handling to preserve
antigenicity and ensure accurate staining results.
• Electron microscopy specimens: Electron
microscopy specimens are tissue samples
processed for ultrastructural analysis at the cellular
level. These specimens require specialized handling
techniques to preserve fine structures and maintain
sample integrity for electron microscopy imaging.
Conti....

• Overall, proper handling and processing of

histopathological specimens are essential to ensure
accurate diagnosis and interpretation of tissue
abnormalities and diseases.
 Specimen collection and handling..
• In histopathology laboratory, proper biopsy preparation
and correct diagnosis depend on proper tissue collection
and handling.

• Proper tissue handling avoids the grave mistakes

committed by incorrect labeling and mixed up of the
tissue biopsy and request form.

• The integrity and the quality of the specimen are vital to

good histopathological diagnosis.
• Laboratory personnel must have appropriate knowledge
of proper tissue handling.
 Histopathological protocols
• Taking a specimen
• The surgeon must be familiar with the conventions
of, and use of,orientation-marker suturing on tissue
(for example, lateral = long suture and superior =
short suture).
• The nature of the tissue, site and side must be
communicated.
• For certain diagnoses (for example, possible
malignant lymphomas equiring impression smears
or flow cytometry), fresh tissue is required and
must be available.
Taking a specimen....
• Desiccation of tissue should be minimized by
immediately placing it into fixative.

• The specimen container should be large enough not

to distort the specimen in the container,
• The fixative should have free access to all surfaces
of the specimen.
Histopathological protocols…

• Containers
• Containers should be of glass with screw-top lids
(reusable) or plastic, with leak-proof lids
(disposable).
• There should be a range of sizes available, and
containers should be clean and uncontaminated,
• particularly by other tissue, (labels should not be
affixed to containers prior to placing the specimen
in prelabelled containers.
Histopathological protocols…

•A
Histopathological protocols…
• Labelling should be performed after the specimen is
placed in the container.
•  Two patient identifiers are required through the
whole processing of the specimen. These should be on
the original specimen container as a minimum.
• Labelling requires at least two of:
•  patient's name
•  patient's date of birth
•  laboratory number
•  hospital number. That to be on the container and
remain on all the cassettes and slides in the
subsequent processing.
Histopathological protocols…
Histopathological protocols…
• The preservation of biopsy, surgical and cytological samples becomes
a priority in order to ensure a correct and complete diagnosis.
• Good preservation of tissue is the most important factor in the
production of satisfactory histology slides.
• If the preservation has been inadequate good sections are
absolutely impossible.
• The preservation required is of a special type known as fixation.
• There are three essentials to good fixation:
•  Fresh tissue,
•  Proper penetration of the fixative, and
•  the right choice of a correctly formulated fixative.

• Surgical tissues should be placed in fixative immediately they are

removed and sent to the laboratory as soon as possible, where
proper fixation can be supervised.
 Histopathological protocols…
• Steps required in preparing tissues for light microscopy include:
• 1. TISSUE ACCESSION:
•  Upon reception of tissue specimens, the following points should
be noted:
• 1. The laboratory staff receiving the biopsy specimen must always
match the ID of the patient on the request form with that on the
specimen container.
• 2.Tissue and organ specimens should be submitted in a wide-
mouthed
• capacious container in order to avoid distortion.
• 3. The tissue or organ submitted should be immersed with adequate
• amount of fixative.
• 4. Label the tissue or organ specimen properly.
• 5. Record all the relevant information on the laboratory record book.
 Specimens processing and handling
• TISSUE ACCESSION:
•  For routine tissue processing by paraffin-embedding
technique, the
• tissue must be put in either;
Specimens processing and handling...
• GROSS EXAMINATION: Done at the gross room
•  Gross examination of the specimen received in the
laboratory is the next most important step.
•  Proper gross tissue cutting, gross description and
selection of representative tissue sample in larger
specimens is a crucial part of the pathologic examination of
tissue submitted.
•  Calcified tissues and bone are subjected to decalcification
to remove the mineral and soften the tissue by treatment
with decalcifying agents such as acids and chelating agents
(most often aqueous nitric acid).
•  It is mandatory that all the gross-room personnel follow
strict precautions in handling the tissues infected with
tuberculosis, hepatitis, HIV and others.
Specimens processing and handling...
• Gross Examination
• 1. Proper identification and orientation of the
specimen.
• 2. Unlabeled specimen should never be processed.
• 3. A properly completed histopathology requisition
form containing patient’s name, age, sex, relevant
clinical data, surgical findings, nature of operation
and name of tissue submitted.
• 4. Careful search and examination of all the tissue
submitted in order
Specimens processing and handling
Gross Examination......

• 5. Place the specimen on cutting board in an anatomic

position and
• record the following information: types of specimen,
Structure
• included, Dimensions, Weight, Shape, Colour, Consistency
and
• Surgical margin, whether included or not involved by tumor.
• 6. Measurements are usually given in centimeter unless the
specimen
• is very small in which mm can be used.
• 7. Endometrial and prostatic tissue should be measured by
aggregate
Steps of Histopathological techniques

•  Histopathological technique deals with the preparation of

tissue for microscopic examination. It has series of processes:
• 1. Sample accessation
• 2. Labeling
• 3. Fixation
• 4. Dehydration
• 5. Clearing
• 6.impregnation
• 7. Embedding
• 8. Sectioning
• 9. Staining
• 10. Mounting
• 11. Microscopic examination and reporting
Histopathological Tissue Preparation

•  Various techniques have been developed to

prepare tissues for study so that they closely
resemble their natural, living state.
• Responsibility of a technician/ technologists
•  The technician is responsible for:
• 1. Specimen preservation.
• 2. Specimen labeling, logging and identification.
• 3. Preparation of the specimen to facilitate their
gross and microscopy.
• 4. Record keeping.
Specimen collection and handling

•  In histopathology laboratory, proper biopsy preparation and

correct
• diagnosis depend on proper tissue collection and handling.
•  Proper tissue handling avoids the grave mistakes committed by
• incorrect labeling and mixed up of the tissue biopsy and request
form.
•  Another problem is the tissue placed in narrow mouthed bottle
that
• necessitates breaking of the container in order to take out the
tissue.
•  Sometimes the tissue biopsy may be placed in inadequate
amount of
• formalin.
•  Laboratory personnel must have appropriate knowledge of
proper
Gr-4 Na