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    Alkaline Phosphatase

    Uploaded by

    Yasir Mushtaq
    This document outlines a method for determining alkaline phosphatase levels from a blood sample. The enzymatic method works by using alkaline phosphatase to convert 4-nitrophenolphosphate to phosphate and 4-nitrophenolate, which can be measured photometrically. The procedure involves taking a blood sample, allowing it to clot, and centrifuging it to obtain serum. The serum is incubated with reagent solution and measured at 405nm to determine enzymatic activity levels, which are then used to calculate alkaline phosphatase levels. Elevated levels can indicate various physiological and pathological bone and liver conditions.

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    Alkaline Phosphatase

    Uploaded by

    Yasir Mushtaq
    340 views2 pages
    This document outlines a method for determining alkaline phosphatase levels from a blood sample. The enzymatic method works by using alkaline phosphatase to convert 4-nitrophenolphosphate to phosphate and 4-nitrophenolate, which can be measured photometrically. The procedure involves taking a blood sample, allowing it to clot, and centrifuging it to obtain serum. The serum is incubated with reagent solution and measured at 405nm to determine enzymatic activity levels, which are then used to calculate alkaline phosphatase levels. Elevated levels can indicate various physiological and pathological bone and liver conditions.

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    practical on Alkaline Phosphatase

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    This document outlines a method for determining alkaline phosphatase levels from a blood sample. The enzymatic method works by using alkaline phosphatase to convert 4-nitrophenolphosphate to phosphate and 4-nitrophenolate, which can be measured photometrically. The procedure involves taking a blood sample, allowing it to clot, and centrifuging it to obtain serum. The serum is incubated with reagent solution and measured at 405nm to determine enzymatic activity levels, which are then used to calculate alkaline phosphatase levels. Elevated levels can indicate various physiological and pathological bone and liver conditions.

    Copyright:

    © All Rights Reserved
    Download as DOC, PDF, TXT or read online from Scribd
    Download as doc, pdf, or txt
    340 views2 pages

    Alkaline Phosphatase

    Uploaded by

    Yasir Mushtaq
    This document outlines a method for determining alkaline phosphatase levels from a blood sample. The enzymatic method works by using alkaline phosphatase to convert 4-nitrophenolphosphate to phosphate and 4-nitrophenolate, which can be measured photometrically. The procedure involves taking a blood sample, allowing it to clot, and centrifuging it to obtain serum. The serum is incubated with reagent solution and measured at 405nm to determine enzymatic activity levels, which are then used to calculate alkaline phosphatase levels. Elevated levels can indicate various physiological and pathological bone and liver conditions.

    Copyright:

    © All Rights Reserved
    Download as DOC, PDF, TXT or read online from Scribd
    Download as doc, pdf, or txt
    of 2

    OBJECT: DETERMINATION OF ALKALINE PHOSPHATASE FROM

    GIVEN SAMPLE OF BLOOD


    METHOD: ENZYMATIC METHOD

    Principle:
    4-Nitrophenolphosphate + H2O
    nitrophenolate

    ALP

    Phosphate + 4-

    When enzyme acts depending upon the substance nature, phosphate


    is liberated. The rate of increase of nitrophenolate is directly
    proportional to enzyme activation and this can be measured photo
    metrically.

    REAGENT SOLUTIONS:
    Diethanolamine HC, Ph.9.8 4-nitrophenolphosphate Mg+ chloride

    EQUIPMENTS:
    1. Disposable Syringe, Pipettes, Test tubes, Centrifuge machine,
    Water bath and Spectrophotometer.

    SAMPLE:
    Take 2 cc/ml of blood by disposable syringes in a clean test tube,
    allow to clot and centrifuge it.

    PROCEDURE:
    Reagent temperature
    37 0C, 30 oC,25 oC
    Wavelength
    405nm
    Light path
    1 cm
    Measuring Temperature 37 0C,
    Take 0.02 ml (20Ul) of serum and add I ml of reagent solution. Now
    mix well and put it in water bath to maintain it on 37 0c. Now after
    one or two minutes, measure in absorbance using photometer at
    405nm and K-20 against the air blank on a factor 2760.

    CALCULATION:
    Enzymatic Activity X Factor
    RESULT:
    Given known sample contains--------------------NORMAL VALUE:
    Female = 65-306 U/ L at 37 oC.
    Male =
    80-306 U/ L at 37 oC

    CLINICAL INTERPRETATION:
    INCREASE ALKALINE PHOSPHATASE:
    Physiological Causes.
    During bone growth in childhood and in Pregnancy
    Pathological Causes.
    Obstructive Jaundice, Billary Diseases (Steatorrhea)
    Extra hepatic and intrahepatic cholestasis, Infectious hepatitis.
    Bone conditions osteomalacia, Rickets,
    Bone Tumors Osteoblastic Sarcoma, Pagets, Bone Metastases
    Diseases hyperthyroidism
    Diseases hyperparathyroidsm (Increase paratharmone)

    GENERAL COMMENTS
    Iso-forms
    Liver, Brain, Heart Muscle, Skeletal Muscle And RBC.

    Alkaline Phosphates (ALP) a hydrolytic enzyme acting optimally at


    alkaline pH, exists in blood in numerous distinct forms which originate
    mainly from liver and bone but also from other tissues as kidney placenta
    testes thymus, lung and tumors.

    EDTA anticoagulant (Ethylene diamine Tetra Acetol) also inhibits


    enzyme by taking Mg++ ions

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