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    Amylase

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    Dinesh Sreedharan
    This document provides instructions for determining α-Amylase activity using a CNP G3 substrate method. α-Amylase cleaves CNP G3, producing CNP which is monitored at 405nm. Elevated levels of α-Amylase are useful for diagnosing acute pancreatitis. The linear range of the assay is up to 2000 IU/L of α-Amylase. The reagent is stable stored between 2-8°C and ready to use.

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    Amylase

    Uploaded by

    Dinesh Sreedharan
    53 views2 pages
    This document provides instructions for determining α-Amylase activity using a CNP G3 substrate method. α-Amylase cleaves CNP G3, producing CNP which is monitored at 405nm. Elevated levels of α-Amylase are useful for diagnosing acute pancreatitis. The linear range of the assay is up to 2000 IU/L of α-Amylase. The reagent is stable stored between 2-8°C and ready to use.

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    This document provides instructions for determining α-Amylase activity using a CNP G3 substrate method. α-Amylase cleaves CNP G3, producing CNP which is monitored at 405nm. Elevated levels of α-Amylase are useful for diagnosing acute pancreatitis. The linear range of the assay is up to 2000 IU/L of α-Amylase. The reagent is stable stored between 2-8°C and ready to use.

    Copyright:

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

    Amylase

    Uploaded by

    Dinesh Sreedharan
    This document provides instructions for determining α-Amylase activity using a CNP G3 substrate method. α-Amylase cleaves CNP G3, producing CNP which is monitored at 405nm. Elevated levels of α-Amylase are useful for diagnosing acute pancreatitis. The linear range of the assay is up to 2000 IU/L of α-Amylase. The reagent is stable stored between 2-8°C and ready to use.

    Copyright:

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

    α-AMYLASE

    (CNP G3 Method)
    For the determination of α-Amylase activity in serum, plasma or urine.
    (For In vitro Diagnostic Use Only)

    SUMMARY:
    α-Amylase is most frequently measured in the diagnosis of acute pancreatitis when serum levels may be grossly
    elevated and therefore is useful diagnostic tool in critical care management. In acute pancreatitis α-Amylase levels
    reach a peak at 24 hours and remain elevated from 3-7 days. Hyperamylasemia is also associated with other acute
    abdominal disorders, biliary tract diseases, diabetic ketoacidosis, severe glomerular dysfunction, salivary gland
    disorders, and ruptured ectopic pregnancy.

    PRINCIPLE:

    Substrate CNP G3 is directly hydrolyzed by α-Amylase to produce CNP monitored at 405 nm during incubation of
    the reagent with a sample.

    α-Amylase
    CNP G3 CNP + CNP G 2+ G 3+ G

    CNP G3 = 2-Chloro-4-nitrophenyl-α-Maltotrioside
    CNP G2 = 2-Chloro-4-nitrophenyl-α-Maltoside
    CNP = 2-Chloro-4-nitrophenol

    CONTENTS:

    PACK SIZE AMYLASE REAGENT(A1)

    4x5ml 4x5ml

    1x25ml 1x25ml

    STORAGE & STABILITY:

    Reagent is stable at 2 – 8⁰C till the expiry mentioned on the label.

    REAGENT PREPARATION:

    Reagents are ready to use.

    SAMPLE MATERIAL:

    Serum, heparinised plasma, urine.α-Amylase is reported to be stable in the sample for 5 days at 2–8ºC. Separate
    serum from clot as soon as possible.

    GENERAL SYSTEM PARAMETERS:


    Reaction Mode Kinetic Sample Volume 25 µl
    Wavelength 405 nm Reagent volume 1000 µl
    Blank Distilled Water Factor 4640
    Temperature 37˚C Reaction Slope Increasing
    Delay Time 60 sec Linearity 2000 IU/L
    Read Time 60 sec Units IU/L

    ASSAY PROCEDURE:

    Wavelength/ Filter : 405 nm


    Temperature : 37˚C
    Light Path : 1 cm
    Pipette into clean dry test tube labelled as Test (T):

    Addition Sequence T(µl)


    Amylase Reagent 1000
    Sample 25
    Mix well and read the initial absorbance A0 after 1 minute and subsequently 2 more readings with 30 seconds
    interval at 405 nm. Calculate the mean absorbance change per minute
    (∆ Abs/30 seconds x 2)

    CALCULATIONS:
    α-Amylase activity in IU/L (Serum) = ∆A/min. x 4640

    NORMAL VALUE:
    Serum upto 140 IU/L

    LINEARITY:

    The procedure is linear upto 2000 IU/L at 37OC. If the amylase activity is above 2000 IU/L, dilute the specimen
    suitably with normal saline. In such case the results obtained should be multiplied by dilution factor to obtain
    correct amylase activity.

    NOTE:

    Anticoagulants like Oxalate and EDTA bind Calcium, which is needed for α-Amylase activity and should not be
    used. Heparin may be used. Saliva and sweat contain α-Amylase. Avoid contamination of reagent and sample
    during use. Reagent should not be used if its absorbance exceeds 0.8 at 405 nm against distilled water.

    REFERENCE:

    1.Jungi W.et.al. Biochem.22,109 (1089)


    2.Winn-Deen E.S. David H.Signett(1988)

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