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    Lab Assays PDF

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    Meevie Toledo
    The document describes several laboratory assays for measuring various enzymes and substrates. Amylase is measured through disappearance of starch, appearance of reducing sugars, or increasing color or absorbance. Lipase assays include titrimetric measurement of fatty acid liberation from olive oil or turbidimetric clearing of substrate. AST and ALT are measured via coupled enzymatic reactions monitoring NADH. ALP assays detect p-nitrophenol production. Creatine kinase assays use coupled reactions to monitor NADH or NADPH. Lactate dehydrogenase is measured via forward or backward reactions detecting NADH. GGT is assayed through detection of p-nitroaniline production. Cholinesterase assays use Ellman's reagent

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    Lab Assays PDF

    Uploaded by

    Meevie Toledo
    107 views2 pages
    The document describes several laboratory assays for measuring various enzymes and substrates. Amylase is measured through disappearance of starch, appearance of reducing sugars, or increasing color or absorbance. Lipase assays include titrimetric measurement of fatty acid liberation from olive oil or turbidimetric clearing of substrate. AST and ALT are measured via coupled enzymatic reactions monitoring NADH. ALP assays detect p-nitrophenol production. Creatine kinase assays use coupled reactions to monitor NADH or NADPH. Lactate dehydrogenase is measured via forward or backward reactions detecting NADH. GGT is assayed through detection of p-nitroaniline production. Cholinesterase assays use Ellman's reagent

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    LAB ASSAYS.pdf

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    The document describes several laboratory assays for measuring various enzymes and substrates. Amylase is measured through disappearance of starch, appearance of reducing sugars, or increasing color or absorbance. Lipase assays include titrimetric measurement of fatty acid liberation from olive oil or turbidimetric clearing of substrate. AST and ALT are measured via coupled enzymatic reactions monitoring NADH. ALP assays detect p-nitrophenol production. Creatine kinase assays use coupled reactions to monitor NADH or NADPH. Lactate dehydrogenase is measured via forward or backward reactions detecting NADH. GGT is assayed through detection of p-nitroaniline production. Cholinesterase assays use Ellman's reagent

    Copyright:

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

    Lab Assays PDF

    Uploaded by

    Meevie Toledo
    The document describes several laboratory assays for measuring various enzymes and substrates. Amylase is measured through disappearance of starch, appearance of reducing sugars, or increasing color or absorbance. Lipase assays include titrimetric measurement of fatty acid liberation from olive oil or turbidimetric clearing of substrate. AST and ALT are measured via coupled enzymatic reactions monitoring NADH. ALP assays detect p-nitrophenol production. Creatine kinase assays use coupled reactions to monitor NADH or NADPH. Lactate dehydrogenase is measured via forward or backward reactions detecting NADH. GGT is assayed through detection of p-nitroaniline production. Cholinesterase assays use Ellman's reagent

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    LABORATORY ASSAYS

    AMY
    1. Amyloclastic measures disappearance of starch
    (substrate)

    2. Saccharogenic measures the appearance of the


    reducing sugars product (Classic method)

    3. Chromogenic measure the increasing color or


    absorbance due to product formation

    4. Continuous Monitoring measures the change in


    absorbance of NAD (340nm; pH 6.9)

    LPS
    1. CHERRY-CRANDALL (Titrimetric)
    Substrate: Olive oil
    Product: Liberated fatty acids

    2. Turbidimetric measures the hydrolysis of fates by its rate


    of clearing substrate

    3. Colorimetric uses peroxidase or glycerol kinase (pH 8.8)

    AST
    1. KARMEN (Coupled enzymatic)
    Indicator enzyme: Malate dehydrogenase
    Measures coenzyme: NADH (340nm; pH 7.3-7.8)

    ALT
    1. WROBLEWSKI-LADUE (Coupled enzymatic)
    Indicator enzyme: Lactate dehydrogenase
    Measures coenzyme: NADH (340nm; pH 7.3-7.8)

    2. REITMAN-FRANKEL (Colorimetric enzymatic)


    Uses: alkaline medium
    Detects: 2,4-Dinitrophenly hydrazone (BROWN)

    ALP
    1. BOWERS AND MCCOMB (continuous monitoring)
    Substrate: p-nitrophenly phosphate
    Product: p-nitrophenol (405 nm; pH 10.2)

    2. BESSEY-LOWRY-BROCK

    ACP
    1. MODIFIED BOWERS AND MCCOMB (continuous
    monitoring)
    Substrate: p-nitrophenly phosphate
    Product: p-nitrophenol (405 nm; pH 5.0)

    2. QUANTITATIVE ENDPOINT

    Nitrophenylphosphate = Nitrophenol + Phosphate


    Thymolphthalein Monophosphate =
    Thymolphthalein + Phosphate

    3. CONTINUOUS-MONITORING

    Alpha-naphthyl phosphate = Alpha-naphthol + Phosphate


    Creatine Kinase (CK)
    1. TANZER-GILVARG (Forward Reaction)
    Uses: Creatine Kinase; Pyruvate Kinase
    Indicator enzyme: Lactate Dehydrogenase
    Coenzyme: ADP (coupled in the second reaction)
    Measures: NADH (340 nm; pH 9.0)

    2. OLIVER ROSALKI (Backward Reaction)


    Uses: Creatine Kinase; Hexokinase
    Indicator enzyme: G6PD
    Coenzyme: ATP (coupled in the second reaction)
    Measures: NADPH (340 nm; pH 6.8)

    Lactate Dehydrogenase
    1. WACKER REACTION (Forward)
    Substrate: Lactate
    Measures: NADH (340nm; pH 8.3 to 8.9)

    2. WROBLEWSKI-LADUE (Backward)
    Substrate: Pyruvate
    Measures: NADH (340nm; pH 7.1 to 7.4)

    3. Alpha-Hydroxybutyrate Dehydrogenase Activity


    uses alpha-hydroxybutyrate as substrate which has
    more affinity to the H subunit thus making it specific for
    LD-1.

    GGT
    1. SZASZ ASSAY
    Substrate: Gamma-Glutamyl-p-Nitroanilide
    Measures: p-nitroaniline (405-420nm; pH 8.2)

    5-nucleotide
    Uses the chemical reaction of 5-NT and detects
    ribonucleoside production

    G6PD
    Uses the chemical reaction catalysed by G6PD in the
    body and detects NADPH @ 340nm

    Cholinesterase
    Substrates:
    Acylthiocholine
    Acetylthiocholine
    Proprionylthiocholine
    Succinylthiocholine
    Reagent: Ellmans Reagent (Dithiobisnitrobenzoic
    acid)
    Product: 5-mercapto-2-nitrobenzoic acid @ 410nm

    ACE
    Detects Hippuric Acid @ 228nm

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