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  • Determination of Plasma Total-And Hdl-Choleterol

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    趙偉光
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    This document describes a method for determining total and HDL cholesterol levels in plasma. It involves using cholesterol esterase and cholesterol oxidase to convert cholesterol to a compound that reacts with other reagents to form a colored complex, which is then quantified spectrophotometrically. Specifically, it precipitates LDL and VLDL from plasma to obtain the HDL fraction, and measures total cholesterol without precipitation to determine overall levels. The absorbance readings are used to calculate cholesterol concentrations based on a standard reference value.

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    胆固醇测定

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    This document describes a method for determining total and HDL cholesterol levels in plasma. It involves using cholesterol esterase and cholesterol oxidase to convert cholesterol to a compound that reacts with other reagents to form a colored complex, which is then quantified spectrophotometrically. Specifically, it precipitates LDL and VLDL from plasma to obtain the HDL fraction, and measures total cholesterol without precipitation to determine overall levels. The absorbance readings are used to calculate cholesterol concentrations based on a standard reference value.

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    15 views9 pages

    Determination of Plasma Total-And Hdl-Choleterol

    Uploaded by

    趙偉光
    This document describes a method for determining total and HDL cholesterol levels in plasma. It involves using cholesterol esterase and cholesterol oxidase to convert cholesterol to a compound that reacts with other reagents to form a colored complex, which is then quantified spectrophotometrically. Specifically, it precipitates LDL and VLDL from plasma to obtain the HDL fraction, and measures total cholesterol without precipitation to determine overall levels. The absorbance readings are used to calculate cholesterol concentrations based on a standard reference value.

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    © All Rights Reserved
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    of 9

    DETERMINATION OF PLASMA

    TOTAL-AND HDL-CHOLETEROL
    Introduction

    free cholesterol
    Plasma cholesterol
    cholesterol ester (70%~80%)

    cholesterol esterase （胆固醇酯酶）

    Cholesterol ester + H2O free cholesterol + fatty acid


    （ hydrolysis ）
    cholesterol oxidase （胆固醇氧化酶）

    Free cholesterol + O2 ∆ 4-cholestenone + H2O2


    （ ∆ 4- 胆甾烯酮）

    peroxidase
    2H2O2 + 4-aminoantipyrine + phenol red compound + 4H2O
    （ 4- 氨基安替比林）

    Quantified by spectrophotometer
    Principle
    HDL Magnesium
    phosphotungstate
    LDL HDL
    VLDL
    +
    LDL
    VLDL

    HDL
    Material and method

    Tube H: 0.2ml plasma+ 0.2ml precipitant


    Tube SH: 0.2ml standard cholesterol+0.2ml precipitant

    Stand for 15 min

    Centrifuge at 2000rpm.10min

    Standard cholesterol (50mg/dl)


    Material and method

    unit ml
    Tube NO. T ST H SH B
    Supernatant -- -- 0.2 0.2 --
    Plasma 0.02 -- -- -- --
    Cholesterol standard -- 0.1 -- -- --
    Distilled water 0.18 0.1 -- -- 0.2
    Enzyme reagent 2.0 2.0 2.0 2.0 2.0
    Material and method

    Mix

    Incubate at 37℃.20min

    Read the absorbance at 520nm


    Calculation

    O.D.T 0.1
    Total cholesterol (mg/dl)= * * 50
    O.D.ST 0.02

    O.D.H
    HDL-cholesterol (mg/dl)= * 50
    O.D.SH
    Standard cholesterol solution (50mg/dl)

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