Edta is the traditional choice bc the anticoagulation

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EDTA is the traditional choice b/c the anticoagulation was thought to enhance stability by chelating metal ions; but, it has potential disadvantages; microclots can form in plasma after storage and could plug the sampling probes on modern chemistry analyzers; EDTA also osmotically draws water from red cells, diluting the plasma constituents, and the dilution effect can vary depending on such factors as fill volumes, the analyte being measured, and the extent of mixing. B/c NCEP cutpoints are based on serum values, cholesterol measurements made on EDTA plasma require correction by the factor of 1.03 6. Describe the principles, reagents, reactions and sources of error involved in the following methods: Cholesterol-Lieberman-Burchard; enzymatic; triglycerides- enzymatic; HDL cholesterol-separation by precipitation of LDL and VLDL and direct measurement (LAB STUDENTS ONLY). Dietetics students: List methods associated with cholesterol, triglycerides, HDL, LDL and VLDL. LIEBERMAN-BURCHARD---cholesterol measurement; a reagent is used to create a color formation; uses hexane extraction after hydrolysis w/ alcoholic KOH followed by reagent, w/ Lieberman-Burchard color reagent which comprises sulfuric and acetic acids and acetic anhydride
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Lipids and Lipoproteins ENZYMATIC---cholesterol measurement; selected for specifity to the analyte of interest; most common sequence---the enzyme cholesteryl ester hydrolase cleaves the fatty acid residue from cholesteryl esters, which comprises about 2/3 of circulating cholesterol, converting them to unesterified or free cholesterol. The free cholesterol is reacted by the 2 nd enzyme cholesterol oxidase, producing hydrogen peroxidase, a substrate for common enzymatic color reaction using horseradish peroxidase to couple 2 colorless chemicals into a colored compound; the intensity of the resulting color is proportional to the amount of cholesterol and can be measured by a spectrophotometer ENZYMATIC---triglyceride measurement; lipases cleave fatty acids from glycerol backbone, the freed glycerol participates in any one of several enzymatic sequences HDL AND LDL CHOLESTEROL – SEPERATION BY PRECIPITATION OF LDE AND VLDL AND DIRECT MEASUREMNT---precipitaion: add reagent---HDL/LDL precipitates---measure supernant; Inc. VLDL and chylomicrons interefere----false elevations; direct measurement: reagent added blocks non HDL/LDL lipoproteins---second reagent---enzymatic reation----quantify LDL/HDL 7. List NCEP values for cholesterol, HDL cholesterol, LDL cholesterol, and triglycerides. Borderline high triglycerides: 150-200 mg/dl; high: 200-500; very high: >500 Adult Reference Ranges for Lipids Total Cholesterol: 140-200 HDL Cholesterol: 40-75 LDL Cholesterol: 50-130 Triglyceride: 60-150
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