HDL-CHOLESTEROL DIRECT

PRINCIPLE

DIRECT
Enzymatic colorimetric method
FIXED TIME

The assay is based on a modified polyvinyl sulfonic acid (PVS) and polyethylene-glycol-methyl ether (PEGME) coupled classic precipitation method with the improvements in using optimized  quantities of PVS/PEGME and selected detergents.

LDL, VLDL, and chylomicron (CM) react with PVS and PEGME and the reaction results in inaccessibility of LDL, VLDL and CM by cholesterol  oxidase (CHOD) and cholesterol esterase (CHER). The enzymes selectively react with HDL to produce H2O2 which is detected through a Trinder reaction.

REAGENT COMPOSITION

40 ml
R1 Reagent 1. MES buffer (pH 6.5), TODB N,N-Bis(4-
sulfobutyl)-3-methylaniline, polyvinyl sulfonic acid,
polyethylene-glycol-methyl ether, MgCl2, detergent,
EDTA. 1 x 30 ml.
R2 Reagent 2: MES buffer (pH 6.5), cholesterol esterase,
cholesterol oxidase, peroxidase, 4-aminoantipyrine,
detergent. 1 x 10 ml.
CAL LDL/HDLc calibrator:  Optative. Ref. 1972005. Concentration
value is traceable to NIST SRM 1951b.

STORAGE AND STABILITY
Store at 2-8ºC

REAGENT PREPARATION
Reagents R1 and R2 are ready to use. Stability open on board the
analyzer at 2-8ºC is of 2 months.
LDL/HDLc calibrator. Lyophilized. Reconstitute contents with
distilled water per instructions on vials. Mix gently and let stand
for 5 minutes before use. The reconstituted material is stable for
7 days at 2-8ºC or for 1 month at –20ºC. Discard if it becomes
turbid or if there is any evidence of microbial contamination.

SAMPLES
Serum, EDTA or heparinized plasma obtained by the patient after
an overnight fast. Remove from cells within 3 hours of
venipuncture. Samples may be kept at 4-8ºC for 2 weeks or at
–20ºC for 3 months.

MATERIALS REQUIRED

• Photometer or spectrophotometer with a thermostatic cell compartment set at 37ºC, capable of reading at 600 +/- 10 nm.
• Stopwatch, strip-chart recorder or printer.
• Cuvettes with 1-cm pathlength.
• Pipettes to measure reagent and samples

HDL-CHOLESTEROL DIRECT Reagents per box