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www.biomaghreb.com
IVD
FT En 23 Date of updating : 10/ 2020 Version B
IN VITRO DIAGNOSTIC USE
Manufacturer Use by In Vitro Diagnostic Temperature
Limitation
Catalogue number See insert Store away from light Sufcient
for < n > essais
Batch number
CLINICAL SIGNIFICANCE
High cholesterol is considered a risk factor for atherosclerosis. It can be primary or secondary to
thyroid insufficiency, pancreatitis, or nephrotic syndrome.... It will be necessary to type this anomaly
by determining the respective levels of the different lipid fractions (HDL, LDL, VLDL).
A high level of HDL cholesterol protects against coronary heart disease complications. It can increase
with age, gender, physical activity, a diet low in cholesterol and high in polyunsaturated fatty acids, or
when taking certain drugs such as lipid-lowering drugs (fibrates), vitamin C, anti-epileptics, insulin
and estrogen-progestin or alcohol consumption.
PRINCIPLE
Chylomicrons and very low density (VLDL) and low density (LDL) lipoproteins contained in the
sample are precipitated by addition of phosphotungstic acid in the presence of magnesium ions.
The supernatant obtained after centrifugation contains the high density lipoproteins (HDL) whose
cholesterol is determined by the cholesterol enzyme reagent of the complementary kit.
REAGENT COMPOSITION
Precipitant
Reagent
Phosphotungstic Acid
MgCl2 6H2O pH 6,2
13,9 mmol/l
490 mmol/l
SAFETY CAUTIONS
Biomaghreb reagents are intended for use by qualified personnel for in vitro use (do not pipette by
mouth).
• Refer to the current SDS available on request or at www.biomaghreb.com;
• Verify the integrity of the reagents before use; and
• Disposal of waste: comply with the legislation in force.
For safety reasons, treat any specimen or reagent of biological origin as potentially infectious.
Respect the legislation in force.
REAGENT PREPARATION
The reagent is ready to use.
SAMPLE PREPARATION
Serum or plasma collected on EDTA.
PRESERVATION AND STABILITY
The reagent must be stored until the expiry date indicated on the package label between 2-8°C.
LIMITS
If the reagent is cloudy or colored, it must be discarded.
ADDITIONAL EQUIPMENT
• Basic equipment of the medical analysis laboratory ;
• Spectrophotometer or Clinical Biochemistry Analyzer.
QUALITY CONTROL
External quality control program.
It is recommended to control in the following cases:
• At least one test per series.
• Change of reagent bottle.
• After maintenance work on the analyzer.
If a control value is outside the confidence limits, repeat the procedure using the same control.
Use normal and pathological control sera.
PROCEDURE
• Precipitation
Dilute in 9 g/l NaCl solution the sera with triglyceride more than 3,5 mmol/l.
Serum...........................................................................500 μl
Precipitant reagent .......................................................50 μl
Mix well, wait 10 minutes;
Centrifuge for 15 minutes at 5000 rpm.
• Cholestérol-HDL (CHOD-PAP) Assay:
Reconstitute the enzymatic cholesterol reagent according to the notice of the additionnal kit.
Wavelength: 505 nm (492-550);
Temperature: 37°C;
Adjust the spectrophotometer zero on the reagent blank.
Blank Standard Assay
Distilled water
10 µl - - - -
Standard Cholesterol
- - 10 µl - -
Supernatant
- - - - 10 µl
Cholesterol enzyme Reagent
1 ml 1 ml 1 ml
Mix, read absorbances after incubation for 5 minutes at 37°C. Staining is stable 30
minutes.
N.B : Once the vial of HDL reagent is opened, there is a possibility of the formation of shiny crystals
at the bottom of the vial which do not affect the quality of the product.
CALCULATION
[HDL-Cholesterol] =
DO. Assay
DO. Standard
x n
with n = Standard value
n = 5,17 mmol/l ;
n = 2 g/l.
Multiply the result obtained by 1.1 to take into account the dilution made during precipitation: this
gives the HDL cholesterol concentration .
REFERENCE VALUES
Cholesterolemia Evaluation du risque
< 2g/l
< 5,2 mmol/l
Low risk
2,0 à 2,5 g/l
5,2 à 6,5 mmol/l
Moderate risk especially if
HDL cholesterol < 0,35 g/l ;
< 0,9 mmol/l.
> 2,5 g/l
> 6,5 mmol/l
High risk especially if
HDL cholestErol < 0,35 g/l ;
< 0,9 mmol/l.
REFERENCES
BURSTEIN M. et al. Lipid Res.11. 583.(1970) ;
Study Group, European Atherosclerosis Society, European Heart Journal, 9, 571 (1988) ;
ARCOL, ISB, 1989, 15, 121 -124.
CHOLESTEROL-HDL
Precipitation method
Reagent for the quantitative determination of
HDL-cholesterol in human plasma
