Cholesterol

CHOD/POD, Liquid 1x25 mL, 5x25 mL

Quantitative determination of Cholesterol by means d. Interfering Substances

of photometric method. - Ordinary anticoagulants except EDTA and heparin
Only for in vitro use in clinical laboratory (IVD) interfere with the test
- Turbidity may be observed in strongly hyperlypemic
Clinical Significance samples. In theis case dilue final reaction 1:2 or 1:3
Cholesterol is a fat like substance that is found in all body with reagent, repeat readings and multiply the dilution
cell. The liver makes all the cholesterol the body needs to ratio.
form cell membranes and to make certain hormones. - no interference has been observed by the following
The determination of Serum Cholesterol is one of the bilirubin upto 10 mg/dL. Ascorbic acid upto 7.5 mg/dL,
important tools in the diagnosis and classification of lipemia. uric acid upto 20 mg/dL, hemoglobin upto 200 mg/dL
High blood cholesterol is one of the major risk factor of - For comprehensive list of interfering substances see
heart disease5.6. Young et. al
Clinical Diagnostic should not be made on a single test
result. It should integrate clinical and other laboratory data. Assay Procedure
1. Assay conditions:
Principle Wavelength: . . . . . . . . . . . .. . . . . . . . . . . 505 nm
Cholesterol esters in Serum are hydrolysed by cholesterol Cuvette: . . … . . . . . . . . . . . . . . . . ... 1 cm light path
esterase (CHE). The free cholesterol produced is oxidized by Constant temperature . . . . . . . . . . . . . . . . . . . 37ºC
cholesterol oxidase (Co) to form Cholest 4en- 3-one with
simultaneous production of hydrogen peroxide (H2O2) In three colorimetric test tubes labeled B (Blank), S
(Standard) and U (Unknown Sample) place
which oxidatively couples with 4-aminoantipyrine and
phenol in the presence of peroxidase (POD) to yield a red Unknown
chromophore. Blank (B) Standard (S)
Samples (U)
CHE
Cholesterol ester + H2O Cholesterol + Fatty acid
Reagent 1000 µL 1000 µL 1000 µL
Cholesterol + O2 CHOD Cholesterol-3-one + H2O2
Distilled Water 10 µL -- --
POD
H2O2 + 4-AAP + Phenol Red Quinoneimine + H2O
Standard -- 10 µL --
The red Quinoneimine dye formed is measured
Sample -- -- 10 µL
spectrophotometrically at 505 nm as an increase in
absorbance. 2. Mix thoroughly.
3. Incubate for 5 minutes at 37 °C or 15 min. at Room
Reagents Temperature (15 -25°C).
Each Cholesterol kit contains 4. Read in Spectrophotometer at 505 nm or in photo
colorimeter with green filter (490-530nm) setting
R: Enzyme Reagent instrument to zero with reagent blank.
Cholesterol Standard: 200mg/dL
Calculation
Reagent Preparation (A) Sample
Reagent 1 is ready for use. Cholestero l (mg/dl) = X 200 (Standard Conc.)
(A) Standard
Stability Application sheets for automated systems are
All the components of the kit are stable until the expiration available on request.
date on the label when stored at 2–8 °C, protected from
light and contamination prevented during their use. Do not QUALITY CONTROL
use reagent over the expiry date. Reagent may develop a Commercially available normal and pathological controls
slight pink color which does not affect results provided the sera are recommended to monitor the performance of the
blank is run with every batch of tests and a standard procedure.
included periodically. If control values are found outside the defined range, check
Cholesterol STD: Stable up to the expiry when stored the instrument, reagent and standard for problems
tightly closed at 2–8 °C. Protected from light and Serum controls are recommended for internal Quality
contamination prevented during their use. control. Each laboratory should establish its own Quality
Signs of reagent deterioration Control scheme and corrective actions
- presence of particles and turbidity
- Blank Absorbance (A) at 505 nm > 0.160 Reference Range
- Standard reading is abnormally low. Risk Evaluation:

Specimen and Stability Less than 200 mg/dL Normal/Recommended

Serum or plasma 200 – 240 mg/dl Borderline
a) Collection: Obtain Serum or Plasma in the Usual way >240 mg/dL High Risk
b) Additives use only Heparin as anticoagulant to obtain (These values are for orientation purpose)
Plasma.
c) Stability of the sample for 7 days at 2 - 8°C or freezing
These values are for orientation purpose
at - 20°C will keep the sample for few months.
Each laboratory should establish its own reference range
Reagent Performance.
Measuring range
Reaction is linear up to 750 mg/dl under the described
assay conditions.
If the results obtained were greater than linearity limit,
dilute the sample 1:2 with NaCl 9 g/L (Normal Saline) and
multiply the result by 2

SYSTEM PARAMETERS

Mode : End Point

Reaction : Ascending
Wavelength : 505 nm
Blank with : Reagent
Sample Volume : 10 µL
Reagent Volume : 1000 µL
Incubation : 5 min/ 10 min
Standard : 200
Linearity limit : 750 mg/dL
Unit : mg/dL

Notes

1. Use clean disposable pipette tips for dispensation

Only for invitro use in Clinical laboratory (IVD)

Literature
1. Trinder P. Ann. Clin. Biochem, 6: 24 (1969)
2. Lieberman. C. Ber. 19-8. 1803 (1885)
3. Young D.S. et al. Clin Chem 21:1D (1975)
4. Castell W.P. Current Prescribing 6/77 (1977)
5. Flegg A. S. Ann Clin Biochem 10:79 (19730)
6. I.F.C.C.- Clin CheM Acta 87/3 : 459F(1978)
7. Friedwald W.T. et al Clin Chem 18: 498 (1972)

Manufactured by: Prism Diagnostics Pvt. Ltd.

Marketed By: Euro Diagnostic Systems Pvt. Ltd.,
Millennium House, M.K.Srinivasan Nagar Main Road,
No. 144, Old Mahabalipuram Road, Perungudi, Chennai –
600 096; Email: eurods@vsnl.net, www.eurods.in