Cholesterol FS*
Diagnostic reagent for quantitative in vitro determination of cholesterol in serum or
plasma on photometric systems

Order Information Storage Instructions and Reagent Stability

Cat. No. Kit size The reagent is stable up to the end of the indicated month
1 1350 99 10 021 R 5x 25 mL + 1 x 3 mL Standard of expiry, if stored at 2 – 8 °C, protected from light and
1 1350 99 10 026 R 6x 100 mL contamination is avoided. Do not freeze the reagents!
1 1350 99 10 023 R 1x 1000 mL The standard is stable up to the end of the indicated month
1 1300 99 10 030 6x 3 mL Standard of expiry, if stored at 2 – 25 °C.
Summary [1,2] Note: It has to be mentioned, that the measurement is not
influenced by occasionally occurring color changes, as long
Cholesterol is a component of cell membranes and a
as the absorbance of the reagent is < 0.3 at 546 nm.
precursor for steroid hormones and bile acids synthesized
by body cells and absorbed with food. Cholesterol is Warnings and Precautions
transported in plasma via lipoproteins, namely complexes 1. The reagent contains sodium azide (0.95 g/L) as
between lipids and apolipoproteins. There are four classes preservative. Do not swallow! Avoid contact with skin
of lipoproteins: high density lipoproteins (HDL), low density and mucous membranes.
lipoproteins (LDL), very low density lipoproteins (VLDL) 2. Standard is harmful. R22: Harmful if swallowed. R43:
and chylomicrons. While LDL is involved in the cholesterol May cause sensitisation by skin contact. S2: Keep out
transport to the peripheral cells, HDL is responsible for the of the reach of children. S13: Keep away from food,
cholesterol uptake from the cells. The four different drink and animal feedingstuffs. S24: Avoid contact
lipoprotein classes show distinct relationship to coronary with skin. S37: Wear suitable gloves. S46: If
atherosclerosis. LDL-cholesterol (LDL-C) contributes to swallowed, seek medical advice immediately and show
atherosclerotic plaque formation within the arterial intima this container or label. S64: If swallowed, rinse mouth
and is strongly associated with coronary heart disease with water (only if the person is conscious).
(CHD) and related mortality. Even with total cholesterol 3. In very rare cases, samples of patients with
within the normal range an increased concentration of gammopathy might give falsified results.
LDL-C indicates high risk. HDL-C has a protective effect 4. Please refer to the safety data sheets and take the
impeding plaque formation and shows an inverse necessary precautions for the use of laboratory
relationship to CHD prevalence. In fact, low HDL-C values reagents. For diagnostic purposes, the results should
constitute an independent risk factor. The determination of always be assessed with the patient’s medical history,
the individual total cholesterol (TC) level is used for clinical examinations and other findings.
screening purposes while for a better risk assessment it is
Waste Management
necessary to measure additionally HDL-C and LDL-C.
In the last few years several controlled clinical trials using Please refer to local legal requirements.
diet, life style changes and/or different drugs (especially Reagent Preparation
HMG CoA reductase inhibitors [statins]) have The reagent and the standard are ready to use.
demonstrated that lowering total cholesterol and LDL-C
levels reduce drastically CHD risk [2]. Materials required but not provided
NaCl solution 9 g/L
Method General laboratory equipment
“CHOD-PAP”: enzymatic photometric test
Specimen
Principle Serum, heparin plasma or EDTA plasma
Determination of cholesterol after enzymatic hydrolysis and Stability [6]: 7 days at 20 – 25 °C
oxidation [3,4]. The colorimetric indicator is quinoneimine 7 days at 4 – 8 °C
which is generated from 4-aminoantipyrine and phenol by 3 months at -20 °C
hydrogen peroxide under the catalytic action of peroxidase Discard contaminated specimens! Freeze only once!
(Trinder’s reaction) [3].
Assay Procedure
Cholesterol ester + H2 O CHE > Cholesterol + Fatty acid
Application sheets for automated systems are
Cholesterol + O2 CHO > Cholesterol-3-one + H2O2 available on request.
2 H2O2 + 4-Aminoantipyrine + Phenol POD > Quinoneimine + 4 H2O
Wavelength 500 nm, Hg 546 nm
Optical path 1 cm
Temperature 20 - 25 °C/37 °C
Reagents Measurement Against reagent blank
Components and Concentrations
Blank Sample or standard
Reagent: Sample or standard - 10 µL
Good's buffer pH 6.7 50 mmol/L Dist. water 10 µL -
Phenol 5 mmol/L Reagent 1000 µL 1000 µL
4-Aminoantipyrine 0.3 mmol/L Mix, incubate for 10 min. at 20 – 25 °C or for 5 min. at
Cholesterol esterase (CHE) 200 U/L 37 °C. Read absorbance within 60 min against reagent
Cholesterol oxidase (CHO) 100 U/L blank.
Peroxidase (POD) 3 kU/L
Standard: 200 mg/dL (5.2 mmol/L)

Cholesterol FS – Page 1 * fluid stable

  

Calculation Reference Range [5]

With standard or calibrator Desirable " 200 mg/dL (5.2 mmol/L)
Borderline high risk 200 – 240 mg/dL (5.2 – 6.2 mmol/L)
A Sample
Cholester o l [mg / dL] ! x Conc. Std / Cal [mg / dL] High risk > 240 mg/dL (> 6.2 mmol/L)
A Std / Cal
Each laboratory should check if the reference ranges are
Conversion factor
transferable to its own patient population and determine
Cholesterol [mg/dL] x 0.02586 = Cholesterol [mmol/L] own reference ranges if necessary.
Calibrators and Controls Clinical Interpretation
For the calibration of automated photometric systems the The European Task Force on Coronary Prevention
DiaSys TruCal U calibrator is recommended. The assigned recommends to lower TC concentration to less than
values of the calibrator have been made traceable to the 190 mg/dL (5.0 mmol/L) and LDL-cholesterol to less than
reference method gas chromatography-isotope dilution 115 mg/dL (3.0 mmol/L) [2].
mass spectrometry (GC-IDMS). For internal quality control
DiaSys TruLab N and P or TruLab L controls should be Literature
assayed. Each laboratory should establish corrective action 1. Rifai N, Bachorik PS, Albers JJ. Lipids, lipoproteins and
in case of deviations in control recovery. apolipoproteins. In: Burtis CA, Ashwood ER, editors.
Cat. No. Kit size Tietz Textbook of Clinical Chemistry. 3rd ed.
TruCal U 5 9100 99 10 063 20 x 3 mL Philadelphia: W.B Saunders Company; 1999. p. 809-
5 9100 99 10 064 6 x 3 mL 61.
TruLab N 5 9000 99 10 062 20 x 5 mL
2. Recommendation of the Second Joint Task Force of
5 9000 99 10 061 6 x 5 mL
TruLab P 5 9050 99 10 062 20 x 5 mL
European and other Societies on Coronary Prevention.
5 9050 99 10 061 6 x 5 mL Prevention of coronary heart disease in clinical
TruLab L Level 1 5 9020 99 10 065 3 x 3 mL practice. Eur Heart J 1998;19: 1434-503.
TruLab L Level 2 5 9030 99 10 065 3 x 3 mL 3. Artiss JD, Zak B. Measurement of cholesterol
concentration. In: Rifai N, Warnick GR, Dominiczak
Performance Characteristics MH, eds. Handbook of lipoprotein testing. Washington:
AACC Press, 1997:99-114.
Measuring range 4. Deeg R, Ziegenhorn J. Kinetic enzymatic method for
The test has been developed to determine cholesterol automated determination of total cholesterol in serum.
concentrations within a measuring range from Clin Chem 1983;29:1798-802.
3 - 750 mg/dL (0.08 – 19.4 mmol/L). When values exceed 5. Schaefer EJ, McNamara J. Overview of the diagnosis
this range samples should be diluted 1 + 4 with and treatment of lipid disorders. In: Rifai N, Warnick
NaCl solution (9 g/L) and the result multiplied by 5. GR, Dominiczak MH, eds. Handbook of lipoprotein
testing. Washington: AACC press, 1997:25-48.
Specificity/Interferences
6. Guder WG, Zawta B et al. The Quality of Diagnostic
No interference was observed by ascorbic acid up to Samples. 1st ed. Darmstadt: GIT Verlag; 2001.
5 mg/dL, bilirubin up to 20 mg/dL, hemoglobin up to p. 22-3.
200 mg/dL and lipemia up to 2,000 mg/dL triglycerides. 7. Young DS. Effects of Drugs on Clinical Laboratory
For further information on interfering substances refer to Tests. 5th ed. Volume 1 and 2. Washington, DC: The
Young DS [7]. American Association for Clinical Chemistry Press
Sensitivity/Limit of Detection 2000.
The lower limit of detection is 3 mg/dL (0.08 mmol/L).
Manufacturer
Precision (at 37°C) DiaSys Diagnostic Systems GmbH
Intra-assay precision Mean SD CV IVD Alte Strasse 9 65558 Holzheim Germany
n = 20 [mg/dL] [mg/dL] [%]
Sample 1 108 1.76 1.62
Sample 2 236 1.45 0.61
Sample 3 254 1.57 0.62

Inter-assay precision Mean SD CV

n = 20 [mg/dL] [mg/dL] [%]
Sample 1 104 1.19 1.14
Sample 2 211 2.57 1.22
Sample 3 245 2.28 0.93
Method Comparison
A comparison of DiaSys Cholesterol FS (y) with a
commercially available test (x) using 78 samples gave
following results: y = 1.00 x – 2.50 mg/dL; r= 0.995.

Cholesterol FS – Page 2 844 1350 10 02 00 November 2012/7