UMA CO., LTD.

MEASURE HDL
2-19-6 Yokosuka Reagent for measuring HDL Cholesterol
Matsudo, Chiba, Japan Inhibition/Direct

á 2 ~ 8 °C IVD In vitro Diagnostics Packages

R1 1 ´ 90 mL R2 1 ´ 30 mL
T DO NOT freeze 6 18 months/block from light R1 1 ´ 60 mL R2 1 ´ 20 mL

1. PURPOSE OF USE 6. MEASUREMENT PRINCIPLE

In vitro Determination of HDL cholesterol in serum or This method enables determination of HDL Cholesterol
plasma. only in a selective manner by combination of surfactant,
inorganic and organic phosphorus compounds which
2. GENERAL INSTRUCTION
inhibits reaction against LDL, VLDL and chylomicron.
1. For in vitro diagnostics use only.
st
In the 1 reaction, LDL, VDL and chylomicron except for
2. Diagnosis should be made in a comprehensive manner,
in accordance with other related test results and clinical HDL-Cholesterol are bound to surfactant, inorganic and
nd
organic phosphorous compounds. In the 2 reaction, only
symptoms by the doctor in attendance.
HDL-Cholesterol can be determined by addition of CHER
3. For guaranteed results, usage of this product must
and CHOD without doing fractionation.
comply with the instruction in this manual.
"#$%,/()*+,-.,/.,
4. For automatic analyzers: follow their instructions. 0*1,/2-/ 2/3*1,/2- -3453)/6(
HDL Cholesterol + H2O
3. MATERIALS REQUIRED BUT NOT INCLUDED
Cholesterol + fatty acid
- Saline 0.9 % and high grade purified water "#07
Cholesterol + O2 Cholestenon + H2O2
- Micropipet and other basic laboratory equipment. + 807
2 H2O2 + 4-AA + ADPS + H3O
- Calibrators and Controls (separatedly sold)
blue-violet quinone pigment + 5H2O
4. REAGENT COMPOSITION & PREPARATION (λmax = 583 nm)
- Reagent R-1: HDAOS; ascorbate oxidase, H3PO4.
7. STANDARD MEASUREMENT OPERATION
Reagent R-1 is ready for use
Specimen Calibrator Blank
- Reagent R-2: CHOD, Cholesterol Esterase (CHER),
(S) (Std) (B)
Peroxidase (POD), 4-aminoantipyrine (4-A-A).
Specimen (µL) 20 - -
Reagent R-2 is ready for use
Calibrator (µL) - 20 -
- Calibrators (separatedly sold): Ready for use
Saline (µL) - - 20
- Controls Lyo-1 & Lyo-2 (separatedly sold): Put 1 mL of
R-1 (µL) 1500 1500 1500
purified water to the vials of calibrator and controls (L, H);
Incubate at 37 °C in 5 minutes
leave at room temparature for 30 minutes before use. After
R-2 (µL) 500 500 500
reconstituted, calibrators/control can be use without dilution.
Mix well; incubate at 37 °C for 5 minutes; measure
5. SAMPLE PREPARATION & STORAGE absorbance at 600 nm
- Serum: Collect blood after 12 ~ 14 hours fasting. Wait Note: See sample preparation for details of specimen
until sample completly coagulated. Take the supernatant to
use as specimen. 8. CALCULATION & UNIT CONVERSION

- Plasma: Collect blood after 12 ~ 14 hours fasting. Treat Calculation

sample by anticoagulant (i.e heparin lithium); leave sample - Calculate ∆Abs of specimen & standards vs blank

to stand for 3 hours or centrifuge at 2000 rpm for 2 minutes; - Plot a calibration curve HDL (mg/dL) = f(∆Abs)

take the plasma layer (supernatant) and use as specimen. - Calculate HDL concentration in specimen using the

- Analyze sample soon after collection. In case, it could curve (doing same procedure for Controls)

not be analyzed soon, store sample 2 ~ 8 °C and analyze Unit conversion

within 4 days. 1 mg/dL = 0.026 mmol/L

1/2 Revision 06/2013

9. PERFORMANCE & CORRELATION TEST
Performance
- Sensitivity: Change in absorbance when using purified
water is 0.001 ~ 0.015, and change in absorbance using
standard solution (cholesterol 50 mg/dL) as specimen is
0.05 ~ 0.15.
- Specificity: The accuracy is within ±10.0%.
- Reproducibility: CV value < 3.0%.
- Measuring range: 5 ~ 120 mg/dL.
Correlation test
Same measuring principle
- Serum:
Regression equation: Y = 1.034X－2.70 (n = 100)
Correlation coefficient: R = 0.999
(Y: Value obtained from using this method)
Reference Materials for Calibration
- JCCRM 224
10. EXPECTED VALUES
Normal reference range
- 40 - 70 mg/dL;
Reference range should be established at each facility and
judgement should base on measurement results in a
comprehensive manner together with clinical symptoms
and other measurement results.
11. INTERFERENCES
- Ascorbic acid up to 50 mg/dL: within ±3%
- Conjugated bilirubin upto 20 mg/dL: within ±3%
- Hemoglobin 500 mg/dL: within ± 1%
- Formazin turbidity upto 1600 unit: within ± 1%.
- Abnormal lipoprotein contained sample may affect to the
determination result.
12. INFORMATION FOR AUTOANALYZERS
Calculation Method 2-point (fix)
13. HANDLING, USAGE & DISPOSAL
Handling
1. Specimen can be potentially positive for infectious agents
including hepatitis B virus and HIV. Wear glove and goggle
when needed.2. In case reagents got into skin, eye or
mouth by mistake, wash it immediately with plenty of water
and consult the doctor if needed.
3. If reagents are spilled, dilute with water and wipe it out. If
specimen is spilled, spray 80% of alcohol over the
specimen and wipe it out.
Usage
1. Store reagents under specified condition. Do not use
after expiration date.
2. Do not use the container and auxiliaries included in this
kit for other purposes.
3. Do not mix reagents of different lot for use.
4. Do not add to the reagent being used even if it is the
same lot number.
Disposal
1. All specimens, as well as all instruments (e.g. test tubes)
that come in contact with the specimens, must be treated by
the following methods, or they must be treated according to
the manual for infectious medical waste provided in each
facility.
・ Sterilize with an autoclave, subjecting them to high
pressure saturated steam at 121 °C for more than 20
minutes. Do not process waste containing sodium
hypochlorite solution with an autoclave.
・Immerse at least one hour in sodium hypochlorite solution
(active chloride concentration of over 1000 ppm).
2. This reagent contains sodium azide. Sodium azide can
react with lead pipe and/or steel pipe and can generate
explosive metal azide. Make sure to use plenty of water at
disposal. Concentration of sodium azide in R-2 is 0.05%.

Temperature 37 °C 14. OTHER INSTRUCTIONS AND CAUTION

Specimen 2.0 - Results may differ depending on the sample/reagent
Volume (μL) R1 150 ratio. Adjust parameters for different analyzer.
R2 50 - Prepare the calibration curve on the day of
Main 600 determination.
Wavelength (nm)
Sub- 800
Point 1 10
Measurement
Point 2 16
(cycle) Point 3 34

Calibration type Linear

Unit mg/dL

2/2 Revision 06/2013