CKMB Creatine kinase isoenzyme (CKMB) assay kit (Enzymatic method)

In vitro test for the quantitative determination of creatine kinase-MB (CK-MB) activity in serum on photometric systems. Creatine kinase (CK) is an enzyme, which consists of isoenzymes mainly of the muscle (CK-M) and the brain (CK-B). CK exists in serum in dimeric form as CK-MM, CK-MB, and CK-BB and as macroenzyme. Determination of CK-MB values possesses highly specificity in myocardial damages. So, measurement of CK-MB is used for diagnosis and monitoring of myocardial infarction.

Storage

Up to expiration date indicated on the label, when stored unopened at 2-8℃ and protected from light. Once opened, the reagents are stable for 28 days when refrigerated on the analyzer or refrigerator. Contamination of the reagents must be avoided. Do not freeze the reagents. Once dissolved, the calibrator are stable for 7 days at 2–8℃, the control are stable for 7 days at 2–8℃，do not freeze.

Principle

In the reaction, the creatine kinase-B catalyzes the transfer of a phosphate group from the creatine phosphate substrate to ADP. The subsequent formation of ATP is measured through the use of two coupled reactions catalyzed by hexokinase (HK) and glucose-6-phosphate dehydrogenase (G-6-PDH) which results in the production of the reduced form of NADPH from NADP. The rate of absorbency increase at 405nm is directly proportional to the activity of creatine kinase-B.

Reagents

Sample requirements

1. Serum, heparin or EDTA plasma, and urine are suitable for samples. Whole blood, hemolysis is not recommended for use as a sample. Freshly drawn serum is the preferred specimen.

2. Stability: Serum/plasma: 1 week at 2-8℃;

Calibrator preparation

Carefully open the bottle, avoiding the loss of lyophilizate, and pipette in exactly 1.0 mL of distilled/deionized water. Carefully close the bottle and dissolve the contents completely by occasional gentle swirling within 30 minutes. Avoid the formation of foam. The dissolved calibrator can be used without any other Pretreatment.

Method

1. Reagent preparation：Liquid reagent can be used when opened

2. Measurement：

ΔA/min = [ΔA/min sample]- [ΔA/min blank]

Quality control preparation

Calibration

It is recommended to use the Calibrator from Hyasens and distilled/deionized water for two-point calibration.

Calibration frequency: After reagent lot changed.

As required following quality control procedures.

Quality control

At least two levels of control material should be analyzed with each batch of samples.Each laboratory

should establish its own internal quality control scheme and procedures for corrective action if controls do not recover within the acceptable tolerances.

Reference intervals

Each laboratory should establish its own reference intervals based upon its patient population. The reference intervals measured at 37 °C listed below were taken from literature. 

Serum / Plasma: ≤ 24 U/L

Interferences/specificity

The following substances were tested for interference with this methodology. Criterion: Recovery within ±10 % of initial value.

Reagent blank absorbance

The absorbance of reagent blank at 340 nm should be ≤0.6 A.

Sensitivity/detection limit

The lowest measurable CK activity that can be distinguished from zero is 10 U/L with 99.7% confidence.

Precision 

Within-run: CV≤5%

Between-run: CV≤10%

Linearity range

Conventional Units: 10-600 U/L

If the value of sample exceeds 600 U/L, the sample should be diluted with 9 g/L NaCl solution (e.g. 1+ 9) and rerun; the result should be multiplied by 10.

Warnings and precautions

1. For in vitro diagnostic use.

2. Take the necessary precautions for the use of laboratory reagents.

3. Preservative contained. Do not swallow. Avoid contact with skin and mucous membranes.

4. Disposal of all waste material should be in accordance with local guidelines.

5. Material safety data sheet is available on request for professional users.

References

[1] Tietz Textbook of Clinical Chemistry, 3^rd edition. Burtis CA, Ashwood ER. WB Saunders Co., 1999.

[2] Gerhardt, W., Ljungdahl, L., Borjesson, J., Hovendahl, S., Hedenas, B., Clin. Chem. Acta, 78:29 (1979).

[3] National Committee for Clinical Laboratory Standards, How to Define, Determine, and Utilize Reference Intervals in the Clinical Laboratory, Approved Guideline, CLSI publication C28-A,Villanova, PA (1995).