Experimental instrument
Test tube, Micropipettor, Vortex mixer, 37℃ incubator, Spectrophotometer (340nm)
Application
This kit can measure the activity of LDH isoenzymes in serum or plasma samples.
Detection principle
There is guanidine thiocyanate as an inhibitor to suppress the activity of LDH2, LDH3, LDH4 and LDH5, so this kit can be used to test the activity of LDH1. Measure the formation rate of NADH at 340nm wavelength and then the activity of LDH1 can be calcμLated.
Kit components
Item | Composition | Specification |
Reagent 1 | N-acetyl-D-glucamine solution(pH9.3) | 40 mL ×1vial |
Guanidine thiocyanate | ||
Lithium L-lactate | ||
Reagent 2 | Nicotinamide adenine dinucleotide cofactor | 10 mL ×1vial |
Operation steps
1. Determination condition
Dominant wavelength | 340 nm | Auxiliary wavelength | 405 nm |
React temperature | 37℃ | Sample | 6 μL |
React method | Velocity method | Reagent 1 | 200 μL |
React direction | Up | Reagent 2 | 50 μL |
Optical path | 1.0 cm | R1+S incubation time | 3~5 minutes |
R1+S+R2 incubation time | 120 seconds | R1+S+R2 react time | 2 minutes |
2. Operation of Automatic biochemical analyzer
| Blank | Sample |
Double distilled water | 6 μL |
|
Sample |
| 6 μL |
Reagent 1 | 200 μL | 200 μL |
Mix fully, incubate at 37℃ for 3~5 minutes, then add R2 | ||
Reagent 2 | 50 μL | 50 μL |
Mix fully, incubate at 37℃ for 120 seconds. Adjust zero with double distilled water. Measure the variation of absorbance of each tube within 2 minutes. Calculate the △A/min. | ||
3. Operation of Spectrophotometer
| Blank | Sample |
Double distilled water | 24 μL |
|
Sample |
| 24 μL |
Reagent 1 | 800 μL | 800 μL |
Mix fully, incubate at 37℃ for 3 minutes, then add R2 | ||
Reagent 2 | 200 μL | 200 μL |
Mix fully, incubate at 37℃ for 120 seconds, adjust zero with double distilled water, measure the variation of absorbance of each tube within 2 minutes, calculate the △A/min. | ||
Limitation of the test method
1. Hemolysis of the samples will influence the result.
2. The activity of LDH isoenzymes in serum or plasma is one of the tested items for clinic and the final result should be in accordance with life signs of patients, medical history and other tested items.
Performance index
1. The absorbance of blank is A340nm (10mm) ≥0.7, △ABlank/min ≤0.01.
2. The linearity range is 0~500U/L(r2≥0.995);
3. The precision: intra CV＜5.0%,inter CV＜10.0%;
4. Sensitivity: The minimum detection limit ≤ 9.0U/L.
Notes
1. Choose the approximate optical filter if there is no one that this kit is required.
2. The kit is only for use in vitro. Do not touch it with skin, eyes or take in it.
3. Do not use the reagents from different batch.
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