2. Standard solution preparation: Add 0.5ml of distilled water before use and mix well to form a solution of 4000pg/ml. Set 8 tubes of standard tube, and add 200 ul of standard dilution solution to each tube. Add 200 ul of the standard solution of 4000 pg/ml to the first tube, mix and aspirate 200 ul with the sampler, and transfer to the second tube. Repeat the dilution as described above, and aspirate 200 ul from the seventh tube and discard it. The eighth tube is a blank control.

3. The 10× specimen dilution was diluted 1:10 with distilled water (example: 1 ml concentrated dilution + 9 ml distilled water).

4. Washing solution: diluted 1:20 with distilled water (example: 1 ml concentrated washing solution added to 19 ml of distilled water)

Test procedure

1. Loading: Add 100 ul of standard or sample to be tested in each well. Mix the reaction plate thoroughly and let it stand at 37 °C for 120 minutes.

2. Wash the plate: Wash the plate thoroughly with washing solution 4-6 times, and dry it on the filter paper.

3. Add 100 ul of the first antibody working solution to each well. The reaction plate was thoroughly mixed and placed at 37 ° C for 60 minutes.

4. Wash the board: the same as before.

5. Add 100 ul of enzyme-labeled antibody working solution per well. The reaction plate was placed at 37 ° C for 30 minutes.

6. Wash the board: same as before.

7. Add 100 ul of substrate working solution to each well and let it react at 37 ° C for 15 minutes in the dark.

8. Add 100 ul of stop solution to each well and mix.

9. Measure the absorbance at 450 nm using a microplate reader within 30 minutes.

Result calculation and judgment

1. All OD values ​​should be subtracted from the blank value before calculation.

2. Take the standard products 2000, 1000, 500, 250, 125, 62.5, 31.2, 0 pg/ml as the abscissa and OD as the ordinate. Draw on the coordinate paper and draw the standard curve.

3. Find the corresponding MCP-1 content on the graph based on the OD value of the sample, and multiply by the dilution factor.

Kit performance

1. Sensitivity: The minimum MCP-1 detection concentration is less than 15pg/ml.

2. Specificity: Recombinant or natural rat MCP-1 can be detected simultaneously. Does not cross-react with other cytokines in rats.

3. Repeatability: The coefficient of variation in the plate and plate is less than 10.6%.

Precautions

1. It is recommended to make double holes for the above standard holes and samples to be tested. The standard curve should be made at the same time for each measurement.

2. The washing process is critical. Insufficient washing will result in an accuracy error and an erroneous rise in the OD value.

3. After the slats are opened, the remaining slats should be sealed again to keep the slats dry .

4. This kit should be stored in a 4oC refrigerator.

5. This kit is for scientific research only and cannot be used for clinical diagnosis!