Operation steps of ADIPOR2 pig adiponectin receptor 2 enzyme-linked immunosorbent assay kit:

1. Dilution and sample addition of standard: Set 10 standard wells on the enzyme-linked immunosorbent assay (ELISA) coated plate, add 100 μ l of standard to the first and second wells respectively, and then add 50 μ l of standard diluent to the first and second wells, mix well; Then take 100 μ l from each of the first and second wells of di and add them to the third and fourth wells, respectively. Then add 50 μ l of standard diluent to the third and fourth wells and mix well; Then, take 50 μ l each from the third and fourth wells and discard them. Then, take 50 μ l each and add 50 μ l to the seventh and eighth wells respectively. Add 50 μ l of standard dilution solution to the seventh and eighth wells respectively, mix well, and add 50 μ l of standard dilution solution from the seventh and eighth wells to the fifth and sixth wells respectively. Finally, add 50ul of standard dilution solution to the fifth and sixth wells and mix well; After mixing, take 50 μ l from each of the fifth and sixth wells and add it to the ninth and tenth wells. Then, add 50 μ l of standard diluent to each of the ninth and tenth wells. After mixing, take 50 μ l from each of the ninth and tenth wells and discard. After dilution, the sample volume for each well was 50 μ l, with concentrations of 180 ng/L, 120 ng/L, 60 ng/L, 30 ng/L, and 15 ng/L, respectively.

2. Sample addition: Set up blank wells (blank control wells without sample or enzyme-linked immunosorbent assay, the rest of the steps are the same) and test sample wells respectively. Add 40 μ l of sample diluent to the well of the test sample on the enzyme-linked immunosorbent assay (ELISA) coated plate, and then add 10 μ l of the test sample (with a final dilution of 5 times). Add the sample to the bottom of the enzyme-linked immunosorbent assay plate well, avoiding touching the well wall as much as possible, and gently shake and mix well.

3. Incubation: Cover the plate with a sealing film and incubate at 37 ℃ for 30 minutes.

4. Liquid preparation: Dilute the concentrated washing solution 30 times (20 times 48T) with distilled water and set aside for later use.

5. Washing: Carefully remove the sealing film, discard the liquid, shake dry, fill each hole with washing solution, let it stand for 30 seconds, then discard. Repeat this process 5 times and pat dry.

6. Enzyme addition: Add 50 μ l of enzyme labeled reagent to each well, except for blank wells.

7. Incubation: Follow the same procedure as in 3.

8. Washing: Follow the same procedure as in 5.

9. Color development: Add 50 μ l of color developing agent A50 to each well, then add 50 μ l of color developing agent B50, gently shake and mix well, and develop color at 37 ℃ in the dark for 15 minutes

10. Termination: Add 50 μ l of termination solution to each well to terminate the reaction (at this point, blue will turn yellow).

11. Measurement: Measure the absorbance (OD value) of each well in sequence using blank air conditioner zero and 450nm wavelength. The measurement should be conducted 15 minutes after adding the termination solution.