1. Add samples: Add the sample dilution to the coated plate, dilute the serum in PBS or saline at 1:20, and then add 10 ul to the reaction wells of the microplate.

2. Control addition: 3-well negative control, 1-well positive control, and 100ul control serum.

3. Incubation: Shake the reaction plate to mix the sample well and then place it in a 37 ° C incubator or water bath for 20 minutes.

4. Floor washing: Dilute 15ml of washing solution to 300ml with distilled water. When washing by hand, the contents of the reaction plate air should be poured out and positioned in the hole filled with washing liquid, and then stand 30 to shake violently. Repeat this operation 5 times and beat it dry. The machine needs to perform 5 times.200ul of wash solution can be injected into each well or filled, then left for 30 minutes and blotted.

5. Add enzyme-labeled working solution: Add 100ul of enzyme-labeled working solution to each well in a 37 ° C incubator and wash the plate five times after 20 minutes. The washing plate is the same as in step 4.

6. This is the latter step of the reaction, with color development and reaction termination: 50ul of the substrate is added to the reaction well and the color is developed for 10 minutes in the dark at 37 ° C. The solution in each well was stopped and mixed to terminate the reaction. The use of the microplates shall be performed under sterile conditions. Carefully disinfect and clean the plates after use, but note that some plates need to read the instructions carefully to see if they can be sterilized at high temperatures.