The conditions required for ELISA experimental operation are quite strict, both in terms of hardware conditions and technical requirements for the operators. The following are some issues to be aware of when conducting ELISA experiments.

General rules for ELISA experiments

1. To ensure the accuracy of the pipette, the error should not exceed 2%. It can be determined using water and an electronic balance. But it is best to have professional personnel for correction.

2. To be equipped with one 20ul, one 50ul, one 100ul, one 1000ul, and one shotgun. After absorbing different liquids, the gun head needs to be replaced. Even when absorbing standard samples.

3. One hour before the experiment, remove the reagent kit from the refrigerator and allow all reagents to return to room temperature to ensure more stable results.

4. During the experiment, the substrate should be stored away from light.

5. When using a gun to suck liquid, the speed should not be too fast to avoid the generation of bubbles and inaccurate suction volume.

6. When sucking liquid, use a gun with a range close to the required amount to reduce errors.

7. When adding liquid to the enzyme-linked immunosorbent assay (ELISA) well, avoid contact between the pipette tip and the liquid inside the well. This will allow the droplets on the pipette tip to come into contact with the well wall, and the droplets will naturally flow down.

8. After adding all the liquid, gently shake the enzyme-linked immunosorbent assay plate parallel to the table for 30 seconds and mix the liquid well. You can also use the shaking function of the enzyme-linked immunosorbent assay (ELISA) reader.

9. When taking a warm bath, use adhesive or tape to seal the enzyme-linked immunosorbent assay (ELISA) plate to prevent water evaporation.

10. When washing the board, each time the washing solution is added, it should be left to stand for 1 minute to make the cleaning more thorough. When there is no washing machine, after pouring the liquid, the enzyme-linked immunosorbent assay (ELISA) plate should be vigorously dried on newspaper or parchment paper.

11. When the washing solution is insufficient, distilled water can be used to prepare a pH 7.4, 0.02M phosphate buffer solution, and 0.1% Tween 20 can be added as the washing solution. After adding 1/1000 sodium azide, it can be stored for a long time.

12. The substrate is photosensitive and should be prepared before use.

13. Before testing, turn on the ELISA reader and let it stabilize for more than 10 minutes.

14. The substrate has certain toxicity, and the termination solution is corrosive to the skin, so contact should be avoided as much as possible.

15. The sample to be tested needs to be clarified, otherwise it will affect the results.

16. The warm bath time should comply with the regulations of the reagent kit.

17. Double hole experiments should be conducted as much as possible to ensure the accuracy of the data.

18. Samples with questionable results should be confirmed using other methods.

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