(1) Carp vitellogenin (VTG) ELISA kit is severely hemolyzed. In the ELISA assay marked with HRP, the hemoglobin remaining in the well has peroxidase-like activity, which catalyzes the coloration of the substrate and causes false positives;
(2) The serum mixed with red blood cells is easy to precipitate or adhere to the polyethylene pores and is not easy to wash;
(3) If there is bacterial contamination, the bacteria may contain endogenous HRP, which will also produce false positive reactions;
(4) The specimen is not completely coagulated. Sometimes, in order to obtain time for rapid detection, the serum is often forcibly centrifuged before the blood has started to coagulate, so that some fibrinogen remains in the serum, and fibrin can be visualized during the ELISA measurement process. Block, prone to false positive results;
(5) The blood collection tube is not washed thoroughly, and it is easy to cross-contaminate after repeated use;
(6) The plastic test tube can absorb the antigen substance. If the sample is placed in the plastic tube for a long time, the antigen content in the sample will decrease, resulting in false negative.
Several methods commonly used to determine ELISA results:
1) Visual determination method: after enzymatic hydrolysis and termination of the reaction by adding a substrate, the color of the positive hole is obviously darker (the lighter than the competition rule) of the negative control; the color close to the negative control is judged as negative.
2) The OD value of the sample well is greater than the OD value of the negative control + 2 ~ 3SD as the threshold of positive results.
3) P / N value (positive hole OD value / negative hole OD value) greater than or equal to 2.1 is positive; P / N value is less than 2.1, but greater than 1.5 is suspicious; P / N is less than 1.5 is negative.
4) Quantitative determination results Calculate the content of the analyte in the sample according to the standard curve.
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