How to calculate antibody titre in serum of vaccinated animal using Direct ELISA? The OD values obtained at different dilution how to determine the cut off OD against which the antibody titre calculated?
Direct ELISA used only for estimation of antigens. For antibody is indirect ELISA assay, it depend on type of antibody (IgG or IgM). You need specific antigen for IgG also specific labelled anti-antibody, and standards for quantitative results, or positive, negative and cutoff control for qualitative results.
Could you clarify the scheme of your ELISA? Just Your choice is unusual, but sometimes it is possible.
If your question is more focused on determination of cutoff OD, there are a lot of approaches. I can recommend you to start from the easiest method: OD(cutoff)=2*OD(NegControl)
But there are also calculation in relation to ODblanc and even ODpos.
1 - the correlation should be with reference serums tested in HIT or SNT.
2 - for some viruses, no correlation btw DIRECT-ELISA!!! Only Liquid-phase ELISA should be developed (FMDV like example).
I perform 7 dilution of sera of vaccinated animals and also performed 5 dilutions of the negative control of the same animals. After I Run the ELISA. The dilution at which highest OD recorded in vaccinated samples, I selected OD at same dilution of the negative control as cutoff OD and made calculation. I want to know whether the method is right or not.
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