Indirect ELISA - Interpreting results - (Nov/08/2011 )
I'm a sophomore from Ohio.
For my labs involving indirect ELISA, I was given various sera immunised with different peptides to see which peptide was better at eliciting an immune response. I made titration curves for each serum (optical density on y-axis and serial dilution factor (i.e. neat, 1/2, 1/4, 1/8, 1/6 etc) on x-axis). I was then told to determine the titre by reading the end-point from each curve i.e. obtain the value of the serial dilution factor (from the x-axis) where the curve from serum crosses the baseline provided by the negative control.
So now that I've obtained a value for titre in the form of the highest serial dilution factor, what do I do with this? I know these values can help tell me which sera have worked and have got more antibodies but how can I quantify this further? Some people talk about drawing standard curves or reference curves but does that fit in with titration curves? Do I need to know the values of the original concentrations? I don't understand beyond this point at all.
Help is greatly appreciated, thanks.
What is the information you are trying to obtain? You have a 'titer' the highest dilution of antisera that you distinguish from "0". (Have you run replicates of the 0 to determine the range of the values?). You can determine if the antisera 'sees' different forms of the peptide: coat the wells with different forms of the peptide and compare.
Further work would be to make an competative elisa using your antisera. It depends upon how much work you want to put in