Custom Competitive ELISA - (Apr/15/2010 )

Hi, I'm making a custom ELISA.

We've gotten a quote for custom polyclonal Ab made from a custom peptide. We are going to get a portion of the peptide biotin conjugated for the tracer and have unconjugated peptide for the standards.

My question is; what concentration should the tracer be?

I know how much unconjugated peptide I need to make the standards from but I don't know how much biotin conjugated peptide is going to be required to make the tracer. If I knew what concentration to make the tracer I could figure out how much raw material I need. I realize we need more conjugated peptide than unconjugated. The tracer is going to be stronger than the highest standard and it's going to be used in every single well; therefore I assume the majority of peptide produced is going to be conjugated; I'm just trying to figure out the ratio.

Right now I'm thinking if 10mg of peptide is produced that 3mg should cover the standards and the rest be conjugated to biotin. The conjugation won't be 100% so some will be lost. Perhaps I should go with 2:8 instead of 3:7?

Does anyone know an approximate concentration for the tracer in competitive ELISAs?

Thank you.

"Stronger" ? I think the 2: 8 should be sufficient.

sgt4boston on Apr 20 2010, 03:21 PM said:

I meant the tracer would be of higher concentration than the highest standard.

Thanks for the input.