I am working on an anti-idiotypic ELISA for human clinical trials and would like to know if human serum is diluted in these assays or used neat. My ab of interest is an IgG4.
Whenever we acquire new ELISA kits, we will optimize our kits by titrating our sample with 5 different concentrations and at a minimum of 3 technical replicates to validate a specific sample concentration for precision and reproducibility. Some of our injury models produce extreme amounts of our proteins of interest and it is important that our samples concentrations fall within the range of the kit’s standard curve. Here is a paper that may be helpful. The paper proposes a standard operating procedure for immunoassays and suggests some critical validation parameters. Hope this help. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4541289/
Whenever we acquire new ELISA kits, we will optimize our kits by titrating our sample with 5 different concentrations and at a minimum of 3 technical replicates to validate a specific sample concentration for precision and reproducibility. Some of our injury models produce extreme amounts of our proteins of interest and it is important that our samples concentrations fall within the range of the kit’s standard curve. Here is a paper that may be helpful. The paper proposes a standard operating procedure for immunoassays and suggests some critical validation parameters. Hope this help. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4541289/
I agree with Andrew. When you start to use an ELISA kit. I recommend you make a scheme of what you need. Are you working with human serum? Check if your serum is not hemolysed. I used to make the standard curve and trial 4 or 5 samples. Whit this you see if you need to diluted or not.
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