I'm currently analysing data of Inteleukin measurement with ELISA. Are you using 4PL Method? Which Software are you using? How can you verify if your Standard curve is correct with 4PL method?
Hi Wiem,
Many, but not all, ELISA standard curves can be fit with a 4PL (the kits I use to assay hormones generally recommend a 4PL fit). Both pieces of software that I use, SoftMax Pro (comes with my plate reader) and GraphPad Prism, have a number of different options for curve fitting and will give you a goodness of fit statistic (R2). Generally speaking, I want my curve to have an R2 of 0.9 or higher.
Cayman Chemical, one of the companies whose kits I use, made an excel spreadsheet to help with analysis: https://www.caymanchem.com/analysisTools/immunometric
I hope that helps! Good luck!
Danielle
Hi,
Yes, as previously said, 4PL curve fitting is usual for ELISA standard curve. I'm using GEN5 software from Biotek Instruments since my ELISA plate reader comes from Biotek Instruments too and they purchased everything together to me. To check the quality of my standard curve, I use the fitting data parameters to check that R² is above 0.9, as does Danielle. I'm also used to do a back-calculation of the standards concentration using the curve equation so that I can check the standards concentrations obtained with that assay curve equation are well in the range of +/- 20% of the targeted values. Thanks to Gen5 I was able to include this internal validation step in my reading protocol directly made in the software.
Hope it helps!
Good luck,
Cécile
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