We are currently working with a fluorescent AlphaLISA assay platform and are experiencing issues with detection of the phosphorylated protein of interest, despite numerous attempts at troubleshooting everything from sample stimulus, sample preparation and total protein concentrations. We are able to detect Total levels of the analyte, but phosphorylation is not seen. We have confirmed that the stimuli in use are effective through other means.
Does anyone have any experience working with AlphaLISA assays, and have you had similar issues? If so, what are some things I could try in order to increase the fluorescent signal of the analyte?
if both antibodies recognize the same phosphorylated site, I guess they cannot bind together to produce the fluorescence because they compete with the epitope site.
- Badges
- Science topic
- Similar topics
- Microbiology
- Assays