We have used a competitive immunoassay to determine the concentration of a neuropeptide in CSF samples. According to the literature, the levels of the neuropeptide should fit properly to the standard curve range of the ELISA kit. However, for all the CSF samples, we have obtained absorbance values that are greater than those of the S0 point of the standard curve (which should be the highest one, since it concentration is 0). Therefore, neuropeptide concentrations in our samples are supossed to be negative, and it does not make sense at all. We have followed the protocol rigorously, so we can not understand the outcomes of the experiment. Do you have any idea about what could have happened? Thank you!
Maybe, due to inadequate sample preparation leading to sample containing other solvents or compounds affecting signal generation and/or detection or a blank/control using the wrong solvent, respectively.
Thank you for your answer Andreas Eisenreich
The CSF samples were extraction-free (according to the kit design), and the standards were diluted in peptide-free serum. I am not sure if that could be the cause of the problem.
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