Interfering factors with ELISA/matrix effect

1 post / 0 new
ckalaska
ckalaska's picture
Interfering factors with ELISA/matrix effect

I am trying to validate an HMGB1 assay and ran into some issues.  Initially linearities performed using the kit dilution buffer gave results of 140% (at a 1:2 dil) to 366% (at a 1:32 dil).  I corrected this by using a low serum sample as a diluent.  But my recovery is still a problem.  My buffer spikes recover around 90% for all levels tested but my samples are literally half of the target values.  The next experiment will be to prepare the standards in glass tubes (I have been using PP tubes) and to acidify samples to see how they react.  Papers published talk about HMGB1 binding IgG and giving falsely low ELISA results.