Simultaneous multianalyte ELISA performed on a microarray platform

Background: A logical progression of the widely used microtiter plate ELISA is toward a protein array format that allows simultaneous detection of mul tiple analytes at multiple array addresses within a single well. Here we de scribe the construction and use of such a multiplex ELISA to measure prosta te-specific antigen (PSA), alpha (1)-antichymotrypsin-bound PSA (PSA-ACT), and interleukin-6 (IL-6). Methods: We silanized glass plates and printed the appropriate capture anti bodies to allow for the construction of "sandwich" ELISA quantification ass ays. We examined specificity of the assay for appropriate antigen, assemble d calibration curves, and obtained PSA concentrations for 14 human serum sa mples. We compared the serum PSA concentrations derived through the use of our array with values obtained independently, using a standard ELISA method . Results: R-2 values generated by our microarray for the PSA and PSA-ACT cal ibration curves were 0.989 and 0.979, respectively. Analyte concentrations used for the construction of these curves were 0.31-20 mug of protein/L of diluent. IL-6 calibration curve concentrations were 4.9-300 ng of IL-6/L of diluent. The R-2 value for the IL-6 calibration curve was 0.983. The 14 hu man serum samples screened by this micro-ELISA technique for PSA concentrat ions generated a regression equation (linear) with a slope of 0.83 +/- 0.10 and intercept of 0.74 +/- 0.70 (R-2 = 0.88). Conclusions: Multiplexed ELISA arrays are a feasible option for analyte qua ntification in complex biologic samples. (C) 2001 American Association for Clinical Chemistry.