Rapid analysis and quantification of proteins in high throughput using PAIA assays

The PAIA technology is a novel approach to quantify proteins. The protein of interest (the analyte) is captured by functionalized beads and detected with a specific fluorescence marker. PAIA assays use protrusions on the bottom of 384-well microplates to separate captured analyte-marker complexes from the unbound fluorescent marker which is then measured (Figure 1). The assay is automation-friendly since no washing steps are needed. Thus, time to result, sample volume, and hands-on time are substantially reduced compared to other immunoassay formats. The PAIA approach has been successfully used for IgG quantification in supernatants of antibody producing cells and a PAIA IgG kit has already been launched. Because it is possible to choose a marker against a selective epitope of the analyte the PAIA technique can address a broad range of analytes. Here, we present assay systems that are currently under development e.g. for analytes that contain affinity tags. Analytes carrying two tags can be captured via the first tag while the fluorescence marker binds the second tag. Figure 2 shows an example of proteins that were captured by their Strep-tag and detected by a Flag-tag specific marker. For proteins with just one tag or epitope, competitive assays are more suitable. In these assays the fluorescence marker contains the same tag or epitope as the analyte and both compete for binding sites on the capture beads. We believe that the high throughput, the small sample volume, and the low overall costs of PAIA assays represent substantial advantages over existing methods.