ProteoChip: A highly sensitive protein microarray prepared by a novel method of protein immobilization for application of protein‐protein interaction studies

Abstract

We have developed a highly sensitive microarray protein chip, ProteoChip, coated with ProLinker™, novel calixcrown derivatives with a bifunctional coupling property that permits efficient immobilization of capture proteins on solid matrixes and makes high‐throughput analysis of protein‐protein interactions possible. The analysis of quartz crystal microbalance showed that both monoclonal antibody (mAb) and antigen (Ag) bound to the gold film of the sensor surface coated with ProLinker™ B and that it is useful for studies of Ab‐Ag interactions. ProteoChip, aminated glass slide coated with ProLinker™ A, was also demonstrated to be useful for preparation of high‐density array spots by using a microarrayer and for analysis of analyte Ags either by direct or sandwich methods of fluorescence immunoassay. The detection sensitivity of ProteoChip was as low as 1–10 femtogram/mL of analyte protein, useful for detection of tumor markers. ProteoChip was also useful for studies of direct protein‐protein interactions as demonstrated by analysis of integrin‐extracellular matrix protein interaction. These experimental results suggest that ProteoChip is a powerful tool for development of chip‐based lead screening microarrays to monitor protein‐protein interactions (i.e. drug target) as well as for biomarker assays which require high detection sensitivity.