Development of a “reverse capture” autoantibody microarray for studies of antigen-autoantibody profiling

Abstract

Diagnosing cancers based on serum profiling is a particularly attractive concept. However, the technical challenges to analysis of the serum proteome arise from the dynamic range of protein amounts. Cancer sera contain antibodies that react with a unique group of autologous cellular antigens, which affords a dramatic amplification of signal in the form of antibodies relative to the amount of the corresponding antigens. The serum autoantibody repertoire from cancer patients might, therefore, be exploited for antigen‐antibody profiling. To date, studies of antigen‐antibody reactivity using microarrays have relied on recombinant proteins or synthetic peptides as arrayed features. However, recombinant proteins and/or synthetic peptides may fail to accurately detect autoantibody binding due to the lack of proper PTMs. Here we describe the development and use of a “reverse capture” autoantibody microarray. Our “reverse capture” autoantibody microarray is based on the dual‐antibody sandwich immunoassay platform of ELISA, which allows the antigens to be immobilized in their native configuration. As “proof‐of‐principle”, we demonstrate its use for antigen‐autoantibody profiling with sera from patients with prostate cancer and benign prostate hyperplasia.