Simultaneous detection of multiple proteins with an array-based enzyme-linked immunosorbent assay (ELISA) and...

Protein arrays hold a promise in basic and clinical applications. As the first step to develop such array system, I used an array-based enzyme-linked immunosorbent assay (ELISA) and enhanced chemiluminescence (ECL) to demonstrate the feasibility of simultaneous detection of multiple proteins. In the direct ELISA system, different known immunoglobulin Gs (IgGs) were immobilized onto polyvinylidine difloride (PDVF) membrane through 96-well format Bio-Dot unit. The antigens were then individually and collectively detected by incubation of membranes with different antibodies coupled with ECL. In the sandwich ELISA system, the cytokine capture antibodies were immobilized onto PDVF membranes. The membranes were then incubated with single cytokine or a combination of different cytokines. The captured cytokines were detected by biotin-conjugated antibodies coupled with ECL system. Experiments demonstrated that multiple IgGs and cytokines could be simultaneously detected using this approach with high specificity and sensitivity. More importantly, cytokines from biological samples were detected using this approach, which can be used in any general laboratory setting without any sophisticated equipment. This concept could be extended to develop a protein-based array system.