A High-Throughput Solid-Phase Microplate Protein-Binding Assay to Investigate Interactions between Myofilament Proteins

Figure 2

Representative design of a 96-well microtitering plate assay for binary binding consisting of an immobilized protein incubated with two variant soluble interacting proteins in 8 serial concentrations of triplicate wells. Immobilized protein is equally and maximally coated in the experimental wells, rows A to H and columns 4 to 9. Columns 1 to 3 and 10 to 12 are left uncoated (incubated with coating buffer only) as controls. Following coating all wells are blocked, and the wells in columns 1 to 6 are incubated with serial dilutions, one dilution per row, of one of the soluble proteins, and wells in columns 7 to 12 are incubated with serial dilutions of the other soluble protein variant. All wells are then processed uniformly for ELISA detection.