Excessive Primary Antibody Concentration Saturating Wells (ELISA (High Background))

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High background signal with poor signal-to-noise ratio. Standard curve may show compressed dynamic range or plateau at lower concentrations than expected. Common causes: 1) Primary antibody concentration exceeds optimal working dilution 2) Antibody titration not performed to determine optimal concentration 3) Non-specific binding increases proportionally with antibody excess Solutions: 1) Dilute primary antibody to optimal concentration listed in datasheet 2) Perform titration experiments to determine optimal working dilution 3) Start with manufacturer-recommended dilution and optimize systematically