A technician observes high background in many wells of an ELISA plate. Which change to the protocol most directly addresses nonspecific binding and why?

Eliminate the blocking step entirely and instead reduce the concentration of capture antibody so fewer binding sites are available for the target antigen; removing blocking actually forces better specificity by lowering antibody availability.

Increase blocking time using casein or BSA and add more thorough wash steps between incubations; blocking occupies uncoated plastic and washes remove unbound reagents, both reducing nonspecific adsorption that produces background signal.

Switch to a fluorescent substrate without changing blocking or washes; fluorescence detection eliminates the need for blocking because fluorescent dyes are not affected by nonspecific protein binding on the plastic surface.

Replace washes with a single brief rinse in distilled water between steps because detergent-containing wash buffers are known to permanently inactivate antibody binding, and water will remove all loosely bound proteins without altering antibody-antigen affinity.