The use of beta-lactamase in enzyme immunoassays for detection of microbial antigens

Abstract

The sensitivity and performance characteristics of enzyme immunoassays (EIA) depende to a great extent on the kinetics of the enzyme-substrate system used as indicator. We labeled a variety of polyclonal and monoclonal immunoglobulins with purified beta-lactamase and used them in sensitive EIA systems for the detection of a number of microbial antigens. Polyclonal antibodies to rotavirus, adenovirus, and Haemophilus influenzae type b polyribitol phosphate and monoclonal antibodies to dengue virus were labeled with beta-lactamase and used to provide sensitive direct EIA systems for the detection of the corresponding antigens. In addition, antibodies directed at animal immunoglobulins were labeled with beta-lactamase and used in indirect EIA for the detection of viral antigens with unlabeled anti-viral monoclonal and polyclonal antibodies. Similarly, avidin from Streptomyces was labeled with beta-lactamase and used to detect viral antigens tested for in an avidin-biotin format. Enzyme immunoassay systems with beta-lactamase-labeled antibodies were also used to detect rotaviral and adenoviral antigens in rectal swab specimens from children with acute gastroenteritis. The sensitivity of the beta-lactamase EIA compared favorably with that of analogous EIA systems using alkaline phosphatase or horseradish peroxidase. The results of a beta-lactamase EIA were easily determined by naked eye and a permanent record of the qualitative results obtained by the use of a standard office photocopier, obviating the need for an expensive colorimeter. Enzyme immunoassays using beta-lactamase have potential as practical assay systems for the detection of a wide range of microbial antigens using monoclonal and polyclonal antibodies.