Development and testing of an enzyme-linked immunosorbent assay (ELISA) for the detection of Staphylococcal Enterotoxin A
1998
Cantera, J.J.L.
A sandwich enzyme-linked immunosorbent assay (ELISA) was developed in this study. Purified staphylococcal enterotoxin A (SEA) from Staphylococcus aureus FRI-100 was used as antigen for the production of anti-enterotoxin A IgG. The anti-enterotoxin A IgG was purified by ion-exchange chromatography. Two types of antibody-enzyme conjugates the Fab'-horseradish peroxidase and the whole antibody-HRP, were prepared using N-N'-0-phenylene-dimaleimide as the cross-linking reagent. Checkerboard titrations to find the optimal working dilutions of immunoreactants, and cross-adsorption to inhibit non-specific reactions, were done. The Fab-HRP was more sensitive than the whole antibody-HRP conjugate and can detect 0.98 mg SEA per ml of sample. The system is less sensitive than those developed by other authors, but can already be used to monitor the safety of food even at a level which can not generally effect intoxication. Enterotoxin A production of several S. aureus strains grown in brain heart infusion using the shake-flask method was quantified using the developed system. A maximum amount of 0.187 ug of SEA per ml of undiluted culture medium was produced by strain NRRL B314 (BIOTECH No. 1582). Enterotoxin production and growth of S. Aureus FRI-100 (BIOTECH No. 1351) in full cream milk incubated at ambient room temperature was monitored during the 18-hr incubation period. No significant correlation was observed between toxin concentration and cell population due to sub-optimal temperature and short incubation time for enterotoxin production
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