A RECEPTOR-BASED IMMUNOASSAY TO DETECT STAPHYLOCOCCUS ENTEROTOXIN-B IN BIOLOGICAL-FLUIDS

A rapid, simple, and inexpensive sandwich enzyme-linked receptor based immunodot assay was developed for the detection of staphylococcal ent erotoxin B (SEE) in human fluids by using purified glycosphingolipid d igalactosylceramide (diGalCer) receptor for SEE. Three micrograms of d iGalCer was immobilized on a polyvinyledene difluoride membrane and th e membrane was subsequently incubated with primary and secondary alkal ine-phosphatase-labeled antibodies, A positive reaction was discerned as a blue spot. As little as 1 ng/ml of SEE could be detected in the a ssay. SEE did not bind to structurally related glycosphingolipids, suc h as glucosylceramide, galactosylceramide, and lactosylceramide in thi s assay. Of five monoclonal anti-SEE antibodies and commercial anti-SE B antiserum tested, the latter was the most sensitive in our assay. Th e specificity of SEE assay was assessed by comparison with structurall y related toxins, for example, staphylococcal enterotoxin A, and toxic shock syndrome toxin 1 (TSST-1), TSST-1 was not detected in the assay . This was because these toxins were not recognized by the anti-SEE an tibody and did not bind to diGalCer. In conclusion, we believe that th is assay may be widely applicable because it is highly specific for SE E, it does not require special equipment, and the results can be obtai ned within few hours with the naked eye. Since the receptor for SEE ha s a long shelf life, it can be easily stored and used for a long time. (C) 1997 Academic Press.