Development of a fluorescent latex microparticle immunoassay for the detection of staphylococcal enterotoxin B (SEB).
Staphylococcus aureus enterotoxin B (SEB) is a highly heat resistant enteric toxin with a potential as a biothreat agent. A sensitive method for the detection of staphylococcal enterotoxins is needed for food safety and food defense monitoring. The objectives of this research were to develop a competitive fluorescent immunoassay with detection of SEB below toxic levels of 1 ng/mL and to minimize sample preparation. Anti-SEB was immobilized onto carboxylated polystyrene microparticles, and SEB was labeled with fluorescein isothiocyanate (FITC). The concentrations of these reagents were optimized for the detection of SEB below 1 part per billion (1 ng/mL), and other assay conditions (sample volumes and incubation periods) were optimized. Drinking water and milk samples were spiked with 0.125-10 ng/mL SEB and were equilibrated overnight prior to analysis. The water and milk samples were directly analyzed, but heating the milk samples for 10 min at 90 degrees C improved the assay performance. SEB in samples bound with the anti-SEB linked to the latex followed by the competitive binding of SEB-FITC tracer. The excess, unbound tracer was separated by centrifugation, and the fluorescence density of the supernatant was measured. SEB was detected at levels as low as 0.125 ng/mL in drinking water and 0.5 ng/mL in whole milk. This fluorescent latex particle immunoassay will be utilized for the detection of SEB in various foods matrices.
Medina
Marjorie B
MB
Microbial Biophysics and Residue Chemistry Research Unit, Eastern Regional Research Center, Agricultural Research Service, U.S. Department of Agriculture, 600 East Mermaid Lane, Wyndmoor, Pennsylvania 19038, USA. mmedina@ errc.ars.usda.gov
D000818:Animals; D004768:Enterotoxins / Q000032:analysis*; D016650:Fluorescein-5-isothiocyanate; D005456:Fluorescent Dyes; D015200:Fluoroimmunoassay / Q000379:methods*; D005506:Food Contamination / Q000032:analysis*; D007840:Latex; D008863:Microspheres; D008892:Milk / Q000737:chemistry; D014867:Water / Q000032:analysis
D004768:Enterotoxins; D005456:Fluorescent Dyes; D007840:Latex; D014867:Water; C031818:enterotoxin B, staphylococcal; D016650:Fluorescein-5-isothiocyanate