Identification of Staphylococcus aureus enterotoxins A and B genes by PCR-ELISA.
We developed PCR-enzyme linked immunosorbent (ELISA) assays to detect Staphylococcus aureus enterotoxins A and B genes. The assays use internal biotin-labelled oligonucleotides as capture probes for immobilizing and subsequently detecting target sequences on microtiter plates. The detection limits of the PCR-ELISAs were approximately 250 gene copies, versus 2500 gene copies by agarose gel analysis. The sensitivity of the assays, as determined from a reference panel of 46 coded samples that included DNA purified from 31 different bacterial species and strains, SEA and SEB plasmid controls, and no-template controls was 100%. No cross-reactivity was observed with DNA from non-staphylococcal species. Using 27 clinical isolates of S. aureus, the SEA PCR-ELISA identified the enterotoxin A (sea) gene in 26 samples, and the SEB PCR-ELISA identified the enterotoxin B (seb) gene in all 27 samples. Compared with conventional antigen capture ELISAs for SEA and SEB toxins, the PCR-ELISAs showed overall superior detection limits. The sensitivity and specificity levels of the SEA PCR-ELISA and the SEA toxin ELISA were comparable within their respective detection thresholds, but the sensitivity and specificity of the SEB PCR-ELISA was much greater than that of SEB toxin ELISA.
Gilligan
K
K
United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD 21702-5011, USA.
Shipley
M
M
Stiles
B
B
Hadfield
T L
TL
Sofi Ibrahim
M
M
D001419:Bacteria / Q000235:genetics; D001710:Biotin / Q000737:chemistry; D003124:Colorimetry / Q000379:methods; D003429:Cross Reactions; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics; D004768:Enterotoxins / Q000235:genetics* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013211:Staphylococcus aureus / Q000235:genetics
D015342:DNA Probes; D004768:Enterotoxins; C026066:enterotoxin A, Staphylococcal; C031818:enterotoxin B, staphylococcal; D001710:Biotin