Enzyme-linked immunosorbent assay for Escherichia coli heat-stable enterotoxin

Abstract

The sensitivity of an enzyme-linked immunosorbent assay (ELISA) to detect pure native Escherichia coli heat-stable toxin (ST) and to identify ST-producing strains among clinical isolates was determined. Two synthetically produced ST preparations were used to raise hyperimmune antisera in rabbits and goats: ST(S), which has the same antigenicity as native ST; and ST(C), which is 15-fold more immunogenic. These antisera were used in the double-sandwich technique as either crude double-species antisera or pure singlespecies antibody. The sensitivity of the assay was increased by using either a purer antibody preparation or the antiserum to the more potent immunogen; the assay in which pure antibody to ST(C) was used was 2,857-fold more sensitive in detecting ST than the assay in which crude antiserum to ST(S) was used. The minimum amount of ST detectable by the ST(C) ELISA was 140 pg/ml, which was an amount 285-fold smaller than that detectable by the suckling mouse assay. Among 50 human E. coli isolates examined by both the ST(C) ELISA and an ELISA for heat-labile toxin (LT), which had a sensitivity of 290 pg/ml for LT, the respective toxins were consistently identified in broth cultures of 10 LT' and ST-, 15 LT' and ST+, and 10 LT-and ST' strains, and there were no false-positive responses. The ST(C) ELISA also detected ST in all of seven ST-producing E. coli strains tested of human origin, which had been shown elsewhere by DNA hybridization probes to have ST-coding genes of either human or porcine origin, and in all of three ST-producing E. coli strains tested of porcine origin. These results indicate that the sensitivity of the ST(C) ELISA is the same as that of previously described LT ELISAs. The concomitant use of both ST and LT ELISAs provides a rapid, simple, and sensitive method for identifying among clinical isolates enterotoxigenic strains of E. coli which produce either toxin.

Properties of crude Campylobacter jejuni heat-labile enterotoxin

“…To determine whether toxigenic strains of C. jejuni can be identified by ELISA of unconcentrated broth supernatants, these fractions of the eight clinical isolates were tested in both the LT/LT and GM/LT ELISAs. We have found in previous studies that unconcentrated broth supernatants of LT-producing enterotoxigenic E. coli strains always give an optical density of >0.200 in both of these ELISAs, whereas those of nontoxigenic strains consistently give values of <0.200 (29). Only the broth supernatants of the three strains on July 10, 2020 by guest http://iai.asm.org/ Downloaded from that produced the most CJT gave a positive response (i.e., an optical density .0.200) in the LT/LT ELISA, whereas the GM/LT ELISA distinguished between CJT-producing strains and those which produced either no or a negligible quantity of toxin ( Fig.…”

Pathologicalstudy on Colibacillosis in Chickens and Detection of Escherichia Coli by PCR

“…Over 700 antigenic types (serotypes) are recognized based on 0, H and K antigens. Different serological test such as fluorescent antibody test (FAT), enzyme-linked immunosorbent assay (ELISA) test, serum agglutination test (SAT), growth inhibition test (GIT) are used for the detection of enterotoxigenic and enteropathogenic E. coli (Klipstein, 1984). Different molecular techniques such as nucleic acid hybridization and polymerase chain reaction (PCR) studies can also be used to detect pathogenic E. coli and DNA probes directed at a number of genes have also been developed (Wani et al, 2003).…”

Evaluation of a nonisotopically labeled oligonucleotide probe to detect the heat-stable enterotoxin gene of Escherichia coli by the DNA colony hybridization test

“…This procedure requires experimental animals and is troublesome. Elaborate immunological techniques to detect ST in vitro have been developed recently (13,14,23). These tests require special equipment and are therefore unsuitable for small laboratories and field studies.…”