Premium
Rapid enumeration of Escherichia coli in oysters by a quantitative PCR‐ELISA
Author(s) -
González I.,
García T.,
Fernández A.,
Sanz B.,
Hernández P. E.,
Martín R.
Publication year - 1999
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.1999.00659.x
Subject(s) - enumeration , escherichia coli , biotinylation , microbiology and biotechnology , streptavidin , primer (cosmetics) , polymerase chain reaction , digoxigenin , chemistry , oyster , biology , taq polymerase , peroxidase , chromatography , enzyme , biochemistry , biotin , gene , thermus aquaticus , gene expression , in situ hybridization , mathematics , organic chemistry , combinatorics , fishery
Direct enumeration of Escherichia coli from oysters was achieved using a polymerase chain reaction (PCR) amplification of the lam B gene coupled with an enzyme‐linked immunosorbent assay (ELISA). Amplified PCR products generated using a digoxigenin‐labelled primer were heat denatured before being quantified by an ELISA. A biotinylated probe immobilized onto streptavidin‐coated microplates was used to capture the digoxigenin‐labelled fragments that were detected with a peroxidase antidigoxigenin conjugate. Subsequent enzymic conversion of substrate gave distinct absorbance differences when assaying oyster samples containing E. coli in the range 10–10 5 cfu g −1 .