A sandwich-type assay based on quantum dot/aptamer bioconjugates for analysis of E. Coli O157:H7 in microtiter plate format

An optical assay based on CdSe/ZnS quantum dots (QD)/NH₂-Apt bioconjugates for the pathogen detection was presented. QDs with carboxyl functional groups and 72-mer aptamer (against E. coli outer membrane proteins) were used as probes and sensing element. E. coli O157:H7 was selected as a model pathogen and 96-well plate assay in the sandwich hybridization format was constructed. Poly-L-lysine-coated 96-well plate surfaces were used as support material where thiol functionalized aptamers were immobilized by 4-(N-Maleimidomethyl)cyclohexane-1-carboxylic acid 3-sulfo-N-hydroxysuccinimide (sulfo-SMCC). After incubation with the bacteria, CdSe/ZnS QDs/aptamer bioconjugates were added. The fluorescence signals were followed before and after addition of bioconjugates. Probe concentrations, incubation time with E. coli O157:H7 were also optimized. The bioassay could detect the pathogen down to 10² CFU/mL with high selectivity. The detection system was successfully employed in samples, in the presence of interfering compounds.