title: Subtractive inhibition assay for the detection of E. coli O157:H7 using surface plasmon resonance.
abstract: A surface plasmon resonance (SPR) immunosensor was developed for the detection of E. coli O157:H7 by means of a new subtractive inhibition assay. In the subtractive inhibition assay, E. coli O157:H7 cells and goat polyclonal antibodies for E. coli O157:H7 were incubated for a short of time, and then the E. coli O157:H7 cells which bound antibodies were removed by a stepwise centrifugation process. The remaining free unbound antibodies were detected through interaction with rabbit anti-goat IgG polyclonal antibodies immobilized on the sensor chip using a BIAcore 3000 biosensor. The results showed that the signal was inversely correlated with the concentration of E. coli O157:H7 cells in a range from 3.0 × 10(4) to 3.0 × 10(8) cfu/mL with a detection limit of 3.0 × 10(4) cfu/mL. Compared with direct SPR by immobilizing antibodies on the chip surface to capture the bacterial cells and ELISA for E. coli O157:H7 (detection limit: both 3.0 × 10(5) cfu/mL in this paper), the detection limit of subtractive inhibition assay method was reduced by one order of magnitude. The method simplifies bacterial cell detection to protein-protein interaction, which has the potential for providing a practical alternative for the monitoring of E. coli O157:H7 and other pathogens.
authors.affiliation: College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310029, Zhejiang, China. wang_yi_xian1986@sina.com
authors.forename: Yixian
authors.initials: Y
authors.lastname: Wang
authors.forename: Zunzhong
authors.initials: Z
authors.lastname: Ye
authors.forename: Chengyan
authors.initials: C
authors.lastname: Si
authors.forename: Yibin
authors.initials: Y
authors.lastname: Ying
mesh_terms: D000818:Animals; D000907:Antibodies, Bacterial / Q000276:immunology; D001681:Biological Assay / Q000379:methods*; D002498:Centrifugation; D004797:Enzyme-Linked Immunosorbent Assay; D019453:Escherichia coli O157 / Q000166:cytology / Q000276:immunology / Q000302:isolation & purification*; D055767:Immobilized Proteins / Q000276:immunology; D015203:Reproducibility of Results; D020349:Surface Plasmon Resonance / Q000295:instrumentation / Q000379:methods*
chemical_list: D000907:Antibodies, Bacterial; D055767:Immobilized Proteins