Development of a surface plasmon resonance-based immunoassay for Listeria monocytogenes.
A polyclonal antibody was produced against Internalin B (InlB)-enriched extract and used to develop an inhibition assay to detect Listeria monocytogenes cells in solution using surface plasmon resonance. The gene sequence encoding for the InlB protein was cloned into a Qiagen pQE-60 vector, expressed in Escherichia coli, and purified by immobilized metal affinity chromatography. Protein G-purified anti-InlB-enriched extract polyclonal antibody was incubated with various concentrations of L. monocytogenes cells and subsequently injected over a purified-recombinant InlB (rInlB)-immobilized CM5 sensor chip surface. A decrease in antibody binding response was observed with increasing L. monocytogenes cell concentrations. Intraday and interday assay variability studies were carried out to evaluate precision and reproducibility. The assay had a limit of detection of less than 2 x 10(5) cells per ml and could be successfully reproduced with coefficients of variation of between 2.5 and 7.7%.
Leonard
Paul
P
School of Biotechnology and National Centre for Sensor Research, Dublin City University, Dublin 9, Ireland.
Hearty
Stephen
S
Wyatt
Gary
G
Quinn
John
J
O'Kennedy
Richard
R
D000818:Animals; D000906:Antibodies; D004269:DNA, Bacterial / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005506:Food Contamination / Q000032:analysis*; D005516:Food Microbiology; D008089:Listeria monocytogenes / Q000276:immunology / Q000302:isolation & purification*; D008970:Molecular Weight; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance / Q000379:methods*
D000906:Antibodies; D004269:DNA, Bacterial