Immunomagnetic nanoparticle-based sandwich chemiluminescence-ELISA for the enrichment and quantification of E. coli

Rapid methods for the quantification of Escherichia coli are required for the monitoring of faecal contamination in water to secure public health. The immunomagnetic separation (IMS) offers rapid enrichment and purification of bacteria in complex matrices and is compatible with immunoassays. By means of this technique, non-target cells and matrix components which might interfere with subsequent analytical methods are removed. We present the synthesis of magnetic nanoparticles (MNPs) and covalent coupling to antibodies against the enterobacterial common antigen (ECA) for use with IMS. Quantification was carried out with a chemiluminescence-based sandwich enzyme-linked immunosorbent assay (ELISA). Our anti-ECA-MNPs allow for a group-specific enrichment of bacterial cells, which can be combined with a species-specific analytical method. The particles were used along with commercially available magnetic columns for the selective enrichment of E. coli from 10-mL water samples. The volumetric enrichment factor was 9. For enriched samples, the limit of detection was reduced from 5.0 x 10(6) cells center dot mL(-1) to 2.6 x 10(5) cells center dot mL(-1). Using 200 A mu L anti-ECA-MNPs, we determined a recovery of 97 A +/- 6% for a sample containing 10(6) cells center dot mL(-1) and 89 A +/- 2% for a sample containing 10(7) cells center dot mL(-1). The overall time for cell enrichment and detection was 3 h 45 min.