Development and evaluation of aptamer magnetic capture assay in conjunction with real-time PCR for detection of Campylobacter jejuni

- Nucleic acid aptamers are emerging ligands for pre-analytical sample preparation.
- The DNA aptamer used in aptamer-magnetic capture (AMC) is a novel capture ligand.
- First report on AMC combined with qPCR for detection of Campylobacter jejuni.
- Detection limits of the AMC-qPCR were 1.0-2.0 log10 CFU per sample (0.3-10 ml).
- AMC-qPCR was more efficient than comparative immunomagnetic separation-qPCR.

A prototype method for the concentration and detection of Campylobacter jejuni was developed using a previously reported biotinylated DNA aptamer in conjunction with qPCR. The so-called aptamer-based magnetic capture-qPCR (AMC-qPCR) assay was compared to a similar immunomagnetic separation (IMS)-qPCR assay. In small volume experiments (300 μl) applied to serially diluted C. jejuni suspended in buffer containing a mixed culture of other common food borne pathogens, the lower detection limit of the AMC-qPCR method was 1.1 log10/300 μl C. jejuni cells, one log10 better (lower) than that of IMS-qPCR (2.1 log10 CFU/300 μl). AMC-qPCR capture efficiency was 10-13% at assay detection limit. In 10 ml scale-up experiments, the lower detection limit of AMC-qPCR was 2.0 log10 CFU/10 ml with corresponding capture efficiency of 4-7%. Nucleic acid aptamers are promising alternatives to antibodies for magnetic bead-based capture followed by qPCR detection.