Label-free and high-sensitive detection of Salmonella using a surface plasmon resonance DNA-based biosensor

• Published in: Journal of biotechnology 2012-08, Vol.160 (3-4), p.123-128
• Main Authors: Zhang, Decai, Yan, Yurong, Li, Qing, Yu, Tianxiao, Cheng, Wei, Wang, Long, Ju, Huangxian, Ding, Shijia
• Format: Article
• Language: English
• Subjects: Biosensing Techniques - instrumentation; Biosensor; biosensors; biotechnology; Colony Count, Microbial - instrumentation; detection limit; dextran; DNA; DNA - genetics; Equipment Design; Equipment Failure Analysis; Escherichia coli; genes; hybridization; invA gene; nucleotide sequences; oligonucleotides; Polymerase chain reaction; Salmonella; Salmonella - genetics; Salmonella - isolation & purification; Sensitivity and Specificity; serotypes; Staining and Labeling; streptavidin; Surface plasmon resonance; Surface Plasmon Resonance - instrumentation
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2012.03.024

► A label-free and high-sensitive method based on surface plasmon resonance DNA biosensor has been proposed for detection of Salmonella. ► A modified asymmetric PCR has been introduced to produce predominantly ssDNA target. ► 102CFUmL−1 of Salmonella cells form culture samples can be detected within 4.5h. ► No significant signal was observed to detect E. coli and S. aureus. A method based on surface plasmon resonance (SPR) DNA biosensor has been developed for label-free and high-sensitive detection of Salmonella. A biotinylated single-stranded oligonucleotide probe was designed to target a specific sequence in the invA gene of Salmonella and then immobilized onto a streptavidin coated dextran sensor surface. The invA gene was isolated from bacterial cultures and amplified using a modified semi-nested asymmetric polymerase chain reaction (PCR) technique. In order to investigate the hybridization detection, experiments with different concentration of synthetic target DNA sequences have been performed. The calibration curve of synthetic target DNA had good linearity from 5nM to 1000nM with a detection limit of 0.5nM. The proposed method was applied successfully to the detection of single-stranded invA amplicons from three serovars of Salmonella, i.e., Typhimurium, Enterica and Derby, and the responses to PCR products were related to different S. typhimurium concentrations in the range from 102 to 1010CFUmL−1. While with this system to detect E. coli and S. aureus, no significant signal was observed, demonstrating good selectivity of the method. In addition, the hybridization can be completed within 15min, and the excellent sensor surface regeneration allows at least 300 assay cycles without obvious loss of performance.