| Abstract | Aptamer based microfluidic platforms have been developed rapidly in recent years, and strategies to improve detection sensitivities of such platforms have attracted a significant amount of attention. To achieve whole cell sensitive detections by microfluidic devices, a new dual-rolling circle amplification (RCA) detection approach is presented in this study. This dual-RCA approach includes a capturing RCA (cRCA) reaction that is designed to modify microfluidic channel surfaces with long tandem repeating aptamers (i.e. poly-aptamers) to effectively capture target E. coli O157:H7 cells. We demonstrate that this poly-aptamers modified microchannels capture 3-fold more target cells in comparison with microchannels modified with mono-aptamers against the target cells. In addition, signalling RCA (sRCA) is employed in the dual-RCA design to further enhance detection signals. Our results show that the detection signals are enhanced by up to 50 times by sRCA when compared with those with single fluorescence probes. Furthermore, by combing both the cRCA and the sRCA in one dual-RCA detection system, we demonstrate that the detection signals can be significantly enhanced by ∼250-fold. We also show that E. coli O157:H7 detections with the dual-RCA approach can be used in different food matrices, including orange juice and milk where the limit of detection of 80 cells/mL is achieved. In conclusion, this microfluidic device in combination with a dual-RCA to enhance both target capturing and detection signals is a simple and promising approach to sensitive whole-cell detections for food safety inspections. |
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