In situ rolling circle amplification surface modifications to improve E. coli O157:H7 capturing performances for rapid and sensitive microfluidic detection applications

Abstract We investigated the application of rolling circle amplification (RCA) to modify microfluidic channels for potential sensitive detection applications. To this end, a novel in situ capturing RCA (cRCA) strategy was used to modify the inner surfaces of microfluidic channels with cRCA products that featured repeating tandem capturing aptamers specific for E. coli O157:H7 cells. We showed that the in situ cRCA reaction modified microfluidic channels demonstrated significantly enhanced capturing efficiency in a wide range of flow rates when compared with the unit-aptamer approach. We demonstrated for the first time that microfluidic surfaces modified with the in situ cRCA products showed peak capturing performances both in terms of target capturing efficiency and specificity, and this was likely due to unexpected base-pairing that resulted in altered secondary structures of the capturing aptamers. Our data suggest that the in situ cRCA surface modification is a promising strategy to improve capturing performances in microfluidic devices in sensitive detection applications that also require high throughput. However, cRCA reaction conditions, particularly reaction time and concentrations of initial circular template, must be carefully investigated before the potentials of the in situ cRCA surface modification approach can be fully realized.