PCR-assisted impedimetric biosensor for colibactin-encoding pks genomic island detection in E. coli samples

Abstract

A fast PCR-assisted impedimetric biosensor was developed for the selective detection of the clbN gene from the polyketide synthase (pks) genomic island in real Escherichia coli samples. This genomic island is responsible for the production of colibactin, a harmful genotoxin that has been associated with colorectal cancer. The experimental protocol consisted of immobilizing the designated forward primer onto an Au electrode surface to create the sensing probe, followed by PCR temperature cycling in blank, positive, and negative DNA controls. Target DNA identification was possible by monitoring changes in the system's charge transfer resistance values (R ct ) before and after PCR treatment through electrochemical impedance spectroscopy (EIS) analysis. Custom-made, flexible gold electrodes were fabricated using chemical etching optical lithography. A PCR cycle study determined the optimum conditions to be at 6 cycles providing fast results while maintaining a good sensitivity. EIS data for the DNA recognition process demonstrated the successful distinction between target interaction resulting in an increase in resistance to charge transfer (R ct ) percentage change of 176% for the positive DNA control vs. 21% and 20% for the negative and non-DNA-containing controls, respectively. Results showed effective fabrication of a fast, PCR-based electrochemical biosensor for the detection of pks genomic island with a calculated limit of detection of 17 ng/μL.

An Impedimetric Biosensing Strategy Based on BicyclicPeptides as Bioreceptors for Monitoring h-uPA Cancer Biomarkers

“…The impedimetric platform responses show a good linearity (R 2 > 0.9997). The LOD calculated as the ratio between three times the standard deviation of the blank and the curve slope [63] is equal to 0.009 µg mL −1 with a LOQ of 0.030 µg mL −1 . These values suggest that this platform has a higher sensitivity than the voltammetric one in simple matrices spiked with h-uPA.…”