Label-free electrochemical quantification of microRNA-375 in prostate cancer cells

Abstract

Clinical applicability of a label-free electrochemical method was investigated, where prostate cancer (CaP) biomarkers miR-107 and miR-375 were examined in two different groups of prostate cell lines, metastatic CaP cells (PC-3, DU145, and LNCaP) and immortalized prostate epithelial cells (267B1, X/267B1, and Ki/267B1). First, the expression levels of miR-107 and miR-375 in these cells were analyzed by quantitative real-time polymerase chain reaction (PCR), resulting in useful discernable expression patterns from miR-375. Choosing miR-375 as a target CaP biomarker, a label-free electrochemical analysis was performed based on the optimized conditions of 100nmol/mL miR-375 capture probe for surface immobilization and 30-minute hybridization. Standard curves were prepared both in buffer and in 100-fold diluted serum resulting in lower detection limit of 11.7aM and 12.4aM respectively. The regression equations for buffer and diluted serum were I=−15.1349 log c+250.9735 and I=−11.8179 log c+193.1639, respectively, where I is the peak current in μA and c is the concentration of miR-375 in fM. The selectivity of the sensor was confirmed by a one-base mismatch test in 3 different concentrations of miR-375 under diluted serum, 10aM, 10fM, and 10pM. Finally, miR-375 expression in the metastatic CaP cells was quantified to range between 10 and 30fM, while its expression in the prostate epithelial cells was negligible, demonstrating close agreement with the results from quantitative real-time PCR, and thereby potential viability of the electrochemical evaluation method in clinical environment.