A simple and highly-efficient approach to monitor the expression of
P-glycoprotein (P-gp) in cells was urgently needed to demonstrate the drug resistance of
cancer cells. Herein, a competitive method-based electrochemiluminescent (ECL) assay with a single ECL
indicator was proposed for the first time to efficiently estimate the concentration ratio of two
proteins. By converting the different
proteins to partially coincident nucleotide sequences via a sandwich type immunoassay on magnetic beads, the concentration ratio related ECL signals could be obtained via competitive
nucleotide hybridization on an
electrode surface. This method could thoroughly overcome the limitations of simultaneous ECL assays via multiple ECL indicators with inevitable cross reactions. At the same time, rolling circle amplification was employed to improve the detection performances, especially the detection limit and sensitivity. With P-gp and
glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as a model, the proposed ECL assay was successfully employed to monitor the drug resistance of
cancer cells. Compared with conventional technologies, improved sensitivity and accuracy were achieved with a correlation coefficient of 0.9928 and a detection limit of 0.52%. Success in the establishment of the competitive method-based ECL assay offered an efficient strategy to demonstrate the concentration ratio of two
proteins and a potential approach for detecting other
proteins and nucleotide sequences, revealing a new avenue for ultrasensitive biomolecule diagnostics, especially in cell function research.