An integrated and multifunctional homemade cell sensor platform based on Si-d-CQDs and CRISPR-Cas12a for CD31 detection during endothelial-to-mesenchymal transition.

Endothelial-to-mesenchymal transition (EndMT) plays a crucial role in the initiation and progression of atherosclerosis and various disease processes. Cluster of differentiation 31 (CD31) is a significant marker in EndMT. Detecting CD31 is essential for early-stage monitoring of EndMT and diagnosing atherosclerosis. Herein, we propose a homemade electrochemical array sensor comprising four electrodes, applied for cell cultivation, electrical stimulation, and simultaneous electrochemical detection, offering a three-in-one approach for CD31 detection during EndMT. To enhance the analytical performance of the cell sensor, indium tin oxide/chitosan-MXene/polyaniline (ITO/CS-MXene/PANI) composites were synthesized. The synthesis process involved the polymerization of PANI on the surface of the CS-MXene-modified ITO electrode, resulting in the creation of highly biocompatible active sites for cell immobilization. Si-d-CQDs@acDNA-AptCD31-Fc, with exceptional photophysical and chemical properties, was integrated into the array sensor setup, which enabled the dual-mode detection of fluorescent and electrochemical signals in cultured cells. A CRISPR-Cas12a system was employed to cleave Si-d-CQDs@acDNA-AptCD31-Fc. Subsequently, the fragmented Fc molecules were enriched via electrochemistry to further amplify the electrochemical signals. Through the unique combination of programmable Si-d-CQDs@acDNA-AptCD31-Fc, the CRISPR-Cas12a system, and voltage enrichment, a novel "signal-on-off-super on" signal amplification strategy was developed. The cell sensor exhibited a wide linear range from 1 × 101 particles mL-1 to 1 × 106 particles mL-1 (R2 = 0.9912) and a detection limit of 4 particles mL-1. The proposed strategy presents a promising approach for developing CRISPR-Cas12a-based tools for detecting various stages of EndMT and opens a new window for dual-mode analysis applications.