CRISPR/Cas12a Integrated with a Microfluidic System Enhanced Analysis of Programmed Cell Death Ligand 1 Expression in
Bingjie Zeng1, Anzhi Sheng2, Xiao Zhang3
1Department of Clinical Laboratory, Shanghai Chest Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200030, China.
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The detection of programmed cell death ligand 1 (PD-L1) positive circulating tumor cells (CTCs) in peripheral blood has significant clinical value for predicting and evaluating the efficacy of immunotherapy in patients with non-small cell lung cancer (NSCLC). However, traditional methods remain limited by low sensitivity and the precise quantification remains a challenge. A dual-mode microfluidic analysis chip was constructed here that included clustered regularly interspaced short palindromic repeats/Cas12a quantification and immunofluorescence visualization. Quantification of the PD-L1 protein on the surface of CTCs (20 to 107 cell/mL) was achieved selectively and sensitively by amplifying the nucleic acid target to generate a strong fluorescent signal, even with very low levels of target cells. The system effectively detected PD-L1+ CTCs expression in peripheral blood samples from patients with NSCLC and monitored the efficacy of PD-1/PD-L1 targeted immune checkpoint inhibitors in real time. It exhibited excellent performance for clinical applications in monitoring the prognosis in patients with NSCLC.
