Proximity ligation assay mediated rolling circle amplification strategy for in situ amplified imaging of glycosylated PD-L1

Abstract

Glycosylated PD-L1 is a more reliable biomarker for immune checkpoint therapy and plays important roles in tumor immunity. Glycosylation of PD-L1 hinders antibody-based detection, which is partially responsible for the inconsistency between PD-L1 immunohistochemical results and therapeutic treatment response. Herein, we present a proximity ligation assay mediated rolling circle amplification (PLA-RCA) strategy for amplified imaging of glycosylated PD-L1 in situ. The strategy relies on a pair of DNA probes: an aptamer probe to specifically recognize cellular surface protein PD-L1 and a glycan conversion (GC) probe for metabolic glycan labeling. Upon proximity ligation of sequence binding to the two probes, the proximity ligation-triggered RCA occurs. The feasibility of the as-proposed strategy has been validated as it realized the visualization of PD-L1 glycosylation in different cancer cells and the monitoring of the variation of PD-L1 glycosylation during drug treatment. Thus, we envision the present work offers a useful alternative to track protein-specific glycosylation and potentially advances the investigation of the dynamic glycan state associated with the disease process.

Keywords: Accurate imaging analysis; Glycosylated PD-L1; Metabolic glycan labeling; Proximity ligation assay; Rolling circle amplification.