Ru(bpy)₃²⁺‐Enabled Cell‐Surface Photocatalytic Proximity Labeling toward More Efficient Capture of Physically Interacting Cells

Abstract

Intercellular proximity labeling has emerged as a promising approach to enable the study of cell‐cell interactions (CCIs), but the efficiency of current platforms is limited. Here, we use Ru(bpy)32+ to construct an efficient photocatalytic proximity labeling (PPL) system on the cell surface that allows the highly discriminative CCI detection with spatiotemporal resolution. Through the mechanism study and quantitative characterization on living cells, we demonstrate that the singlet‐oxygen (1O2) mechanism is more efficient and specific than the single electron transfer (SET) mechanism in Ru‐mediated PPL. Ru(bpy)32+ catalysts with different cell‐anchoring moieties are prepared to facilitate the catalyst loading on primary cells. Finally, based on this system, we develop a “live” T cell receptor (TCR) multimer with TCR‐T cells that could sensitively identify and discriminate cells presenting antigens of different affinity, providing a powerful tool to better understand the heterogeneity of antigen presenting cells.

Painting cell-cell interactions by horseradish peroxidase and endogenously generated hydrogen peroxide

“…Since HRP showed no toxicity issue in their expression in live animal model in these studies [28, 33] , we expect that our cell-cell interaction labeling technique can be employed in these HRP-expressed animal models. We believe that HRP-mediated proximal cell labeling has advantages in overcoming the limitations of other approaches, such as utilization of high molecular weight of probe (LIPSTIC, EXCELL, PUP-IT, FucoID) [9, 10, 11, 12] , light irradiation (μMap, PhoTag, PhoXCELL, Ru- 1 O 2 -hydrazide) [15, 16, 17, 18] , low concentration of extracellular ATP (TurboID) [37] , and exogenous treatment of H 2 O 2 (APEX2).…”

“…Oslund et al have designed riboflavin tetraacetate (RFT) mediated labeling of proteins, called photocatalytic cell-tagging (PhoTag) [16] . Lie et al reported antigen-specific T cell detection using the photosensitizer dibromofluorescein (DBF) termed PhoXCELL [17] , and Qiu et al developed Ru- 1 O 2 -hydrazide system for photocatalytic cell labeling [18] . However, light irradiation for photocatalytic reactions critically limits their use to in vitro or cell-level applications rather than in vivo .…”