Drawing on the linker architectures we have previously deployed for in vivo mul=plexed imaging , we synthesized AZ647-iTCO-NHS for an=body labeling using fluorescent AZDye 647 (AZ647). In parallel, we also constructed a highly water-soluble Pyr2(PEG24)3 conjugate for facile intravenous delivery and op=mal extracellular reac=vity (Supplementary Fig. ). Intravascular click-to-release results in cleavage of AZ647 from the surface of the circula=ng cells, making RBC brightness a direct readout of the overall reac=on kine=cs. Our mouse imaging pla‚orm also uses fluorescent dextran for primary iden=fica=on of the vascular tree and neutrophil staining (α-Ly6G) to assist visualiza=on of intravenous flow during =me-lapse imaging (Fig. ). Simultaneous staining of the RBC popula=on in the shared phycoerythrin (PE) reference channel provides a direct control to validate the α-TER119 iTCO probe and confirm bioorthogonal cleavage specificity.