Extracellular vesicle (EV)/exosome secretion is a dynamic process that tunes the cellular communication for response to internal and external cues. The selective enrichment of a newly synthesized EV/exosome has been hindered by the basic fact that all EVs/exosomes, new and old, share the similar inherent parameters and thus are indistinguishable.
Researchers at Xiamen University have developed a method by cotranslational introduction of azide groups into EV/exosome proteins as a timestamp and label them with biotin tag by click chemistry, to separate the newly synthesized EVs/exosomes from preexisting populations by streptavidin-modified herringbone microfluidic chip. For mouse model of anti-PD-L1 immunotherapy, the level of newly synthesized PD-L1+ EVs detected by the developed approach was superior to the total PD-L1+ EVs from mixed time sources (quantified by classical method) for tumor progression. This method makes it possible to address the temporal characteristics of newly synthesized EVs/exosomes in cell and in vivo, for studying EV/exosome secretion to respond to specific stimuli.
Schematic illustrating the strategy to study dynamic changes of newly synthesized EVs in vivo after immunotherapy based on metabolic labeling
Mice were subcutaneously injected with 4T1 cells, and then every week treated with PD-L1 antibody once or injected with PBS as a control, simultaneously injection Ac4ManNAz for unnatural glucose metabolism labeling for three consecutive days. At this time, the azide-labeled EVs in the mice were newly synthesized. After reacting with DBCO-PEG4-Biotin linker, the newly synthesized EVs with biotin labeled can be captured by streptavidin (SA)-modified chips.