Exosomes are cellular components with promising uses in
cancer diagnostics and
therapeutics, and their imaging and tracking are essential to study their
biological properties. Herein, we report on an in situ one-step fluorescence labeling strategy for exosomes via bioorthogonal click chemistry. First, exosome donor
cancer cells were treated with tetraacetylated N-azidoacetyl-
d-mannosamine (
Ac4ManNAz) to generate unnatural
azide groups (-N3) on their surface via metabolic glycoengineering. Then, the
azide groups were labeled with near-infrared
fluorescent dye-conjugated
dibenzylcyclooctyne (DBCO-Cy5) via bioorthogonal click chemistry. After 2 days of incubation, the DBCO-Cy5-labeled exosomes (Cy5-Exo) were successfully secreted from the donor
cancer cells and were isolated via classical ultracentrifugation, providing a high-yield of
fluorescent dye-labeled exosomes. This in situ one-step bioorthogonal click chemistry offers improved labeling efficiency, biocompatibility, and imaging sensitivy compared to standard exosomes (ST-Exo), purified with classical ultracentrifugation or
carbocyanine lipophilic
dye (DiD)-labeled exosomes (DiD-Exo) in vitro. In particular, the Cy5-Exo were successfully taken up by A549 cells in a time-dependent manner, and they could escape from lysosome confinement, showing their possible use as a delivery carrier of therapeutic drugs or imaging agents. Finally, intraveneously injected Cy5-Exo were noninvasively tracked and imaged via near-infrared fluorescence (NIRF) imaging in
tumor-bearing mice. This new fluorescence labeling strategy for natural exosomes may be useful to provide better understanding of their
theranostic effects in many biomedical applications.