bc0c00216_si_001.pdf (598.04 kB)
In Situ One-Step Fluorescence Labeling Strategy of Exosomes via Bioorthogonal Click Chemistry for Real-Time Exosome Tracking In Vitro and In Vivo
journal contribution
posted on 2020-05-11, 19:35 authored by Sukyung Song, Man Kyu Shim, Seungho Lim, Yujeong Moon, Suah Yang, Jinseong Kim, Yeonsun Hong, Hong Yeol Yoon, In-San Kim, Kwang Yeon Hwang, Kwangmeyung KimExosomes
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.