期刊
ADVANCED BIOSYSTEMS
卷 4, 期 12, 页码 -出版社
WILEY-V C H VERLAG GMBH
DOI: 10.1002/adbi.201900307
关键词
droplet digital PCR; extracellular vesicles; microfluidics
资金
- NIH [PO1 CA069246, RO1 CA204019, R21 CA236561]
- Schmidt Science Fellow program
- Rhodes Trust, Oxford, UK
- QuantBio graduate student award at Harvard
There is a need for novel analytical techniques to study the composition of single extracellular vesicles (EV). Such techniques are required to improve the understanding of heterogeneous EV populations, to allow identification of unique subpopulations, and to enable earlier and more sensitive disease detection. Because of the small size of EV and their low protein content, ultrahigh sensitivity technologies are required. Here, an immuno-droplet digital polymerase chain reaction (iddPCR) amplification method is described that allows multiplexed single EV protein profiling. Antibody-DNA conjugates are used to label EV, followed by stochastic microfluidic incorporation of single EV into droplets. In situ PCR with fluorescent reporter probes converts and amplifies the barcode signal for subsequent read-out by droplet imaging. In these proof-of-principle studies, it is shown that multiplex protein analysis is possible in single EV, opening the door for future analyses.
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