Multiplexed Live-Cell Visualization Of Endogenous Proteins With Nanometer Precision By Fluorobodies [article]

Alina Klein, Susanne Hank, Anika Raulf, Felicitas Tissen, Mike Heilemann, Ralph Wieneke, Robert Tampé
2017 bioRxiv   pre-print
The visualization of endogenous proteins in living cells is a major challenge. A fundamental requirement for spatiotemporally precise imaging is a minimal disturbance of protein function at high signal-to-background ratio. Current approaches for visualization of native proteins in living cells are limited by dark emitting, bulky fluorescent proteins and uncontrollable expression levels. Here, we demonstrate the labeling of endogenous proteins using nanobodies with site-specifically engineered
more » ... ight organic fluorophores, named fluorobodies. Their fast and fine-tuned intracellular transfer by microfluidic cell squeezing allowed for low background, low toxicity, and high-throughput. Multiplexed imaging of distinct cellular structures was facilitated by specific protein targeting, culminating in live-cell super-resolution imaging of protein networks. The high-throughput delivery of engineered nanobodies will open new avenues in visualizing native cellular structures with unprecedented accuracy in cell-based screens.
doi:10.1101/145698 fatcat:4uvvbylemndgxac7zzrjv3eiim