Chemistry

Streptolysin

Bacterial protein

Fluorescence

Physics

Cell

Fluorescence microscope

Gene

Optics

Biophysics

Cytoplasm

Transfection

Fluorescence-lifetime imaging microscopy

Quantum mechanics

Cell biology

Membrane

Biology

Organelle

Microscopy

Live cell imaging

Biochemistry

eLife

Site-specific fluorescent labeling of proteins inside live mammalian cells has been achieved by employing Streptolysin O, a bacterial toxin which forms temporary pores in the membrane and allows delivery of virtually any fluorescent probes, ranging from labeled IgG’s to small ligands, with high efficiency (&gt;85% of cells). The whole process, including recovery, takes 30 min, and the cell is ready to be imaged immediately. A variety of cell viability tests were performed after treatment with SLO to ensure that the cells have intact membranes, are able to divide, respond normally to signaling molecules, and maintains healthy organelle morphology. When combined with Oxyrase, a cell-friendly photostabilizer, a ~20x improvement in fluorescence photostability is achieved. By adding in glutathione, fluorophores are made to blink, enabling super-resolution fluorescence with 20–30 nm resolution over a long time (~30 min) under continuous illumination. Example applications in conventional and super-resolution imaging...

Labeling proteins inside living cells using external fluorophores for microscopy