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Fig. 7 | BMC Biology

Fig. 7

From: A genetically encoded toolkit of functionalized nanobodies against fluorescent proteins for visualizing and manipulating intracellular signalling

Fig. 7

Nanobody-SNAPf fusion proteins allow labelling of RFP-tagged and GFP-tagged proteins with fluorescent O6-benzylguanine derivatives in live cells. a, b Schematics of RNb-SNAPf fusion bound to RFP, and GNb-SNAPf fusion bound to GFP, after labelling with SNAP-Cell-647-SiR (magenta circles). cf HeLa cells co-expressing RNb-SNAPf and mitochondrial TOM20-mCh (c), RNb-SNAPf and lysosomal LAMP1-mCh (d), GNb-SNAPf and TOM20-GFP (e) or GNb-SNAPf and LAMP1-GFP (f) were treated with SNAP-Cell-647-SiR (0.5 μM, 30 min at 37 °C) and imaged using TIRFM. Scale bars 10 μm (main images) or 2.5 μm (enlarged images of yellow boxed regions). Colocalization values: RNb-SNAPf + TOM20-mCh (r = 0.95 ± 0.02, n = 6 cells); RNb-SNAPf + LAMP1-mCh (r = 0.84 ± 0.06, n = 8 cells); GNb-SNAPf + TOM20-GFP (r = 0.78 ± 0.09, n = 10 cells); and GNb-SNAPf + LAMP1-GFP (r = 0.85 ± 0.10, n = 11 cells)

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