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Blinking fluorescent probes for tubulin nanoscopy in living and fixed cells

Gerasimaite, R.; Bucevicius, J.; Kiszka, K. A.; Kostiuk, G.; Koenen, T.; Lukinavicius, G.

2021-06-02 cell biology
10.1101/2021.06.01.446685 bioRxiv
Show abstract

Here we report a small molecule probe for single molecule localisation microscopy (SMLM) of tubulin in living and fixed cells. We explored a series of constructs composed of taxanes and spontaneously blinking far-red dye hydroxymethyl silicon-rhodamine (HMSiR). We found that the linker length profoundly affects the probe permeability and off-targeting. The best performing probe, HMSiR-tubulin, is composed of cabazitaxel and 6-regioisomer of HMSiR bridged by a C6 linker. Microtubule diameters of [≤]50 nm can be routinely measured in SMLM experiments on living and fixed cells. HMSiR-tubulin also performs well in 3D stimulated emission depletion (STED) microscopy, allowing a complementary use of both nanoscopy methods for investigating microtubule functions in living cells.

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