The anti-mitotic effect upon release of CA4 was confirmed by staining with the fluorescent probe SiR-tubulin, which enables visualization of intact microtubules. While individual treatment with prodrug 20 and Tz 5 showed no effect on microtubule integrity, almost complete depletion of the fluorescence signal was observed upon in situ reaction of both compounds, analogous to direct treatment with CA4 (Fig. , see Supporting Information for individual images and further control experiments).