Comparative analysis of taxol-derived fluorescent probes to assess microtubule networks in a complex live...

Drosophila oogenesis is an excellent in vivo model for investigating cytoskeletal dynamics because of the rapid cytoskeletal remodeling that occurs at the end of stage 10; however, there are few robust tools for detecting microtubules in live complex tissues. The recent development of membrane permeable taxol-based fluorescent probes to label microtubules is significant technical progress, but the effectiveness of these probes and the potential stabilizing effects of the taxol derivative have not been well characterized in vivo. Here, we compared three commercially available taxol-derived microtubule labels to determine their efficacy and potential artifacts. We found that all three probes labeled microtubules with differences in permeability, brightness, and signal to noise ratio. Like taxol, however, all of the probes disrupted the F-actin cytoskeleton at higher concentrations. We also found that the efflux pump inhibitor, verapamil, increased the intensity of the label and modestly increased the severity of the F-actin defects. Of the three probes, Tubulin Tracker (ThermoScientific) was the most permeable and was brightest, with the highest signal to noise ratio. Furthermore, washing out the probe after a 30-min incubation significantly reduced the F-actin artifacts without compromising signal brightness.