|Always add your plasmid to the buffer before adding TransIT
EB3 microtubule tracking
To visualize live-cell microtubule dynamics, cells were transfected with mApple-EB3, a gift from Michael Davidson (Addgene plasmid #54892), using the TransIT-LT1 transfection reagent (Mirus Bio) per manufacturer's instructions. Cells were imaged with a Nikon Eclipse Ti inverted epifluorescent microscope, maintained at 37°C and 5% carbon dioxide throughout the experiment using an In Vivo Scientific environmental cell chamber and Bioscience Tools CO2 controller. A Photometrics QuantEM CCD camera (Princeton Instruments) was used to minimize exposure time while imaging with a Nikon CFI Apochromat TIRF 100X oil-immersion lens. Videos were captured at 1 Hz for 2 minutes and quantified using the open source u-track software as described27,67. Full paper
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