Phalloidin is a bicyclic peptide that belongs to a family of toxins isolated from the deadly Amanita phalloides “death cap” mushroom and is commonly used in imaging applications to selectively label F-actin in fixed cells, permeabilized cells, and cell-free experiments. Alexa Fluor® 488 phalloidin is one of the most popular and brightest fluorescent phalloidin conjugates. Conjugated with our bright, photostable, green-fluorescent Alexa Fluor® 488 dye, Alexa Fluor® 488 phalloidin clearly outperforms fluorescein phalloidin in brightness and photostability in a side-by-side comparison (Figure 1).
||Figure 1. Photobleaching comparison of fluorescein phalloidin and Alexa Fluor® 488 phalloidin. The cytoskeleton of bovine pulmonary artery endothelial cells (BPAEC) was labeled with (top series) Alexa Fluor® 488 phalloidin and mouse monoclonal anti–alpha-tubulin antibody in combination with Alexa Fluor® 546 goat anti–mouse IgG antibody or (bottom series) fluorescein phalloidin and the anti–a-tubulin antibody in combination with a commercially available Cy3 goat anti–mouse IgG antibody. The pseudocolored images were taken at 30-second intervals (0, 30, 90, and 210 seconds of exposure). The images were acquired with bandpass filter sets appropriate for fluorescein and rhodamine. Even after only 30 seconds of illumination, the fluorescein phalloidin fluorescence was significantly diminished whereas the Alexa Fluor® 488 phalloidin fluorescence was basically unchanged after 210 seconds of exposure.||