Protocol for Taxol Janelia Fluor® 549
This protocol is intended as a guide only, for full experimental details please read the reference provided.
Introduction
Taxol (often referred to as Paclitaxel) is a diterpenoid that binds to tubulin. It promotes and stabilizes tubulin polymerization, which accounts for its anti-mitotic and cytotoxic action. The conjugation of a fluorescent dye to taxol offers a convenient way to image the microtubule cytoskeleton of a cell; well known fluorescent probes based on this principle are: flutax 1 (Catalog # 2226) and flutax 2 (Catalog # 6254).
Taxol Janelia Fluor® 549 is a tubulin fluorescent probe that incorporates the bright photostable yellow dye, Janelia Fluor® 549 (Catalog # 6147). The following protocol provides guidelines for using this product.
Protocol
- Prepare a stock solution of Taxol Janelia Fluor® 549 in DMSO. Stock solutions can be aliquoted and stored for up to 1 month at ≤-20°C.
- Dilute the stock solution into warm media (37°C) and place into a pre-warmed box (for storage in the incubator). Aqueous working solutions should be prepared and used on the same day.
- Apply to live cells at a concentration of 3 μM. Lower concentrations can be used and the concentration should be optimized for individual experiments.
- Incubate for 1 hour at 37°C
- Rinse the cells three times with 1x PBS and 2% BSA and apply fresh media prior to imaging.
- Image cells using appropriate filters for Taxol Janelia Fluor® 549. Excitation maximum = 556 nm; emission maximum = 575 nm.