Flubida-2 

Flubida-2 is a cell permeable dye which can be hydrolyzed to Fubi-2 by endoesterases in cells (after hydrolysis, Ex=492 nm, Em=517 nm). Flubida-2 can be used to detect pH at a specific site in a cell organelle by directing the probe to where avidin fusion proteins are located^[1].

Guidelines (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs)^[1].
1. Dissolve Flubida-2 in DMSO (approximately 2 mM).
2. Mix the stock solution 1: 1 with 20% (w/v in DMSO) Pluronic F-127 (Molecular Probes), and dilute to the desired final concentration (2-4 μM) with serum-free DMEM.
3. 30 to 48 h post-transfection with either AV-KDEL or ST-AV DNA, HeLa cells are rinsed once with serum-free DMEM and loaded with 2-4 μM Flubida-2 for 3-5 h (or overnight for 10-15 h).
4. Chase the labeled cells with normal growth medium for at least 2 h, to allow excess dye-biotin to exit from the cytosol.
5. The strong avidin-biotin interaction ensures stable, specific avidin-Flubi- binding that resisted washing. Biotin starvation of the cells is not necessary before Flubida- loading, as staining was bright and stable.

MCE has not independently confirmed the accuracy of these methods. They are for reference only.

812.93

C₈₆H₉₆N₈O₂₀S₂

517

492

Room temperature in continental US; may vary elsewhere.

Please store the product under the recommended conditions in the Certificate of Analysis.

• [1]. M M Wu, et al. Studying organelle physiology with fusion protein-targeted avidin and fluorescent biotin conjugates. Methods Enzymol. 2000;327:546-64.  [Content Brief]