Fluorescein diacetate

• fluorescein (494/518) is formed by intracellular hydrolysis of FDA
• bacteria can hydrolyze this
• leaks from cells
• fluorescein has a relatively high rate of photobleaching
• pH-sensitive fluorescence ref (pKa ~6.4) that is significantly reduced below pH 7
• relatively broad fluorescence emission spectrum (not good for multicolor use)