Cell experiment:

Prior to the fluorescence measurements, cells are incubated in KRH (Krebs-Ringer-HEPES) buffer containing with 100 nM DiBAC4(3) for 20 min at room temperature. The stained cells are used for experiments without washing. The fluorescence emission is collected using a 505 nm dicroic mirror and a BA filter (>520 nm)[1].

DiBAC4(3) is a voltage-sensitive fluorescent dye (λex=490 nm, λem=505 nm).

The membrane hyperpolarization induced by 10 μM Evans blue (EB) in HEKBKα is clearly observed with DiBAC4(3), while the change in membrane potential (MP) by addition of 3 mM tetraethylammonium chloride (TEA) appears more slowly than that measured with microelectrode. The time to peak of hyperpolarization is 2.3±0.9 s (n=4) and 35.0±2.6 s (n=12, P<0.01) by the measurements with microelectrodes and DiBAC4(3), respectively[1].

[1]. Yamada A, et al. Usefulness and limitation of DiBAC4(3), a voltage-sensitive fluorescent dye, for the measurement of membrane potentials regulated by recombinant large conductance Ca2+-activated K+ channels in HEK293 cells. Jpn J Pharmacol. 2001 Jul;86(3):342-50.