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. Author manuscript; available in PMC: 2011 Jun 15.
Published in final edited form as: Biotechniques. 2011 Feb;50(2):98–115. doi: 10.2144/000113610

Figure 2. Rhod123 used in quenching mode.

Figure 2

11–14-day-old primary rat cortical neurons were loaded for 30 min at 37°C with 10 μM Rhod123 in normal growth media as described (45). The Rhod123-containing loading medium was removed, and the cultures placed in CO2-insensitive, pH-stable Leibovitz’s L-15 medium (Invitrogen Life Technologies) and incubated at 37°C in room air for 15 min to allow for the Rhod123 fluorescent signal to reach steady state. Using a Chroma HQ560/55x:Q595LP:HQ645/75m filter and dichroic set, CCD images were captured 1/min for 5 min to establish a baseline. After 5 min, oligomycin or FCCP was added at the times indicated by the arrows, which respectively caused the characteristic quenching (hyperpolarization of Δψm) and unquenching (depolarization of Δψm) responses in Fwc as expected. Average traces of relative Fwc were taken from n = 4 neurons. This figure and legend is modified from Figure 1C in Reference 45, with permission.