RhoVR 1

Guidelines
(The recommended experimental protocol below is for guidance only and needs to be adjusted according to your specific requirements)^[1].

1. Working Solution Preparation
1.2 Concentration recommendations: 150 nM; 250 nM.

2. Staining Procedure
2.1 Sample types: Adherent cells (HEK 293T, COS7, MCF-7, HeLa).
2.2 Incubation conditions: Incubate with 150 nM RhoVR 1; or incubate with 250 nM RhoVR 1.
2.3 Washing step: After treatment with carbonyl cyanide-p-trifluoromethoxyphenylhydrazone, wash with HBSS.

3. Control Setup
3.1 Negative control: Use SPIRIT RhoVR 0 (lacking voltage-sensitive aniline group).
3.2 Vehicle control: Use ethanol as the solvent for carbonyl cyanide-p-trifluoromethoxyphenylhydrazone treatment.

4. Detection and Analysis
4.1 Instruments: Confocal fluorescence microscope; Wide-field fluorescence microscope.
4.2 Result analysis:
4.2.1 Fluorescence intensity change: Fluorescence intensity decreases upon mitochondrial depolarization (induced by carbonyl cyanide-p-trifluoromethoxyphenylhydrazone or antimycin A); fluorescence intensity returns to baseline after elution of carbonyl cyanide-p-trifluoromethoxyphenylhydrazone.
4.2.2 Fluorescence localization: Localizes to mitochondria; remains in mitochondria after depolarization.
4.2.3 Color change: Tetramethylrhodamine-type fluorescence (appears yellow in multicolor imaging).