BV-1

Guidelines (The following is a recommended experimental protocol for guidance only; please adjust it according to your specific needs)

1. Stock Solution Preparation
1.1 Solvent: Ethanol or DMSO.
1.2 Recommended concentration: 1 × 10^-3 M (1 mM).

2. Working Solution Preparation
2.1 Diluent: Cell culture medium.
2.2 Working concentration: 5 μM.
2.3 Notes: Adjust the working solution concentration as required; prepare and use immediately.

3. Staining Procedures
3.1 Applicable to adherent cells:
3.1.1 Sample type: Adherent cells (A549); seed the cells in a culture plate before staining; trypsinization is not required for staining.
3.1.2 Incubation conditions: Incubate at 37°C for 30 min; for A549 cells, incubate in a humidified environment containing 5% CO₂; no need for light protection is mentioned in literature.
3.1.3 Washing step: Wash 3 times with phosphate-buffered saline (PBS) after incubation.

4. Control Setup
4.1 Unstained control: Used to adjust instrument voltage and eliminate autofluorescence of cells.
4.2 Blank control: Only add BV-1 without drug treatment, used to exclude fluorescence changes caused by the dye itself.

5. Detection and Analysis
5.1 Instrument type: Laser confocal microscope, confocal fluorescence microscope.
5.2 Excitation/emission wavelength: Ex = 488 nm; Em = 525-650 nm.
5.3 Result analysis:
5.3.1 Changes in fluorescence intensity: The higher the environmental viscosity, the stronger the fluorescence intensity; a time-dependent fluorescence enhancement can be observed during the increase of cellular viscosity.
5.3.2 Fluorescence localization: The fluorescence of BV-1 localizes to mitochondria, showing red fluorescence; the fluorescence is weak at low viscosity, while it increases significantly with an emission redshift at high viscosity.

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