Ratio-pHCL-1

Guidelines (The following is a recommended experimental protocol for reference only, and adjustments are required based on your specific needs)

1. Stock Solution Preparation
1.1 Solvent: DMSO.
1.2 Suggested concentration: 1 mM.

2. Working Solution Preparation
2.1 Diluent: PBS buffer.
2.2 Working concentration: 20 μM.
2.3 Notes: Adjust the working solution concentration as needed; prepare and use immediately.

3. Staining Procedures
3.1 Sample Type Description
3.1.1 Adherent cells^[1]: A549 cells.
3.2 Incubation Conditions
3.2.1 For adherent cells: 20 μM Ratio-pHCL-1, incubation duration of 80 min.
3.3 Washing Steps
3.3.1 Wash the cells after incubation to remove excess probes/dyes.

4. Control Setup
4.1 Set up negative controls, positive controls and blank controls.
4.2 Positive controls are used to validate the experimental system; blank controls are used to exclude fluorescent interference from reagents.

5. Detection and Analysis
5.1 Instrument Type
5.1.1 Fluorescence microscope.
5.2 Excitation/Emission Wavelengths
5.2.1 Emission wavelength range: 400−900 nm; monitor emission signals at 530 nm and 580 nm.
5.3.1 Fluorescence intensity changes: Chemiluminescence emission intensity increases with elevated pH; and the luminescence intensity decreases over time.
5.3.2 Fluorescence localization: Accumulates in adherent cells and can be detected using GFP and RFP filter sets.
5.3.3 Color changes: The emission signals correspond to green and orange-red luminescence.

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