Lyso-OC

Guidelines (The following is a recommended operating procedure for guidance only, and adjustments should be made according to your specific requirements)
1. Stock Solution Preparation
1.1 Solvent: For most dyes, organic solvents are commonly used for dissolution, such as anhydrous DMSO.
1.2 Recommended Concentration: It is generally recommended to prepare a high-concentration stock solution of 1-10 mM.
2. Working Solution Preparation
2.1 Diluent: Serum-free medium or PBS is usually used. Proteins and esterases in serum may interfere with staining results or cause dye hydrolysis.
2.2 Working Concentration: 10 μM.
2.3 Notes: Adjust the working solution concentration as needed; prepare it fresh before use.
3. Staining Procedure
3.1 Sample Types^[1]: Adherent cells (MCF-7 cells, HeLa cells, HELF cells, CHO cells).
3.2 Incubation Conditions: Incubate cells with 10 μM Lyso-OC for 45 min.
4. Control Setup
4.1 Set up negative controls, positive controls and blank controls.
4.1.1 Use nutrient-rich culture conditions as the autophagy induction control.
4.1.2 Use starvation + 3-Methyladenine (HY-19312) (autophagy inhibitor) as the autophagy inhibition control.
5. Detection and Analysis
5.1 Instrument Type: Two-photon fluorescence confocal microscope; two-photon fluorescence microscope.
5.2 Excitation/Emission Wavelengths: Excitation light (Ex): 760 nm; emission light (Em): 490-550 nm.
5.3 Result Analysis:
5.3.1 Fluorescence intensity changes: Fluorescence intensity remains stable under nutrient-rich conditions; under starvation-induced autophagy conditions, fluorescence intensity decreases gradually over time (0-4 h); fluorescence intensity remains stable under autophagy inhibition (starvation + 3-MA) conditions.
5.3.2 Fluorescence localization: Localizes to lysosomes in living cells; localizes exclusively to lysosomes.
5.3.3 Color changes: Green fluorescence is observable in lysosomes; fluorescence weakens when lysosomes fuse with autophagosomes to form autolysosomes.

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