Acriflavine (Acriflavinium chloride 3,6-Acridinediamine mix) is a papain-like protease (PL(pro)) inhibitor, a fluorescent dye for labeling high molecular weight RNAs, has antimicrobial and anticancer activity, inhibits SARS-CoV-2 and other β-coronaviruses, and can be used for research on leukemia. be used to study leukemia.

A novel candidate drug, Acriflavine, exhibits significant dose-dependent inhibition of A549 cell viability at low concentrations (1, 2, 5, and 10 μM)[1].

Nude mice bearing xenografts of A549 cells were intraperitoneally injected with 2 mg/kg ACF daily for six weeks. The results indicate that treatment with ACF leads to a reduction in tumor size[1].

1. RNA marker
Experimental steps:
1. Prepare solution: Dissolve Acriflavine in an appropriate solvent, such as DMSO or water, and the concentration is adjusted according to the experimental needs.
2. Label RNA: Add solution (10-100uM) to the RNA sample and incubate for a certain period of time (usually 30 minutes to 1 hour).
3. Fluorescence measurement: Use a fluorescence spectrophotometer or fluorescence microscope to detect the fluorescence signal of RNA and evaluate labeling efficiency.
2. Antibacterial experiments
Experimental steps:
1. Dissolve Acriflavine: Dissolve Acriflavine in an appropriate solvent (usually in PBS or water).
2. Treat bacteria: Contact Acriflavine solution at different concentrations with the bacteria to be tested and incubate for a certain period of time.
3. Detection of inhibitory effect: Measure the antibacterial effect by bacterial growth curve or plate counting method.
III. Anti-cancer research
Experimental steps:
1. Dissolve Acriflavine: Dissolve Acriflavine in an appropriate solvent (such as DMSO).
2. Treat tumor cells: Add different concentrations of Acriflavine solution to cultured tumor cells and incubate for a certain period of time.
3. Cell proliferation detection: Use MTT method, cloning experiments or other cell proliferation detection methods to evaluate the anti-cancer activity of Acriflavine.
IV. SARS-CoV-2 inhibition experiment
Experimental steps:
1. Dissolve Acriflavine: Prepare Acriflavine solution in an appropriate solvent.
2. Contact with the virus: Add the Acriflavine solution to the virus culture system and contact with SARS-CoV-2 at different concentrations.
3. Viral inhibition detection: Viral replication level is detected by RT-PCR, virus titer experiments or immunofluorescence analysis.
5. Papain-like protease inhibition experiment:
Experimental steps:
1. Prepare Acriflavine solution: Dissolve Acriflavine in an appropriate solvent and adjust the concentration.
2. Reaction with enzymes: Add Acriflavine to the reaction system of papain-like protease and incubate.
3. Measuring enzyme activity: Evaluate the inhibitory effect of Acriflavine on enzyme activity by enzyme activity detection methods, such as substrate cleavage experiments.

The above information is based on published literature. Experimental procedures should be appropriately modified to meet specific research demands.

CML mice are treated daily with acriflavine (8 mg/kg) or PBS via intraperitoneal injection, for 10 days starting from day 7 after bone marrow transplantation [2].

H2O: 20 mg/mL (42.65 mM), Sonication is recommended.