Hoechst 33258 (trihydrochloride) [23491-45-4]

Hoechst 33258 trihydrochloride is a marker dye in Hoechst series. Hoechst is A live nuclear marker dye. Hoechst binds to the grooves in the DNA double strand, which tends to be A/ T-rich DNA strand. Although it binds to all nucleic acids, the A/ T-rich double strand DNA significantly enhances fluorescence intensity Therefore,Hoechst dye can be used for living cell labeling. The fluorescence intensity of Hoechst dye increases with the increase of pH of solution^[1].

IC50: 51.31±4.56 μM (HeLa cell), 32.43±3.27 μM (HL60 cell), 15.42±2.16 μM (U937 cell)^[1]

Preparation of Hoechst working solution
1.1 Preparation of stock solution
Take 10 mg and dissolve in 5 mL DMSO
Note: It is recommended that the stock solution be stored at 4°C or - Store at 20°C in the dark and avoid repeated freezing and thawing.
1.2 Preparation of Hoechst working solution
The diluted stock solution was dissolved in serum-free cell culture medium or PBS to obtain Hoechst working solution with a final concentration of 10 μg/mL.
Note: Please adjust the concentration of Hoechst working solution according to the actual situation.
1. Cell staining
2.1 Suspension cells (6-well plate)
a. Centrifuge at 1000 g for 3-5 min at 4°C and discard the supernatant. Wash 2 times with PBS, 5 minutes each. The cell density was 1×10⁶/mL.
b. Add 1 mL of working solution and incubate at room temperature for 3-10 minutes.
c. Centrifuge at 400 g for 3-4 minutes at 4°C and discard the supernatant.
d. Wash 2 times with PBS, 5 min each.
e. Resuspend cells in serum-free cell culture medium or PBS. Fluorescence microscopy or flow cytometry observation.
2.2 Adherent cells
a. Adherent cells were cultured on sterile coverslips.
b. Remove the coverslip from the medium and aspirate excess medium. c. Add 100 μL of working solution, shake gently to completely cover the cells, and incubate at room temperature for 3-10 minutes.
d. Wash 2 times with medium, 5 min each. Fluorescence microscope or flow cytometry observation.

Store
4°C, 1 year. Avoid light

Precautions
1. Please adjust the concentration of Hoechst working solution according to the actual situation.
2. This product is for R&D use only, not for medicine, household or other purposes.
3. For your safety and health, please wear a lab coat and disposable gloves for operation.

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

* Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 6 months; -20°C, 1 month (sealed storage, away from moisture and light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

* Note: If you choose water as the stock solution, please dilute it to the working solution, then filter and sterilize it with a 0.22 μm filter before use.

• [1]. Wang XJ, et al. Newly synthesized bis-benzimidazole derivatives exerting anti-tumor activity through inductionof apoptosis and autophagy. Bioorg Med Chem Lett. 2012 Oct 1;22(19):6297-300.  [Content Brief]

  [2]. Stolić I, et al. Synthesis, DNA/RNA affinity and antitumour activity of new aromatic diamidines linked by 3,4-ethylenedioxythiophene. Eur J Med Chem. 2011 Feb;46(2):743-55.  [Content Brief]