Hoechst 33258 analog 6 

Hoechst 33258 analog 6 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 (Ex/Em = 356/451 nm)^[1].

Guidelines (The following is our recommended protocol. This protocol only provides guidance and should be modified according to your specific needs).
Preparation of Hoechst working solution
1.1 Preparation of stock solution
Dissolve 10 mg in 5 mL DMSO.
Note: It is recommended to store the stock solution at 4°C or -20°C, protected from light, and avoid repeated freeze-thaw cycles.
1.2 Preparation of Hoechst working solution
Dilute the stock solution in serum-free cell culture medium or PBS to obtain a Hoechst working solution with a final concentration of 10 μg/mL.
Note: Please adjust the concentration of the Hoechst working solution according to actual conditions.
Cell staining
2.1 Suspension cells (6-well plate)
a. Centrifuge at 1000 g at 4°C for 3-5 minutes, then discard the supernatant. Wash twice with PBS for 5 minutes each time. Adjust the cell density to 1×10⁶/mL.
b. Add 1 mL working solution and incubate at room temperature for 3-10 minutes.
c. Centrifuge at 400 g at 4°C for 3-4 minutes and discard the supernatant.
d. Wash twice with PBS for 5 minutes each time.
e. Resuspend the cells in serum-free cell culture medium or PBS. Observe using a fluorescence microscope or flow cytometry.
2.2 Adherent cells
a. Culture adherent cells on sterile coverslips.
b. Remove the coverslips from the culture medium and aspirate excess medium.
c. Add 100 μL working solution, gently shake to fully cover the cells, and incubate at room temperature for 3-10 minutes.
d. Wash twice with culture medium for 5 minutes each time. Observe using a fluorescence microscope or flow cytometer.
Notes
1. Please adjust the concentration of the Hoechst working solution according to actual conditions.
2. This product is for R&D use only and is not intended for use in drugs, household, or other purposes.
3. For your safety and health, please wear a lab coat and disposable gloves during operation.

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

536.71

C₃₃H₄₀N₆O

129244-66-2

Solid

Off-white to pink

451

356

OC1=C(C(C)(C)C)C=C(C2=NC3=CC=C(C4=NC5=CC=C(N6CCN(C)CC6)C=C5N4)C=C3N2)C=C1C(C)(C)C

Room temperature in continental US; may vary elsewhere.

4°C, protect from light

*In solvent : -80°C, 6 months; -20°C, 1 month (protect from light)

* 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 (protect from light). When stored at -80°C, please use it within 6 months. When stored at -20°C, please use it within 1 month.

• [1]. Latt SA, Stetten G, Juergens LA, Recent developments in the detection of deoxyribonucleic acid synthesis by 33258 Hoechst fluorescence. The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society 23 (7): 493-505.  [Content Brief]

  [2]. a b c "Hoechst Stains". Invitrogren (Molecular Probes).

  [3]. Portugal J, Waring MJ. Assignment of DNA binding sites for 4',6-diamidine-2-phenylindole and bisbenzimide (Hoechst 33258). A comparative footprinting study. Biochimica et Biophysica Acta 949 (2): 158-68.  [Content Brief]