Hoechst 33342 trihydrochloride (solution) 

Hoechst 33342 trihydrochloride (solution) 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].
Solution Concentration: 10 mg/mL

General Protocol
Preparation of Hoechst working solution
The corresponding stock solution can be diluted according to the actual situation. Note that if the solvent is DMSO, the cytotoxicity of DMSO must be considered, and a solvent control should be prepared; if the solvent is pure water, the working solution needs to be filtered and sterilized before adding cells.
1.Cell staining
2.1 Suspension cells (6-well plate)
a. Centrifuge at 1000 g at 4°C for 3-5 minutes and then discard the supernatant. Wash twice with PBS, 5 minutes each time.The cell density is 1×10⁶/mL.
b. Add 1 mL of working solution, and then incubate at room temperature for 3-10 minutes.
c. Centrifuge at 400 g at 4°C for 3-4 minutes and then discard the supernatant.
d. Wash twice with PBS, 5 minutes each time.
e. Resuspend cells with serum-free cell culture medium or PBS. Observation by fluorescence microscopy or flow cytometry.
2.2 Adherent cells
a. Culture adherent cells on sterile coverslips.
b. Remove the coverslip from the medium and aspirate excess medium.
c. Add 100 μL of working solution, gently shake it to completely cover the cells,and then incubate at room temperature for 3-10 minutes.
d. Wash twice with medium, 5 minutes each time.Observation by fluorescence microscopy or flow cytometry.

Storage
Store in separate containers to avoid repeated freeze-thaw cycles. -20°C, 1 year. Protect from 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 drug, household, or other uses.
3. For your safety and health, please wear a lab coat and disposable gloves to operate.

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

561.94

C₂₇H₃₁Cl₃N₆O

875756-97-1

[H]Cl.[H]Cl.[H]Cl.CN1CCN(C2=CC=C3C(N=C(C4=CC=C5C(N=C(C6=CC=C(OCC)C=C6)N5)=C4)N3)=C2)CC1

Room temperature in continental US; may vary elsewhere.

Please store the product under the recommended conditions in the Certificate of Analysis.

• [1]. Chazotte B. Labeling nuclear DNA with hoechst 33342. Cold Spring Harb Protoc. 2011 Jan 1;2011(1):pdb.prot5557.  [Content Brief]