ortho-iodoHoechst 33258 

ortho-iodoHoechst 33258 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].

Guide (The following is our recommended experimental protocol. This protocol is for reference only and should be adjusted according to your specific needs).
Preparation of Hoechst working solution
1.1 Preparation of stock solution
Prepare a 1 mg/mL Hoechst stock solution in DMSO.
Note: It is recommended to aliquot the Hoechst stock solution and store at -4°C or -20°C protected from light.
1.2 Preparation of working solution
Dilute the stock solution with pre-warmed serum-free cell culture medium or PBS to obtain a final concentration of 10 μg/mL Hoechst working solution.
Note: Adjust the concentration of Hoechst working solution according to actual needs and prepare fresh before use.
2. Cell staining (suspension cells)
2.1 Collect cells by centrifugation and wash twice with PBS, 5 min each time. Cell density: 1×10⁶/mL.
2.2 Add 1 mL Hoechst working solution and incubate at room temperature for 3-10 min.
2.3 Centrifuge at 400 g for 3-4 min and discard the supernatant.
2.4 Wash cells twice with PBS, 5 min each time.
2.5 Resuspend cells in 1 mL serum-free medium or PBS and observe using a fluorescence microscope or flow cytometer.
3. Cell staining (adherent cells)
3.1 Culture adherent cells on sterile coverslips.
3.2 Remove the coverslips from the culture medium and aspirate excess medium.
3.3 Add 100 μL dye working solution, gently shake to fully cover the cells, and incubate for 3-10 min.
3.4 Remove the dye working solution, wash 2-3 times with culture medium (5 min each), and observe using a fluorescence microscope or flow cytometer.


Notes
1. Adjust the concentration of Hoechst working solution according to actual needs.
2. This product is for research use only by professionals and is not intended for clinical diagnosis or treatment, nor for use in food or drugs.
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.

534.39

C₂₅H₂₃IN₆

158013-41-3

Solid

Pink to red

451

356

CN1CCN(C2=CC=C3N=C(C4=CC=C5N=C(C6=CC=CC=C6I)NC5=C4)NC3=C2)CC1

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

  [2]. 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]

  [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]