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Hoechst 33258

Technical documents: SDS CoA Datasheet

Product overview

Name Hoechst 33258
Alternative names bisBenzimide; HO 33258; Hoechst-33258
Biological description


Blue fluorescent DNA stain that is commonly used in immunofluorescent work. It is frequently used as a nuclear stain to stain nuclei. Excited by UV light. 

It is less cell permeable but slightly more water soluble than the similar DNA stain Hoechst 33342. Unlike Hoechst 33342, Hoechst 33258 is not an apoptotic inducer.

The stain is used as a substitute to DAPI and can be used on both live and fixed cells.

As with other Hoechst stains, Hoechst 33258 binds to the AT-rich regions of the minor grove in DNA.


Uses and applications

Hoechst 33258 has similar applications of use to Hoechst 33342 and is suitable for a variety of applications as there is little fluorescent overlap with other used fluorophores/fluorescent proteins that emit in the green/red range.

The stain is frequently used as a nuclear counterstain, for cell cycle analysis and to distinguish apoptotic cells.

As this stain does not induce apoptosis, it may be preferential to Hoecsht 33342 for identification and isolation of the stem cell side population.

Purity >95%
Description Blue fluorescent nuclear DNA stain. Cell permeable.
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Solubility & Handling

Storage instructions +4°C (desiccate)
Solubility overview Soluble in water (100 mM), and in DMSO (20 mM)
Important This product is for RESEARCH USE ONLY and is not intended for therapeutic or diagnostic use. Not for human or veterinary use.



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Chemical Data

Purity >95%
Chemical name Bisbenzimidazole trihydrochloride
Molecular Weight 533.88
Chemical structure Hoechst 33258  [23491-45-4] Chemical Structure
Molecular Formula C25H24N6O 3HCl
CAS Number 23491-45-4
PubChem identifier 31953
SMILES CN1CCN(CC1)C2=CC3=C(C=C2)N=C(N3)C4=CC5=C(C=C4)NC(=C6C=CC(=O)C=C6)N5.Cl.Cl.Cl
MDL number MFCD00012679

References for Hoechst 33258

References are publications that support the biological activity of the product
  • Recognition of RNA duplex by a neomycin-Hoechst 33258 conjugate.

    Willis B et al (2014) Bioorg Med Chem 22(7) : 2327-32.
  • Minor-groove binding drugs: where is the second Hoechst 33258 molecule?

    Fornander LH et al (2013) J Phys Chem B 117(19) : 5820-30.
  • Interaction of DNA minor groove binder Hoechst 33258 with bovine serum albumin.

    Ojha H et al (2009) Chem Pharm Bull (Tokyo) 57(5) : 481-6.

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