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`!DAPI`! (pronounced 'DAPPY', /ˈdæpiː/), or `!4′,6-diamidino-2-phenylindole`!, is a `F33f`_`[fluorescent`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Fluorescence]`_`f `F33f`_`[stain`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Staining_(biology)]`_`f that binds strongly to `F33f`_`[adenine`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Adenine]`_`f–`F33f`_`[thymine`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Thymine]`_`f-rich regions in `F33f`_`[DNA`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=DNA]`_`f. It is used extensively in `F33f`_`[fluorescence microscopy`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Fluorescence_microscopy]`_`f. As DAPI can pass through an intact `F33f`_`[cell membrane`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Cell_membrane]`_`f, it can be used to stain both live and `F33f`_`[fixed`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Fixation_(histology)]`_`f cells, though it passes through the membrane less efficiently in live cells and therefore provides a marker for membrane viability.

>>Contents

• `F0af`_`[History`#history]`_`f

• `F0af`_`[Modelling of absorption and fluorescence properties`#modelling-of-absorption-and-fluorescence-properties]`_`f

• `F0af`_`[Live cells and toxicity`#live-cells-and-toxicity]`_`f

• `F0af`_`[Alternatives`#alternatives]`_`f

• `F0af`_`[References`#references]`_`f

• `F0af`_`[See also`#see-also]`_`f

-─

>>History

DAPI was first synthesised in 1971 in the laboratory of Otto Dann as part of a search for drugs to treat `F33f`_`[trypanosomiasis`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Trypanosomiasis]`_`f. Although it was unsuccessful as a drug, further investigation indicated it bound strongly to DNA and became more fluorescent when bound. This led to its use in identifying `F33f`_`[mitochondrial`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Mitochondria]`_`f DNA in `F33f`_`[ultracentrifugation`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Ultracentrifugation]`_`f in 1975, the first recorded use of DAPI as a fluorescent DNA stain.`:cite-ref-kapuscinski1995-1-0[`F5bf`_`[1`#cite-note-kapuscinski1995-1]`_`f]

Strong fluorescence when bound to DNA led to the rapid adoption of DAPI for fluorescent staining of DNA for `F33f`_`[fluorescence microscopy`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Fluorescence_microscopy]`_`f. Its use for detecting DNA in `F33f`_`[plant`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Plant]`_`f, `F33f`_`[metazoa`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Metazoa]`_`f and `F33f`_`[bacteria`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Bacteria]`_`f cells and `F33f`_`[virus`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Virus]`_`f particles was demonstrated in the late 1970s, and quantitative staining of DNA inside cells was demonstrated in 1977. Use of DAPI as a DNA stain for `F33f`_`[flow cytometry`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Flow_cytometry]`_`f was also demonstrated around this time.`:cite-ref-kapuscinski1995-1-1[`F5bf`_`[1`#cite-note-kapuscinski1995-1]`_`f]

When bound to double-stranded DNA, DAPI has an absorption maximum at a wavelength of 358 nm (`F33f`_`[ultraviolet`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Ultraviolet]`_`f) and its emission maximum is at 461 nm (blue). Therefore, for fluorescence microscopy, DAPI is excited with ultraviolet light and is detected through a blue/cyan filter. The emission peak is fairly broad.`:cite-ref-dapi-nucleic-acid-stain-2-0[`F5bf`_`[2`#cite-note-dapi-nucleic-acid-stain-2]`_`f] DAPI will also bind to `F33f`_`[RNA`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=RNA]`_`f, though it is not as strongly fluorescent. Its emission shifts to around 500 nm when bound to RNA.`:cite-ref-3[`F5bf`_`[3`#cite-note-3]`_`f]`:cite-ref-4[`F5bf`_`[4`#cite-note-4]`_`f]

DAPI's blue emission is convenient for microscopists who wish to use multiple fluorescent stains in a single sample. There is some fluorescence overlap between DAPI and green-fluorescent molecules like `F33f`_`[fluorescein`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Fluorescein]`_`f and `F33f`_`[green fluorescent protein`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Green_fluorescent_protein]`_`f (GFP) but the effect of this is small.

Outside of analytical fluorescence light microscopy DAPI is also popular for labeling of `F33f`_`[cell cultures`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Cell_cultures]`_`f to detect the DNA of contaminating `*`F33f`_`[Mycoplasma`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Mycoplasma]`_`f`* or `F33f`_`[virus`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Virus]`_`f. The labelled `*Mycoplasma`* or virus particles in the `F33f`_`[growth medium`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Growth_medium]`_`f fluoresce once stained by DAPI making them easy to detect.`:cite-ref-5[`F5bf`_`[5`#cite-note-5]`_`f]

>>Modelling of absorption and fluorescence properties

This DNA fluorescent probe has been effectively modeled`:cite-ref-6[`F5bf`_`[6`#cite-note-6]`_`f] using the `F33f`_`[time-dependent density functional theory`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Time-dependent_density_functional_theory]`_`f, coupled with the IEF version of the `F33f`_`[polarizable continuum model`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Polarizable_continuum_model]`_`f. This quantum-mechanical modeling has rationalized the absorption and fluorescence behavior given by minor groove binding and `F33f`_`[intercalation`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Intercalation_(biochemistry)]`_`f in the DNA pocket, in term of a reduced structural flexibility and polarization.

>>Live cells and toxicity

DAPI can be used for fixed cell staining. The concentration of DAPI needed for live cell staining is generally very high; it is rarely used for live cells.`:cite-ref-7[`F5bf`_`[7`#cite-note-7]`_`f] It is labeled non-toxic in its MSDS`:cite-ref-8[`F5bf`_`[8`#cite-note-8]`_`f] and though it was not shown to have mutagenicity to `*E. coli`*,`:cite-ref-9[`F5bf`_`[9`#cite-note-9]`_`f] it is labelled as a known mutagen in manufacturer information.`:cite-ref-dapi-nucleic-acid-stain-2-1[`F5bf`_`[2`#cite-note-dapi-nucleic-acid-stain-2]`_`f] As it is a small DNA binding compound, it is likely to have some `F33f`_`[carcinogenic`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Carcinogenic]`_`f effects and care should be taken in its handling and disposal.

>>Alternatives

The `F33f`_`[Hoechst stains`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Hoechst_stain]`_`f are similar to DAPI in that they are also blue-fluorescent DNA stains which are compatible with both live- and fixed-cell applications, as well as visible using the same equipment filter settings as for DAPI.

>>References

`:cite-note-kapuscinski1995-1`!1.`! `F0af`_`[↑`#cite-ref-kapuscinski1995-1-0]`_`f `:citerefkapuscinski1995`aKapuscinski, J. (September 1995). "DAPI: a DNA-specific fluorescent probe". `*Biotech. Histochem`*. `!70`! (5): 220–233. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.3109/10520299509108199. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 8580206.

`:cite-note-dapi-nucleic-acid-stain-2`!2.`! `F0af`_`[↑`#cite-ref-dapi-nucleic-acid-stain-2-0]`_`f Invitrogen, DAPI Nucleic Acid Stain Archived 2009-03-06 at the `F33f`_`[Wayback Machine`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Wayback_Machine]`_`f. accessed 2009-12-08.

`:cite-note-3`!3.`! `F0af`_`[↑`#cite-ref-3]`_`f Scott Prahl, DAPI. accessed 2009-12-08.

`:cite-note-4`!4.`! `F0af`_`[↑`#cite-ref-4]`_`f `:citerefkapuscinski2017`aKapuscinski, J (2017). "Interactions of nucleic acids with fluorescent dyes: spectral properties of condensed complexes". `*Journal of Histochemistry & Cytochemistry`*. `!38`! (9): 1323–1329. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.1177/38.9.1696951. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 1696951.

`:cite-note-5`!5.`! `F0af`_`[↑`#cite-ref-5]`_`f `:citerefrussellnewmanwilliamson1975`aRussell, W. C.; Newman, Carol; Williamson, D. H. (1975). "A simple cytochemical technique for demonstration of DNA in cells infected with mycoplasmas and viruses". `*Nature`*. `!253`! (5491): 461–462. `F33f`_`[Bibcode`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Bibcode_(identifier)]`_`f:1975Natur.253..461R. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.1038/253461a0. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 46112. `F33f`_`[S2CID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=S2CID_(identifier)]`_`f 25224870.

`:cite-note-6`!6.`! `F0af`_`[↑`#cite-ref-6]`_`f `:citerefbiancardibiverseccomennucci2013`aBiancardi, Alessandro; Biver, Tarita; Secco, Fernando; Mennucci, Benedetta (2013). "An investigation of the photophysical properties of minor groove bound and intercalated DAPI through quantum-mechanical and spectroscopic tools". `*Phys. Chem. Chem. Phys`*. `!15`! (13): 4596–603. `F33f`_`[Bibcode`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Bibcode_(identifier)]`_`f:2013PCCP...15.4596B. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.1039/C3CP44058C. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 23423468.

`:cite-note-7`!7.`! `F0af`_`[↑`#cite-ref-7]`_`f `:citerefzinksadonistelzer2003`aZink D, Sadoni N, Stelzer E (2003). "Visualizing Chromatin and Chromosomes in Living Cells. Usually for the live cells staining Hoechst Staining is used. DAPI gives a higher signal in the fixed cells compare to Hoechst Stain but in the live cells Hoechst Stain is used". `*Methods`*. `!29`! (1): 42–50. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.1016/S1046-2023(02)00289-X. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 12543070.

`:cite-note-8`!8.`! `F0af`_`[↑`#cite-ref-8]`_`f DAPI MATERIAL SAFETY DATA SHEET. kpl.com

`:cite-note-9`!9.`! `F0af`_`[↑`#cite-ref-9]`_`f `:citerefohtatokishitayamagata2001`aOhta T, Tokishita S, Yamagata H (2001). "Ethidium bromide and SYBR Green I enhance the genotoxicity of UV-irradiation and chemical mutagens in `*E. coli`*". `*Mutat. Res`*. `!492`! (1–2): 91–7. `F33f`_`[Bibcode`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Bibcode_(identifier)]`_`f:2001MRGTE.492...91O. `F33f`_`[doi`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Doi_(identifier)]`_`f:10.1016/S1383-5718(01)00155-3. `F33f`_`[PMID`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=PMID_(identifier)]`_`f 11377248.

>>See also

Wikimedia Commons has media related to

DAPI

• `F33f`_`[DNA binding ligand`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=DNA_binding_ligand]`_`f

• `F33f`_`[Hoechst stain`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Hoechst_stain]`_`f

• `F33f`_`[Lexitropsin`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Lexitropsin]`_`f

• `F33f`_`[Netropsin`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Netropsin]`_`f

• `F33f`_`[Pentamidine`:/page/wikibook/entry.mu`zim=wikipedia_en_all_nopic_2025-08.zim|entry_path=Pentamidine]`_`f

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