imported>JWBE: added Category:Biaryls using HotCat

2025-08-07T15:49:42Z

← Previous revision		Revision as of 15:49, 7 August 2025
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	==Live cells and toxicity==		==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.<ref>{{cite journal \|vauthors=Zink D, Sadoni N, Stelzer E \|title=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. \|doi=10.1016/S1046-2023(02)00289-X \|journal=Methods \|volume=29 \|issue=1 \|pages=42–50 \|year=2003 \|pmid=12543070}}</ref> It is labeled non-toxic in its MSDS<ref>[https://web.archive.org/web/20120305183437/http://www.kpl.com/docs/msds/710301.pdf DAPI MATERIAL SAFETY DATA SHEET]. kpl.com</ref> and though it was not shown to have mutagenicity to ''E. coli'',<ref>{{cite journal \|vauthors=Ohta T, Tokishita S, Yamagata H \|title=Ethidium bromide and SYBR Green I enhance the genotoxicity of UV-irradiation and chemical mutagens in ''E. coli''. \|journal=Mutat. Res. \|volume=492 \|issue=1–2 \|pages=91–7 \|year=2001 \|pmid=11377248 \|doi=10.1016/S1383-5718(01)00155-3}}</ref> it is labelled as a known mutagen in manufacturer information.<ref name="DAPI Nucleic Acid Stain"/> As it is a small DNA binding compound, it is likely to have some [[carcinogenic]] effects and care should be taken in its handling and disposal.		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.<ref>{{cite journal \|vauthors=Zink D, Sadoni N, Stelzer E \|title=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. \|doi=10.1016/S1046-2023(02)00289-X \|journal=Methods \|volume=29 \|issue=1 \|pages=42–50 \|year=2003 \|pmid=12543070}}</ref> It is labeled non-toxic in its MSDS<ref>[https://web.archive.org/web/20120305183437/http://www.kpl.com/docs/msds/710301.pdf DAPI MATERIAL SAFETY DATA SHEET]. kpl.com</ref> and though it was not shown to have mutagenicity to ''E. coli'',<ref>{{cite journal \|vauthors=Ohta T, Tokishita S, Yamagata H \|title=Ethidium bromide and SYBR Green I enhance the genotoxicity of UV-irradiation and chemical mutagens in ''E. coli''. \|journal=Mutat. Res. \|volume=492 \|issue=1–2 \|pages=91–7 \|year=2001 \|pmid=11377248 \|doi=10.1016/S1383-5718(01)00155-3 \|bibcode=2001MRGTE.492...91O }}</ref> it is labelled as a known mutagen in manufacturer information.<ref name="DAPI Nucleic Acid Stain"/> As it is a small DNA binding compound, it is likely to have some [[carcinogenic]] effects and care should be taken in its handling and disposal.

	==Alternatives==		==Alternatives==
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	[[Category:Indoles]]		[[Category:Indoles]]
	[[Category:Amidines]]		[[Category:Amidines]]
			[[Category:Biaryls]]

208.40.86.170: /* Fluorescence properties */

2025-03-07T21:06:29Z

Fluorescence properties

New page

{{short description|Fluorescent stain}}
{{chembox
| Watchedfields = changed
| verifiedrevid = 443556310
| ImageFile=DAPI.svg
| ImageSize=200px
| ImageFile1=DAPI Space Fill.png
| ImageSize1=200px
| IUPACName=2-(4-Amidinophenyl)-1''H''-indole-6-carboxamidine
| OtherNames=4′,6-Diamidino-2-phenylindole
|Section1={{Chembox Identifiers
| InChI = 1/C16H15N5/c17-15(18)10-3-1-9(2-4-10)13-7-11-5-6-12(16(19)20)8-14(11)21-13/h1-8,21H,(H3,17,18)(H3,19,20)
| InChIKey = FWBHETKCLVMNFS-UHFFFAOYAH
| SMILES1 = [H]/N=C(/c1ccc(cc1)c2cc3ccc(cc3[nH]2)/C(=N/[H])/N)\N
| ChEMBL_Ref = {{ebicite|correct|EBI}}
| ChEMBL = 48217
| StdInChI_Ref = {{stdinchicite|correct|chemspider}}
| StdInChI = 1S/C16H15N5/c17-15(18)10-3-1-9(2-4-10)13-7-11-5-6-12(16(19)20)8-14(11)21-13/h1-8,21H,(H3,17,18)(H3,19,20)
| StdInChIKey_Ref = {{stdinchicite|correct|chemspider}}
| StdInChIKey = FWBHETKCLVMNFS-UHFFFAOYSA-N
| CASNo_Ref = {{cascite|correct|CAS}}
| CASNo=28718-90-3

| UNII_Ref = {{fdacite|correct|FDA}}

| UNII = 76BFW26YJO
| ChemSpiderID_Ref = {{chemspidercite|correct|chemspider}}
| ChemSpiderID = 2848
| PubChem=2954
| ChEBI_Ref = {{ebicite|correct|EBI}}
| ChEBI = 51231
| SMILES = [N@H]=C(N)c3ccc(c2cc1ccc(cc1[nH]2)C(=[N@H])N)cc3
}}
|Section2={{Chembox Properties
| C=16 | H=15 | N=5
| Appearance=
| Density=
| MeltingPt=
| BoilingPt=
| Solubility=
}}
|Section3={{Chembox Hazards
| MainHazards=
| FlashPt=
| AutoignitionPt =
}}
}}

'''DAPI''' (pronounced 'DAPPY', /ˈdæpiː/), or '''4′,6-diamidino-2-phenylindole''', is a [[fluorescence|fluorescent]] [[staining (biology)|stain]] that binds strongly to [[adenine]]–[[thymine]]-rich regions in [[DNA]]. It is used extensively in [[fluorescence microscopy]]. As DAPI can pass through an intact [[cell membrane]], it can be used to stain both live and [[Fixation (histology)|fixed]] cells, though it passes through the membrane less efficiently in live cells and therefore provides a marker for membrane viability.

==History==
DAPI was first synthesised in 1971 in the laboratory of Otto Dann as part of a search for drugs to treat [[trypanosomiasis]]. 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 [[mitochondria]]l DNA in [[ultracentrifugation]] in 1975, the first recorded use of DAPI as a fluorescent DNA stain.<ref name="Kapuscinski1995">{{cite journal |last=Kapuscinski |first=J. |title=DAPI: a DNA-specific fluorescent probe |journal=Biotech. Histochem. |volume=70 |issue=5 |pages=220–233 |date=September 1995 |pmid=8580206 |doi= 10.3109/10520299509108199}}</ref>

Strong fluorescence when bound to DNA led to the rapid adoption of DAPI for fluorescent staining of DNA for [[fluorescence microscopy]]. Its use for detecting DNA in [[plant]], [[metazoa]] and [[bacteria]] cells and [[virus]] 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 [[flow cytometry]] was also demonstrated around this time.<ref name="Kapuscinski1995" />

When bound to double-stranded DNA, DAPI has an absorption maximum at a wavelength of 358 nm ([[ultraviolet]]) 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.<ref name="DAPI Nucleic Acid Stain">Invitrogen, [http://probes.invitrogen.com/media/pis/mp01306.pdf DAPI Nucleic Acid Stain] {{webarchive|url=https://web.archive.org/web/20090306113851/http://probes.invitrogen.com/media/pis/mp01306.pdf |date=2009-03-06 }}. accessed 2009-12-08.</ref> DAPI will also bind to [[RNA]], though it is not as strongly fluorescent. Its emission shifts to around 500 nm when bound to RNA.<ref>Scott Prahl, [http://omlc.ogi.edu/spectra/PhotochemCAD/html/dapi(H2O).html DAPI]. accessed 2009-12-08.</ref><ref>{{cite journal|last1=Kapuscinski|first1=J|title=Interactions of nucleic acids with fluorescent dyes: spectral properties of condensed complexes.|journal=Journal of Histochemistry & Cytochemistry|volume=38|issue=9|year=2017|pages=1323–1329|pmid=1696951|doi=10.1177/38.9.1696951|doi-access=free}}</ref>

[[File:1D30 DNA DAPI.png|250px|left|thumb|DAPI (magenta) bound to the minor groove of DNA (green and blue). From {{PDB|1D30}}.]]
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 [[fluorescein]] and [[green fluorescent protein]] (GFP) but the effect of this is small.

Outside of analytical fluorescence light microscopy DAPI is also popular for labeling of [[cell cultures]] to detect the DNA of contaminating ''[[Mycoplasma]]'' or [[virus]]. The labelled ''Mycoplasma'' or virus particles in the [[growth medium]] fluoresce once stained by DAPI making them easy to detect.<ref>{{cite journal|last1=Russell|first1=W. C.|last2=Newman|first2=Carol|last3=Williamson|first3=D. H.|title=A simple cytochemical technique for demonstration of DNA in cells infected with mycoplasmas and viruses|journal=Nature|date=1975|volume=253|issue=5491|pages=461–462|doi=10.1038/253461a0|pmid=46112|bibcode=1975Natur.253..461R|s2cid=25224870}}</ref>

==Modelling of absorption and fluorescence properties==
This DNA fluorescent probe has been effectively modeled<ref>{{cite journal|last1=Biancardi|first1=Alessandro|last2=Biver|first2=Tarita|last3=Secco|first3=Fernando|last4=Mennucci|first4=Benedetta|title=An investigation of the photophysical properties of minor groove bound and intercalated DAPI through quantum-mechanical and spectroscopic tools. |doi=10.1039/C3CP44058C |pmid=23423468 |journal=Phys. Chem. Chem. Phys. |year=2013 |volume=15 |issue=13 |pages=4596–603 |bibcode=2013PCCP...15.4596B }}</ref> using the [[time-dependent density functional theory]], coupled with the IEF version of the [[polarizable continuum model]]. This quantum-mechanical modeling has rationalized the absorption and fluorescence behavior given by minor groove binding and [[intercalation (biochemistry)|intercalation]] 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.<ref>{{cite journal |vauthors=Zink D, Sadoni N, Stelzer E |title=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. |doi=10.1016/S1046-2023(02)00289-X |journal=Methods |volume=29 |issue=1 |pages=42–50 |year=2003 |pmid=12543070}}</ref> It is labeled non-toxic in its MSDS<ref>[https://web.archive.org/web/20120305183437/http://www.kpl.com/docs/msds/710301.pdf DAPI MATERIAL SAFETY DATA SHEET]. kpl.com</ref> and though it was not shown to have mutagenicity to ''E. coli'',<ref>{{cite journal |vauthors=Ohta T, Tokishita S, Yamagata H |title=Ethidium bromide and SYBR Green I enhance the genotoxicity of UV-irradiation and chemical mutagens in ''E. coli''. |journal=Mutat. Res. |volume=492 |issue=1–2 |pages=91–7 |year=2001 |pmid=11377248 |doi=10.1016/S1383-5718(01)00155-3}}</ref> it is labelled as a known mutagen in manufacturer information.<ref name="DAPI Nucleic Acid Stain"/> As it is a small DNA binding compound, it is likely to have some [[carcinogenic]] effects and care should be taken in its handling and disposal.

==Alternatives==
[[File:FluorescentCells.jpg|250px|right|thumb|Endothelial cells stained with DAPI (blue), [[phalloidin]] (red) and through [[immunofluorescence]] via an [[antibody]] bound to [[fluorescein isothiocyanate]] (FITC) (green).]]

The [[Hoechst stain]]s 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==
{{Reflist|2}}

== See also ==
{{Commons category}}
* [[DNA binding ligand]]
* [[Hoechst stain]]
* [[Lexitropsin]]
* [[Netropsin]]
* [[Pentamidine]]

{{DEFAULTSORT:Dapi}}
[[Category:Staining dyes]]
[[Category:Fluorescent dyes]]
[[Category:DNA-binding substances]]
[[Category:Indoles]]
[[Category:Amidines]]

DAPI - Revision history

imported>JWBE: added Category:Biaryls using HotCat

208.40.86.170: /* Fluorescence properties */