Ethidiummonoazidebromide(EMA)isafluorescentnucleicacidstainwithaphotoaffinitylabel.Thedye,afterphotolysis,bindscovalentlytonucleicacids.ThedyehasbeenusedtofootprintdrugbindingsitesonDNA2,tomodifyplasmidDNA,3,4andforlabelingyeastcellsforleukocytephagocytosisassays5.Aparticularlyusefulapplicationofthedyeistolabeldeadcellsinamixedpopulationofliveanddeadcells.BecauseEMA isrelativelyimpermeanttolivecells,itselectivelybindsDNAindeadcells.PhotolysisrendersthedeadcellDNAcovalentlylabeledwiththedye.Onecanthenwashandfixthecellpreparationforanalysisbymicroscopy,fluorescenceplatereaderorflowcytometry.Themajoradvantageofthismethodisthatresearcherscanavoidextensivemanipulationoflivepathogenicorganisms.7
EMA hasbeenusedtodifferentiatebetweenviableanddeadbacteriaby5′-nucleasePCR.8
ForviabilityPCRapplications,Biotiumalsoofferspropidiummonoazideiodide(PMA,catalognumbers40013and40019)aswellasournewandimprovedviabilityPCRdye,PMAxx™(catalognumber40069).PMAandPMAxx™arelessmembranepermeantthanEMAandthereforemoreselectivefordeadcelllabelingforqPCRquantitationofviablebacteria.9
ForphotoactivationofPMA,PMAxx™orEMAdye,werecommendtheuseofBiotium’sPMA-Lite™LEDPhotolysisDevice(E90002),whichisdesignedtoconductphotolysisundercontrolledconditions.
- Ex(pH3)=458nm
- λEx/λEm(afterphotocrosslinkingtonucleicacid)=510/600nm6
- OrangesolidsolubleinDMF,orethanol
- Storedesiccatedat4oCor-20oCandprotectfromlight,especiallyinsolution
- C21H18BrN5
- MW:420
- [58880-05-0]
References
1.JMolBiol92,319(1975).
2.EuroJBiochem182,437(1989).
3.JBiolChem257,13205(1982).
4.JBiolChem259,11090(1984);.
5.Cytometry11,610(1990).
6.ForexcitationandemissionofthedyebeforeandafterbindingtoDNA/RNAandphotolysis,pleaseseeNucleicAcidsRes.5,4891(1978).
7.Cytometry,12,133(1991).
8.Biotechniques,34,804(2003)
9.J.Microb.Meth.,67,310(2006).
