What is ethidium bromide?
Ethidium bromide (EtBr) is a fluorescent dye widely supplied in molecular biology research. Early intake was as a vet trypanocide.14 the is a mutagenic compound that intercalates double-stranded DNA and also RNA.1,10 The fluorescence of EtBr boosts 21-fold top top binding come double-stranded RNA, 25-fold on binding double-stranded DNA (although histones block binding of EtBr to DNA). Ethidium bromide has been used in many fluorimetric assays for main point acids.15,16,17 It has actually been presented to tie to single-stranded DNA (although no as strongly)2 and also triple-stranded DNA.18 since of the binding come DNA, EtBr is a powerful inhibitor that DNA polymerase.4
Synonyms: 3,8-Diamino-5-ethyl-6-phenylphenanthridinium bromide; 2,7-Diamino-10-ethyl-9-phenylphenanthridinium bromide; Homidium Bromide; Dromilac; EtBr
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Ethidium bromide product details
Appearance: Red to purple powderMolecular formula: C21H20N3BrMolecular mass: 394.3Melting point: 255-266 °C v decomposition1,2
Spectral data:UV/Vis Absorbance:Maxima at 210 nm, EmM = 0.20-0.5; 285 nm, EmM = 5.0-10.0; 316 nm, EmM = 50.0; 343 nm, EmM = 40.0 (water)3Published absorb spectrum (methanol) v λmax at 296 nm and525 nm1λmax = 475 nm, EmM = 5.76 (0.66 M glycine buffer)4
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Solvent effects on absorbance:λmax = 480 nm (water); 515 (glycerol); 520 nm (methanol): 520 nm (acetone); 532 nm (ethanol); 535 (DMSO) and also 540 nm (pyridine).5 The absorbance maximum relative to water is red-shifted to longer wavelengths ~ above binding to main point acids.6,7
FluorescenceExcitation/emission wavelengths reported because that the EtBr-nucleic acid complex:Ex in ~ 526 nm, Em at 605 nm (aqueous)8Ex in ~ 360 nm, Em at 590 nm (in PBS)9Ex in ~ 525 nm, Em in ~ 600 nm (10 mM TBE, pH 8.0)10Ex (absorption) in ~ 510 nm, Em at 590 nm.11
Ex (absorption) at 482 nm (blue-green), Em in ~ 616 nm (red-orange).12 The fluorescence productivity of EtBr rises as solvent polarity decreases.13
Storage: The solid flour has presented minimal readjust after two years stored in ~ room temperature, defended from light.
The recommended ultimate disposal an approach is through incineration as disputed in the SDS.23 processing solutions through Rohm and also Haas Amberlite XAD-16 resin was presented to be efficient to the border of detection, through none the the fully decontaminated solution discovered to be mutagenic.24,25 We sell XAD-16 together a bulk product, and also several commodities that adsorb the dye for simpler disposal. Check out Rezorian A161 cartridges (Product No. 57611 or 57610) and the Extractor™ for EtBr decontamination (Product No. Z361569). A 5 mL Rezorian A161 cartridge can be provided to treat an ext than 16 liters of systems (0.5 mg/liter of EtBr) prior to dye breakthrough. The luer lock cartridge have the right to be supplied with 4 mm I.D. Tubing, syringes or low-pressure chromatographic systems. The Extractor™ because that EtBr decontamination can procedure up to 10 liters of equipment (0.5 mg/liter of EtBr). Other methods of treatment for handle of aqueous EtBr solutions have been suggested.26,27,28 We have actually not verified and also do no endorse these procedures.
Methylene blue (Product No. M4159) was reported together an different stain.19 Thiazole orange homodimer (monomer = Product No. 390062) was listed as nonmutagenic, but helpful for detecting DNA in agarose gels.20
For usage in cell studies, dihydroethidium (Product No. D7008), additionally known together hydroethidine, is a possible alternative. It is the reduced form of the dye and is convert (dehydrogenated) within the cabinet to the ethidium cation, which climate intercalates right into DNA.21,22
Preparing ethidium bromide
At room temperature, EtBr disappear in water at 10 mg/mL to provide a red solution.2 It have to be soluble approximately 20 mg/mL in water or as much as 2 mg/mL in ethanol.1 that is dissolve at 1 component in 750 components chloroform.3
Stock solutions of EtBr in water or PBS room stable for at the very least two year at room temperature if protected from light.11
Electrophoresis staining with ethidium bromide
Use in Electrophoresis Staining6The dye is usually incorporated into the gel and also the electrophoresis buffer in ~ 0.5 mg/mL. Note: Electrophoresis mobility of direct double-stranded DNA is decreased by about 15 % in the existence of the dye.To stain after gel has been run, immerse gelatin in electrophoresis buffer or water containing EtBr (0.5 mg/mL) for 30-45 minute at room temperature.Destaining is optional. Detection of tiny amounts (