Technology Highlights > Gelred and Gelgreen

GelRed™ and GelGreen™

Environmentally safe and ultra-sensitive nucleic acid gel stains for replacing EtBr

  GelRed™ and GelGreen™ dye flyer                     FAQs for GelRed™ and GelGreen™ Dyes
  GelRed GelGreen Safety Report

GelRed™ and GelGreen™ are a new generation of fluorescent nucleic acid gel stains designed to replace the highly toxic ethidium bromide (EtBr). Developed by scientists at Biotium, GelRed™ and GelGreen™ are superior to EtBr and other EtBr alternatives by having a combination of low toxicity, high sensitivity and exceptional stability.

GelRed™ Is Superior to EtBr

       GelRed                      EtBr

Figure 1. Comparison of GelRed™ and ethidium bromide (EtBr) in precast gel staining using 1% agaose gel in TBE buffer. Two-fold serial dilutions of 1 kb Plus DNA Ladder from Invitrogen were loaded onto each gel in 4 lanes in the amounts of 200 ng, 100 ng, 50 ng and 25 ng, respectively, from left to right. Gels were imaged using a 300-nm transilluminator and photographed with an EtBr filter and Polaroid 667 black-and-white print films.

EtBr has been the predominant dye used for nucleic acid gel staining for decades because of its low price and generally sufficient sensitivity. However, EtBr is a highly mutagenic material. The safety hazard and costs associated with decontamination and waste disposal can ultimately make the dye expensive to use. For this reason, alternative gel stains, such as SYBR dyes, have become commercially available in recent years. Although these alternative dyes have reduced mutagenicity, they often have to sacrifice other aspects of the dyes. For example, SYBR Safe has very limited sensitivity while SYBR Green and SYBR Gold are much less stable than EtBr. SYBR dyes also enter cells rapidly to stain mitochondria and nuclear DNA, making it more likely for the dyes to be toxic at high enough concentrations. Indeed, SYBR Green I is known to strongly potentiate mutation caused by UV light or another mutagen (Ohta, et al. Mutat. Res. 492, 91(2001)).

GelGreen Is Simply Unmatched by SYBR Safe

       GelGreen               SYBR Safe

Figure 2. Comparison of GelGreen and SYBR Safe in post gel staining using 1% agarose gel in TBE buffer. Two-fold serial dilutions of 1 kb Plus DNA Ladder from Invitrogen were loaded onto each gel in 4 lanes in the amounts of 200 ng, 100 ng, 50 ng and 25 ng, respectively, from left to right. Gels were imaged using a 254-nm transilluminator and photographed with a SYBR filter and Polaroid 667 black-and-white print films.

To make GelRed™ and GelGreen™ safe, scientists at Biotium used a novel yet very simple concept: reducing genotoxicity by preventing the dyes from entering living cells. We believe that a DNA-binding dye can be made nonmutagenic or substantially so by denying its chance to be in contact with genomic DNA in living cells. Thus, we engineered the chemical structures of GelRed™ and GelGreen™ such that the dyes are incapable of crossing cell membranes. The Ames test confirmed that GelRed™ and GelGreen™ are nonmutagenic at concentrations well above their working concentrations used for gel staining. Furthermore, environmental safety tests showed that GelRed™ and GelGreen™ are nonhazardous and nontoxic to aquatic life. As a result, GelRed™ and GelGreen™ can be disposed of down the drain or in regular trash. For more information, please download the GelRed™/GelGreen™ Safety Report on Biotium website.

GelRed™ and GelGreen™ are highly sensitive either as precast gel stains or post gel stains. Designed primarily for use with a 312/302 nm UV transilluminator, GelRed™ is much more sensitive than EtBr, and at least as sensitive as or brighter than SYBR Gold in post gel staining. Unlike SYBR Gold, GelRed™ can also be used as a highly sensitive precast gel stain. GelGreen is developed to meet the needs of researchers who use a 488 nm laser-based gel scanner or a Dark Reader that uses a visible blue light for excitation. GelGreen™ is spectrally similar to SYBR Safe, but is far more sensitive than the latter.

Another major advantage of GelRed™ and GelGreen™ is their remarkable stability. You can handle the two dyes the same way you do with EtBr. This means that the dyes are perfectly stable in water at room temperature for long-term storage, and they can be microwaved for making precast gels. Both dyes are also very photostable, permitting their use under normal room light without exercising special precaution.

Figure 3. Normalized excitation and emission spectra of GelGreen (green) and GelRed™ (red) in the presence of dsDNA in PBS buffer.
	SYBR Dyes	GelRed™	GelGreen
A) 5 min. incubation				The best way to make a DNA-binding dye safe is to prevent it from going into cells in the first place.
B) 30 min incubation
Figure 4. HeLa cells were incubated at 37 °C with 1X of SYBR Green I, SYBR Safe, GelGreen™ and GelRed™, respectively. Images were taken following incubation for 5 min (panel A) and 30 min (panel B), respectively. SYBR Green I and SYBR Safe entered into cells rapidly as evident from the bright green nuclear staining (images from SYBR Safe not shown). However, GelRed™ and GelGreen™ were unable to cross cell membranes as shown by the lack of any fluorescence staining.
Figure 5. Comparison of mutagenicity among GelGreen™, GelRed™, SYBR Green I and EB in +1 frameshift Salmonella indicator strain TA98 with the presence of S9 fraction. ""indicates EB was not tested at this concentration. "" indicates SYBR Green I became cytotoxic at this concentration. For more information, you can download GelRed™ and GelGreen™ Safety Report on Biotium website.

* GelGreen™ and GelRed™ are covered by pending US and international patents.
** SYBR is a trademark of Molecular Probes, Inc.

Table A. Ordering Information