The main differences between Safe-Green, Safe-Red and Safe-white is their DNA binding dye and its excitation/emission spectra. Safe-Green has 500/522nm; Safe-White has approximate fluorescence excitation/emission maxima 358/461nm; Safe-Red has 488/562nm. Among these three, Safe-White has higher sensitivity than the other two colours.

If you have a 10ul sample, you should add 2ul of SafeView Dye to your sample before loading onto the gel.

SafeView can be used as a replacement for ethidium bromide as they both work on general agarose. We recommend using SafeGreen for downstream cloning applications as SafeView can interfere with the ligation reaction, yielding fewer colonies.

There are fluorescent compounds in SafeView and these fluorescent compounds have the capability to bind to DNA. There may be some unknown effects of SafeView that have not been documented but that applies to the SYBR set as well; however, SafeView products are definitely not as carcinogenic as ethidium bromide.

The Safe-(Red, Green and White) loading dyes work the same as 6x loading dye, loaded with the sample. SafeView Classic is used directly in the gel and the running buffer.

Under UV, SafeView Classic emits a green fluorescence when bound to both single and double stranded DNA templates, therefore they cannot be differentiated by this method. It will emit a red fluorescence when bound to RNA templates. The SafeView Stains (Red, Green and White) do not perform in this way and will stain all nucleic acid templates one color.

All the SafeView products should be stored at four degrees Celsius.

Under UV light, SafeView Classic emits a green fluroescence when bound to both single and double stranded DNA templates. It will emit a red fluorescence when bound to RNA templates. No filter is necessary for viewing these colours, however a filter may be needed for photographing the gel.

Our in-house testing has shown that SafeView stained gels (>10ng DNA loaded per lane) can still be effectively visualized up to 1 week later with only a slight decrease in brightness. Gels should be stored properly to maintain a good signal, at 4C in the dark, sealed in a plastic bag or pouch with wet paper towel loosely wrapped around the gel.

Make sure you are following the protocol carefully. It is critical that both buffer and gel have SafeView dye in them otherwise it will not work. This is different from ethidium bromide. You can consider to add 2.5ul of SafeView (instead of 5ul) for every 100ml of running buffer, which will reduce background fluorescence and allow the bands to show with more contrast.

2 hours over 100C

Safe-Green has Excitation Wavelength of 490nm and Emission Wavelength of 525nm, and its sensitivity range is between 0.2-0.6ng. Safe-Red has Excitation Wavelength of 540nm and Emission Wavelength of 630nm, and its sensitivity range is between 0.3-0.8ng. Safe-White has Excitation Wavelength of 370nm and Emission Wavelength of 470nm, and its sensitivity range is between 0.2-0.5ng. SafeView Classic emits green fluorescence when bound to dsDNA and ssDNA and red fluorescence when bound to RNA. This stain has one excitation (490 nm) and two emission spectra (520 nm and 635 nm) and the sensitivity range is between 0.1-0.3ng. SafeView Plus has Excitation Wavelength of 490nm and Emission Wavelength of 525nm and its sensitivity range is between 0.05-0.1ng.

Only SafeView Plus (G468) should be used in a post stain. SafeView classic and Safe Stains are not designed for post-staining. SafeView Classic must be added to the gel and the running buffer prior to the loading of the samples. Safe-(Red, Green and White) stains must be added to the sample before loading it to the gel.

A reason for this is that most gel doc systems have been optimized for EtBr so that is why the EtBr signal may be stronger in the pictures.

Yes, it is correct. Safe-Pack (Red, Green and White) has different different protocol from Safeview Claasic. Safe-Pack requires only to be mixed with DNA sample before loading the gel. There is no need to add dye in the gel or buffer.

SafeGreen works best between pH 7 to 9.

We know that gels with Safeview can last a week in the refrigerator. If it is stored longer than a week, the agarose gel will have resolution problem.

We know that gels with Safeview can last a week in the refrigerator. If it is stored longer than a week, the agarose gel will have resolution problem.

Dispose SafeView™ Classic as you would any other non-carcinogenic fluorescent dye (eg. Acridine orange; Propidium iodide). All gels and contaminated “non-sharp” lab debris (e.g., gloves, pads, towels, tubes, etc.) that are processed using this dye can be discarded in the trash. Spent running buffer solutions and destaining solutions that contain the dyes can either be collected and disposed of through the HWMU or collected and run through an approved filter device. The buffer solutions that have been run through the approved filter should be checked under the appropriate light source for complete removal of the dyes, and if it passes (does not fluoresce), the liquid can be disposed of down the drain with a copious amount of water as long as no other materials are present that would cause the material to be a Hazardous Waste. The filters that have been used up and are no longer effective must be disposed of through the HWMU.

The loading dye is included in the products. No additional loading buffer needed.

We recommend using Safe-Green (G108-G) for downstream cloning applications, as testing has shown it yields more colonies following the ligation reaction than SafeView (G108).

The difference is simply in colour of the dye, when viewed under UV the bands in the gel will show up in green, red or white.

Shifting is unavoidable and quite natural for any fragments, regardless of the staining agent. We suggest to use SafeGreen ladders, which will give accurate molecular weight with no additional staining agents needed. http://www.abmgood.com/DNA-Ladder-Safe-Green™-100bp-Opti-DNA-Marker-Invitroge-G473.html http://www.abmgood.com/DNA-Ladder-Safe-Green™-1kb-Opti-DNA-Marker-Invitroge-G474.html

Shifting is unavoidable and quite natural for such fragments, regardless of the staining. We suggest using SafeGreen ladders, which will give accurate molecular weight with no additional staining agents needed. http://www.abmgood.com/DNA-Ladder-Safe-Green™-100bp-Opti-DNA-Marker-Invitroge-G473.html http://www.abmgood.com/DNA-Ladder-Safe-Green™-1kb-Opti-DNA-Marker-Invitroge-G474.html

It is very difficult to detect 100bp and 200bp bands in 1% gel with any stains. Higher gel concentrations should be used, such as 2% agarose.

Yes, we have tested our SafeView products for this application.

We use both UV and LED transilluminators, our gel QC pictures in PCR department are taken with UV.

UV and LED will give the same colors.

All of our SafeView stains have been tested in-house to be compatible with UV light. SafeView Classic, SafeView Plus, and SafeGreen will also work under blue light/LED. SafeRed and SafeWhite will only work under UV light. Safe View Plus should only appear green. SafeView Classic will be red for RNA and green for DNA.

It should be fine to store at -20°C, it will only prolong the shelf life (normally 2 years).

No, please store at 4°C only. Shelf life of 2 years.

20000X (5ul of the G108 in 100ml gel solution).

Yes

Yes, the dye should migrate on a 1% gel.

No.

6X loading dye

No, adding SafeGreen to the DNA ladder will not interfere with the DNA ladder's loading dye. You still need to add SafeGreen to the DNA ladder at the same ratio. To be more technically correct, SafeGreen, and all other EB alternative DNA binding dyes, would alter the migration pattern of DNA on agarose gel by shifting DNA to a larger molecular size; it is just simply because the overall molecular size of a particular DNA band increases as more dye is binding onto it. This is why we recommend mixing the DNA ladder and SafeGreen at the exact same 1:6 ratio, so everything on the gel will be subjected to the same concentration of SafeGreen and thus having the same shift.

Most labs are equipped with UV or LED light sources for viewing agarose gels, which is why we suggest these. As for SafeRed, yes, it does have an additional excitation wavelength of around 300nm. The other wavelength reported is for other people who may decide to use other light sources for the visualization.

Yes, SafeView Classic can penetrate cell membranes.

The dye should be at ~300bp after electrophoresis.

Our Safe-Red™ (G108-R) will not affect the downstream cloning experiments, and thus no extra treatment is required to recover the stained sample DNA.

Yes, our Safe-Green™ stain indeed has the ability to penetrate the nucleus.

Of our SafeView™ series DNA Stains, only SafeView™ (G108) and Safe-White™ (G108-W) can penetrate cell membranes, while the rest cannot.

The complex interaction of SafeView Classic with DNA and RNA means that the emission spectrum is affected by secondary structure, temperature, ionic strength, pH and SafeView Classic concentration. Also, at high polynucleotides to SafeView Classic ratios, most nucleic acids will appear green. If the gels contain 6M urea at pH 3.5, all nucleic acids will appear green. So it is not unusual to observe the RNA bands as green under certain conditions.

The color of the dyes seen during migration would be blue, the same for all 3 dyes (Safe-Green/ Safe-Red/Safe-White). The size of the dye should be at approximately 300bp after electrophoresis. All three loading dyes have different excitation and emission wavelengths, so they will emit different colors. When bound to DNA, Safe-Green emits green fluorescence, Safe-Red emits red fluorescence, and Safe-White emits whitish pale blue fluorescence.

Any imaging system with UV and/or LED should work. No filter for Safeview is required, however a filter at ~500-650nm is optional.