Sensitivity of FluoroVue™

The FluoroVue™ Nucleic Acid Gel Stain (NS1000) shows a green-yellow fluorescence under blue light excitation. The sensitivity of NS1000 is about 0.14 ng (arrow) for a 4 kb fragment. 

Excitation and emission spectrum of FluoroVue™ 

FluoroVue™ Nucleic Acid Gel Stain (NS1000) has a fluorescent excitation maxima of ~250 and ~482 nm, and an emission maximum of ~509 nm. Therefore, it can replace EtBr without the need for changing existing lab imaging systems. 

Non-mutagenicity of FluoroVue™

FluoroVue™ Nucleic Acid Gel Stain (NS1000) is proofed for their safety (non-mutagenicity) using Ames test. However, it must be noted that since solvent may penetrate the skin, it is recommended that users wear gloves when using the fluorescent dyes.

In- gel staining

This protocol is highly recommended.

1. Prepare TAE or TBE buffered molten agarose solution.

40 ml molten agarose solution can cast two mini-gels (5.4 x 5.9 cm) or one landscape-gel (10.9 x 5.9 cm).

2. Dilute FluoroVue™ Nucleic Acid Gel Stain 10,000X with the molten gel solution and mix well prior to being poured into the gel. 

For example: 4 μl in 40 ml molten agarose solution

3. Cool the molten agarose solution until it can be handled by hand and then pour it into gel tray.

Casted gels are stable at 4°C for 3 days in dark. After three days the sensitivity will decrease daily. 

4. Perform agarose gel electrophoresis (avoid light).

The recommended voltage is 4–10 V/cm (distance between anode and cathode). Avoid using high voltage during electrophoresis. High voltage causes excess heat and affects the dye adversely. 

During electrophoresis, the staining dye runs toward the anode, therefore DNA bands with smaller molecular weights may be weaker in intensity due to less staining dye. 

5. Visualize or photograph the gel with UV or blue-light illumination (blue-light is recommended).

Clean the surface of the illuminator before and after each use with deionized water. Accumulation of fluorescent dyes on the surface will create a high fluorescent background.

Video cameras and CCD cameras have a different spectral response compared to the black-and-white print film and therefore may not exhibit the same sensitivity.

Staining during electrophoresis

The sensitivity of this method is slightly lower than the in-gel staining.

1. Prepare agarose gel following your standard protocol.

2. Dilute FluoroVue™ Nucleic Acid Gel Stain 10,000 folds into the running buffer and mix well.

For example: 30 μl in 300 ml running buffer 

3. Perform agarose gel electrophoresis (avoid light).

During electrophoresis, the staining dye runs toward the anode.

4. Visualize or photograph the gel with UV or blue-light illumination (blue-light is recommended).

Clean the surface of the illuminator before and after each use with deionized water. Accumulation of fluorescent dyes on the surface will create a high fluorescent background.

Video cameras and CCD cameras have a different spectral response compared to the black-and-white print film and therefore may not exhibit the same sensitivity.

Staining after electrophoresis (post-staining)

Post-staining method is recommended for polyacrylamide electrophoresis, due to the longer time required for running PAGE. The sensitivity of this method is lower than the in-gel staining method.

1. Performing agarose gel electrophoresis following your standard protocol.

 2. Dilute FluoroVue™ Nucleic Acid Gel Stain 10,000 folds into the TE, TAE, or TBE buffer and mix well.

For example: 10 μl in 100 ml TAE buffer 

Use a plastic container. Glass containers are not recommended, as they absorb fluorescent dye in staining solution. 

Protect the staining container from light by covering it with aluminium foil, or place it in the dark.

The staining solution can be stored for up to one week or more.

3. Immerse the gel in a staining solution (1X) and incubate at room temperature for 10 - 30 minutes. (avoid light).

Staining time varies with the thickness of the gel and percentage of agarose. If needed, agitate the gel gently at room temperature to shorten staining time. 

4. Visualize or photograph the gel with UV or blue-light illumination (blue-light is recommended).

Clean the surface of the illuminator before and after each use with deionized water. Accumulation of fluorescent dyes on the surface will create a high fluorescent background.

Video cameras and CCD cameras have a different spectral response compared to the black-and-white print film and therefore may not exhibit the same sensitivity.

B-BOX™ Blue Light LED Epi-illuminator

• 470 nm long wavelength

• Improved cloning efficiency 

• Compact, light-weight, and portable (less than 1 kg)

• Adjustable and removable filter plate allows for gel cutting, visualization, and documentation

ExcelBand™ DNA Ladder series

• Sharp bands

• Quick reference— enhanced bands 

• Ready-to-use— premixed with loading dye for direct loading

• Stable— room temperature storage over 6 months

FluoroStain™ DNA Fluorescent Staining Dye

• Excellent for post staining

• Sensitivity up to 0.04 ng DNA

• A safe alternative to EtBr

• Suitable for blue or UV light