PRACTICAL # 5

GEL DOCUMENTATION SYSTEM

1. INTRODUCTION:-
A gel doc, also known as a gel documentation system, gel image system or gel imager,
refers to equipment widely used in molecular biology laboratories for the imaging and
documentation of nucleic acid and proteins suspended within polyacrylamide and agarose
gels.

Figure 1: Gel-Documentation System

2. WORKING PRINCIPLE:-
Gel documentation works in the principle of fluorescence. Nuclei acids stained with a
fluorescent substance such as Ethidium Bromide are excited by ultraviolet irradiation and
emit fluorescent light. Depending on the molecular weight and concentration of nucleic acid
the amount of Ethidium Bromide binding is made specifically to nucleic acid.

In other words, the larger the molecular weight of nucleic acid, the higher the binding,
and the fluorescence will shine brighter. Lower the molecular weight of nucleic acid,
fluorescence will be weaker.

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3. COMPOENENTS OF GEL DOCUMENTATION SYSTEM:-
Generally a gel documentation system includes:-

a) Source of Irradiation.
b) Base Plate.
c) Hood.
d) Filters
e) Visible Light.
f) Adjustable Stage.
g) Imaging System.
h) Readout System.

Figure 2: Components of Gel Documentation System

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3.1. Source of Irradiation:-

Transilluminator is the source of irradiation and is used in the observation of DNA

bands. Usually, UV light is used for radiating samples. UV light is used to excite fluorescent
molecules, such as those present in DNA or protein stains, making them visible for imaging.
The excitation range can vary from 250-800 nm.

For UV visualization, 254 nanometer light is optimal for DNA cross-linking, 302
nanometer light is ideal for short exposure gels stained with Ethidium Bromide, and 365
nanometer light is best for gel band cutting.

Nowadays, white light and LED lighting options are also available. The white light
LED option provides viable intensity with a safety cut off that prevents damaging the sample
and operators.

3.2. Base Plate:-

The base plate is a sample tray and can be easily pulled out. It is used to hold the gel
for viewing. It is placed on a non-reflective and black surface.

3.3. Hood:-

It is a darkroom for shielding the skin against UV rays. It is suitable for fluorescence,
chemiluminescence, and visible light applications. There is an electronic auto door lock,
which is used for the prevention of longer exposures to radiation. It ensures proper
documentation of captured images by acting as a dark box. Fold down door and slide-to-
door options are available for less interference.

3.4. Filters:-

Amber filters should be able to shield the UV radiation fully from the eyes. It is also
used in blocking background light, otherwise, the noise will appear on captured images.
Emission filters are used to block the UV radiation for safe viewing of the illumination of
the sample. An extensive variety of filters are available depending on the type of application.

3.5. Visible light:-

Visible light is used for the extension of transmitted light application. For visible light

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 application, visible light converts the screen.

3.6. Adjustable Stage:-

It is used to move the sample closer to the camera.

3.7. Imaging System:-

A camera (CCD with high resolution) unit integrated with darkroom, camera controller,
video monitor, and printer. The resolution of the camera normally ranges from 1.4 MP to
8.3 MP and it is present above the hood for capturing images.

3.8. Readout System:-

A touch screen monitor and a normal monitor comes with inbuilt software that enables
image optimization, analysis, and acquisition. Image editing is also possible.

4. TYPES OF GEL DOCUMENTATION SYSTEM:-

Different types of gel doc systems or gel imaging systems are used depending on the
laboratory applications and detection methods.

 Chemiluminescence is basically used for western blotting.

 Film Auto Radiography is used for radioisotopes
 Laser is used for multiplex fluorescence applications, high sensitivity, and precise
quantitation for radioisotopes
 Multiplex is used to detect and image multiple fluorescent signals simultaneously. This
type of gel imaging system is also used to detect luminescent, colorimetric, and radio
isotopic signals.
 Phosphoimagers is the advance of multiplex fluorescence applications and
radioisotopes. It is more sensitive than multiplex gel imagers. It is more flexible in
choosing emission filters.
 CCD/ Digital Ethidium Bromide (UV), infrared, color fluorescence,
chemiluminescence, and visible light detectors are available to take quantitative reads
of nucleic acid, protein fragments, microplates, and dot blots.

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5. PROCEDURE:-

Ensure you are wearing a lab coat, closed toed shoes, gloves and eye protection.

a) Launch the software:-

 Ensuring you are using an ungloved hand, click the mouse to activate the monitor.
 Open the GelDoc software if it is not already open.
 On the menu bar, select ‘File’, toggle down to ‘Acquire’ and select ‘GelDoc’.

b) Positioning the gel:-

 Open the chamber door with the ungloved hand and with the gloved hand load the
gel into the chamber. Centre the gel, using the monitor to assist in visualization.
 Close door and switch on the UV light.
 With an ungloved hand, adjust the focus, zoom and aperture on the camera to obtain
the optimal image.
 In the GelDoc window, click ‘Capture’. Select the hatched-box icon in this window
and drag it to select the area of interest.
 On the menu bar, select ‘Edit’ and cursor down to ‘Extract’. A new window will
appear with the final picture. You may wish to adjust the image properties such as
brightness and contrast.

c) Printing:-
 On the menu bar, select ‘File’, cursor down to ‘Video Print’ and click to print.

d) Closing the program:-

 Turn off the UV light.
 With a gloved hand, remove your gel from the chamber and wipe down the glass
surface of the GelDoc with a Kimwipe.
 Close the door to the chamber with the ungloved hand.
 Record your name and the number of photos taken in the log book.

e) Decontamination:-
To decontaminate the area, it is recommended that a solution of sodium nitrite and
hypophosphorus acid be used. Add 4.2g sodium nitrite and 20 mL of hypophosphorus
acid to 300mL of water. Alternatively, a 10% bleach solution can be used.

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  Soak a paper towel with the decontamination solution and wash the affected area.
 Use a UV-light to locate any remaining EtBr.
 Use water and paper towel to clean area again. Rinse the area a few times to be sure
any residue has been cleaned up.

6. RESULTS:-
The gel documentation results reveal distinct DNA bands, providing a visual
representation of separated fragments and enabling molecular weight determination and
qualitative assessment of nucleic acid quality. Band intensity and pattern analysis contribute
to quantitative insights into DNA samples.

Figure 3: DNA bands observed under Gel-Doc System.

7. PRECAUTIONS:-
When working with gel documentation systems, it's important to follow certain
precautions to ensure safety, accuracy, and optimal performance. Here are some key
precautions:
 Ethidium bromide is a known mutagen. Always wear a lab coat, goggles and gloves
when handling ethidium bromide solutions and stained agarose gels.
 UV light is harmful to the skin and eyes. Do not look into the UV light source without
face or eye protection.
 Handle gels with care to prevent damage or tearing. Use appropriate tools, such as gel
cutters or spatulas, when necessary. Avoid unnecessary exposure of gels to UV light.
 Dispose of gels and any materials containing hazardous substances, such as ethidium
bromide, in accordance with laboratory safety protocols and regulations.

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8. APPLICATIONS:-
Gel documentation systems play a crucial role in molecular biology and biochemistry
research, allowing scientists to visualize, analyze, and document the results of gel-based
experiments. Here are some key applications of gel documentation systems:

 Monoclonal and polyclonal antibody binding affinities.

 Gel and blot imaging.
 Colony counting.
 Immunoassay.
 Multiplex Protein Detection.
 Post-Translational Modification Characterization.
 2D Electrophoresis.
 Protein Quantitation.

9. SURVEY:-
There is only one gel documentation system in our department.

 Ph. D GENERAL LAB:-

Equipment name: Dolphin-Doc Plus
Company: WEAL TEC
Features: Dolphin-Doc Plus Gel Image system, complete included 16bit 5.0M pixels
camera, lens, MD-25K 302nm UV- transilluminator, Epi white light, high transparent amber
filter, Dolphin-1D software, PC excluded, 100-240V/50-60Hz.
Observations:-
a) The base plate was not properly decontaminated. There were ethidium bromide stains.
b) The log book of the equipment was not present.

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Figure 3: Gel doc system in Ph. D general laboratory.

Figure 4: Inside view of gel doc system.

Figure 5: UV Transilluminator.

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 REFERENCES

Asad, N., Cregg, S., Shakya, S., Stegman, S., & Timmons, L. (2023). Sustainable and Cost-
Effective Gel Documentation. Methods and Protocols, 6(2), 21.
Scott, T. M., Dace, G. L., & Altschuler, M. (1996). Low-cost agarose gel documentation
system. BioTechniques, 21(1), 68-72.