2D Gel Scanning Guide

2D gel electrophoresis is an extremely valuable but also demanding technique for performing global proteomic analysis. Making your own gels takes time and needs hard work. After weeks of adapting the sample preparation setup, optimizing the gel running protocol, and finding the corresponding gel staining technique, the final step on the way to computerized analysis is the generation of images from your 2D gels by using the adequate imaging technology.

This last step in the process is at least as important as all the other previous steps for the successful analysis of your experiment. This guide will help you preserve the information that is contained in the gel and produce the best possible input for computerized 2D gel image analysis.

2D Gel Scanning Check List

Here is our check list for scanning 2D gels. We hope that it helps you to achieve the best possible input for computerized image analysis. Please send your questions and comments to support (at) decodon . com

• Use grayscale instead of color images.
• Try to use the complete available grayscale range. Check this using the image histogram.
• Choose the image resolution such that the smallest spots you want to analyze have a diameter of at least 5 pixels. Use our image resolution table as a guide.
• Scan all gel images using the same orientation.
• Place each gel at the same position on the scanner plate. Avoid scanning too much of the area around the gel.
• Use presets in the scanner's software to make the scanning process easy and reproducible. Save resolution and scanning area as presets and reuse them for all gels in your experiment.
• Avoid Photoshop and similar general image processing software if you do not know exactly what you are doing. Use the software that came with your scanner / imager for image post-processing.
• Limit post-processing of images to crop, mirror, and rotation by 90, 180, 270 degrees.
• Avoid producing TIFF files if you can process calibrated image file formats such as *.IMG/INF and *.GEL. Usually, the TIFF files are produced without grayscale calibration. This means you loose precision or grayscales are distorted nonlinearly, making quantitation questionable.
• Avoid using JPEG files for quantitative analysis.

Background Information and More Tips

In the following sections you will find an overview about imaging technology and some explanations helping you to optimize your scanning results.

• Image Resolution
• Image File Formats
• Using a Consistent Image Orientation
• Scanner Settings
• Avoiding Artifacts and Background

What Do You Think?

Please help us to improve this guide, send your comments to support (at) decodon dot com.