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Download the Protein Blotting Guide
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:: Posted by American Biotechnologist on 09-08-2011
Western Blotting is probably one of the most ubiquitous techniques in the molecular biology lab and relatively easy to perform. Yet many of us have been frustrated with statistically insignificant results or protein bands that appear either too dark or too light to quantitate.
Well, do not despair! There are many things you can do to help improve the quality of your blots and increase your likelihood of obtaining statistically significant results!
In the video below, you will learn about the many factors affecting western blot analysis (such as detection limit and dynamic range limitations of film and overloaded gels) and what can be done to improve your chances of success.
The presentation was given by Bio-Rad Laboratories Field Application Specialist Dr. Sean Taylor as part of an intimate customer training. Some of the references in the presentation may be specific for that particular customer but the general information contained in this presentation is highly valuable to all molecular biology labs.
:: Posted by American Biotechnologist on 03-24-2011
Most people who have run polyacrylamide gels have at one point or another gotten confused with the gel’s orientation. This is especially true following protein transfer from the gel to a membrane for the purpose of western blotting or other downstream processing. This can be extremely frustrating and may even jeopardize your entire experiment if you are unable to tell the right side of the gel from the left or your control samples from your treatment group.
Several methods have been created to help bench scientists avoid this problem. These include the use of multicolored protein ladders and marking a predefined corner of your membrane once the protein has been transfered (I cut the bottom left corner of the membrane).
Today, Arefeh Seyedarabiclose from the Department of Structural and Molecular Biology, University College London published a new and very basic method for labeling polyacrylamide gels on the Nature Protocol Exchange website. Essentially, Arefeh suggests labeling the bottom inside corner of the long glass plate (facing the gel) with permanent marker). During electrophoresis the label will transfer from the plate to the gel, thereby permanently labeling your gel. Simple and brilliant!