LI-COR Biosciences has launched a new piece of free software for the analysis of western blot gels and film called Image Studio Lite. I have provided a tutorial for the use of this new software for simple densitometry analysis.
After some time away, I am back and ready to address some questions regarding western blot quantification. I hope you’ll all excuse the delay, but real life and research have been a bit overwhelming the past month. I will try to reply to all of the backed-up comments as quickly as possible.
Please feel free [...]
As new students start to arrive in your labs for the new school year, don’t forget to direct them to our site. We have free and proven protocols for western blotting. We also include troubleshooting, densitometry tutorials, and
In recent weeks, we’ve seen a large growth of traffic to this site, along with an increase in requests for various protocols and buffers. Because of this, we are expanding our protcol sections to include some basic protein techniques for molecular biology (e.g. immunoprecipitation, gel staining, etc.). Immunoblotting will still be our focus, but we [...]
We have just finished adding a new discussion forum for our site. We sincerely hope that this forum will help the researchers who use our site troubleshoot and share information effectively.
I hesitated to start a forum so quickly, but keep in mind that although this is a relatively [...]
After a long wait, we have finally posted our general western blot overview. This may be useful for beginners who are just learning the basics of western blotting technique, or people outside of the field who want to understand how western blotting works. There are some good figures of how proteins [...]
Western Blot Stripping Buffer:
Making your own western blot stripping buffer is easy and affordable. We essentially always use PVDF membranes for our western blots because of their durability and low background for a variety of primary antibodies. PVDF membranes are useful because they can be stripped and re-probed many times [...]
If you are not currently, reusing primary antibodies for your western blots, you’re washing a lot of money down the drain.
In our lab, we regularly reuse primary antibodies at least 3 times before we get rid of them. Some antibodies will work longer than others. In particular, antibodies for post-translational modifications [...]
I previously shared a review of the Snap i.d. rapid western blotting system from Millipore. I stated that the product definitely works with a good set of antibodies, but individual researchers must decide for themselves whether the saved time was worth the Snap i.d. price tag.
The newest and best polyacrylamide gels for western blotting use bis-tris as a buffer. These gels are sold by Invitrogen under the brand name NuPAGE®. However, these gels can be expensive, and, of course, have a limited shelf life. So it’s really easier if you just make them yourself in your lab.
On [...]
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