this was great for someone like me who hasn't done it yet
@Polecat-qz5om2 жыл бұрын
Key word yet 😃
@JYOtiRaNJanMANgaRaj6 ай бұрын
THANK U FOR A GREAT 😃 ANALYSIS ABOUT WESTERN BLOTTING. 👍👌
@hkicgh72773 жыл бұрын
3:52 After the centrifugation, do you load the samples in the gel without discarding the supernatant layer?
@cellsignaldotcom3 жыл бұрын
Correct - this centrifugation step is just to pellet any insoluble material, and pull down any liquid from the side walls of the tube. Load 20 microliters without discarding any supernatant.
@maretawidiya96729 жыл бұрын
like this protocol,.thanks so much
@thamizh28506 жыл бұрын
On website in the protocol you have mentioned that wash membrane three times for 5 min each after blocking. But here you say only 5 min. So please let me know which one is correct?
@cellsignaldotcom6 жыл бұрын
Thanks for the note. After blocking, 3 washes are optional. We typically rinse the excess milk off with TBS-T; one brief wash is sufficient. After antibody incubation, however, three 5-min washes are recommended.
@hanconway81625 жыл бұрын
Can you recommend a method for storing pancreatic islets after collection from pancreas for western blot? Can the islets be pelleted and stored at -80? What solution should they be left in until western blot is performed? Thanks
@cellsignaldotcom5 жыл бұрын
Thanks for the question, Han. In general, it depends on both the sample type and the stability of the protein of interest. Ideally, lysates should be freshly prepared, but if this is not possible storage at -20C or -80C may be suitable. If not stored in SDS, the buffer should contain protease and phosphatase inhibitors. Some proteins will yield western blot results with little to no change after repeated freeze-thaw cycles, but other will be affected. You can get in touch with a CST scientist, they may have further information for your sample/protein of interest, please use cellsignal.com/support. You may also want to view our Tech Tips video on lysate buffers - kzbin.info/www/bejne/gYeYYqqDqbuBl80
@avanaavan2575 Жыл бұрын
Do you have offers for students to work western blot in your lab??
@cellsignaldotcom Жыл бұрын
Hi Avana. CST has a summer internship program for undergraduate students and rising high school seniors. Please visit www.cellsignal.com/about-us/social-responsibility/internship-program for information.
@giahungtran68222 жыл бұрын
In the protocol on website, after transfer, wash membrane with TBS 5min. However, in this video, it said that we will wash with TBST 5min. So what is the best choice, TBS or TBST?
@cellsignaldotcom2 жыл бұрын
Thanks for the question. Here, either TBS or TBST will work just fine for chemiluminescent WB. Please note that for fluorescent WB, the Tween 20 should be omitted from the optional wash step and the blocking buffer.
@arpitkhanna30394 жыл бұрын
Thats helpful
@p91grwm143 жыл бұрын
I have a question regarding the buffer for the western blott. We use normally 20 % methanol in the solution and we have always used nitril gloves in contact with the solution. But currently I have discovered that these gloves are not suitable for methanol. We had never problems but now I'm afraid from that. Is possible that a 20 % methanol solution in water can penetrate the gloves?
@cellsignaldotcom3 жыл бұрын
Sorry for the delayed response. It is recommended that you contact the glove supplier and your local EHS or lab safety department for guidance.
7 жыл бұрын
thank you for information
@oliviaxu44926 жыл бұрын
is it bad to health while developing x-ray film
@cellsignaldotcom6 жыл бұрын
Hi Olivia, x-rays are not part of the Western Blot protocol. If you are developing film manually in a dark room, please use appropriate PPE (safety glasses, gloves, lab coat).