Hi everybody! I am doing western blots on bovine mammary epithelial cells to determine casein. We have two types of blocking buffer in the lab. PBS: https://www.selectscience.net/products/odyssey-blocking-buffer-(pbs)/?prodID=209631#tab-2
and TBS: https://www.selectscience.net/products/odyssey-blocking-buffer-(tbs)/?prodID=93459#tab-2
The protocol says just to use blocking buffer. Which type is best for me to use in this instance?
Thank you!
For western blot TBS is usually used to make a blocking buffer. it should be TBS-T. You can avoid using PBS if you are working with phosphorylated protein detection in the WB. Regardless, we always use TBS for WB, if you run out of your TBS then you can use PBS for making your blocking buffer so that you could do your experiments running.
For western blot TBS-T (Tris buffer saline with tween 20) is preferred. You can also prepare by you own.
https://www.bio-rad.com/webroot/web/pdf/lsr/literature/Bulletin_6376.pdf
All of them can be used as buffer solutions. But for western blot TBS+0.5% tween 20 is preferred.
Often, a small amount of Tween®20 detergent is added to blocking and washing solutions to reduce background staining, and the buffer is known as PBST or TBST. When choosing between these buffers, it is important to note that TBS/TBST is preferred with AP (Alkaline Phosphatase) labeled antibodies because PBS will interfere with the AP signal.
Please refer to the link below:
https://www.bio-rad-antibodies.com/immunodetection-blocking-antibody-incubation-western-blotting.html
TBS + 0.5% Tween with 5% defatted milke used for preparation of blocking buffer in western blot analysis... while PBS is a general buffer not really used in Western blot analysis blocking buffer preparation...
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