If tween 20 is a detergent that washes of antibodies, why is it sometimes used in blocking buffer. Does its presence prevent nonspecific binding exclusively? Would it not reduce the amount of specific binding and weaken a band?
Thank you.
Yes, it could reduce the amount of specific binding. However, antibody interactions with non-target proteins have a weaker binding affinity. If an optimum concentration of tween is used (I typically use 0.1% tween, but it can vary depending on your antibody) then the background will be reduced without losing too much of your specific signal.
PBST and TBST is essentially used in Blocking buffer to perform western blotting.
blocking buffer or Milk solution is used to prevent the non specific binding of primary antibody on the membrane.! PBST plays a signinficant role in maintaining the epitope of our desired protein on the membrane.
TBST is preferrerd over PBST.
-- Best regards.
I would reccomend you to read this.
https://www.thermofisher.com/in/en/home/life-science/protein-biology/protein-biology-learning-center/protein-biology-resource-library/pierce-protein-methods/blocking-buffers-western-blot-elisa.html
Hi there,
The concentration of Tween 20 used in all buffers used for WB (usually 0.1%) is meant to prevent/limit the non specific binding of antibodies to the membrane without affecting specific interactions.
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