In a lab I rotated in, we always reused primary antibody but never reused secondary. In the lab I've chosen to remain in, it is considered okay to reuse secondary antibodies. I am a bit hesitant to do this. We store the antibody cocktail in PBST in 4 degree refrigeration, and the secondary is wrapped in foil.
The other question on research gate concerning this is specific to Westerns, and I'm concerned specifically with immuno.
Hi Hope,
First of all, the price of secondary Ab is more cheaper than primary Ab, so I usually used the secondary Ab for just one time in all experiments.
Second, catalytic enzyme (e.g. HRP) is conjugated to secondary Ab, there is risk for losing function of color development if you stored secondary Ab in 4 degree refrigeration for a period of time.
On the other hand, reuse primary Ab for performing WB is okay, but for the IHC, you may need to try. Keep in mind for storing the Ab in solution with NaN3 to prevent contamination.
Hope this help.
I would never do that. You can't be sure that the concentration / activity of a reused solution is comparable to the fresh one. If you are only interested in a YES/NO signal it may be ok but but only as a very last option
Hi Hope. This depends on the quality of your antibodies. If they are from a well-known manufacturer with high specificity and sensitivity, you can recycle the antibody dilutions. I have found that my cheap secondary antibody can be used once only, and must be prepared fresh everytime. Meanwhile my more expensive secondary antibody (twice the price than the first one) can be recycled up to three times (max). Also, you must practise aseptic techniques while preparing these dilutions to avoid contamination, i.e. sterile tubes, sterile pipettes, sterile TBST/PBST. In addition, I dilute my antibodies in skimmed milk, so this helps to check for potential contamination as well, because you can see if the milk is spoilt by their colour and smell.
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