I have purified my protein (6kD) from pET28a+. I want to proceed with antibody production against this protein. Will the His Tag have any effect in the antibody production process?
Hi,
His-tag will not hinder in the generation of antibodies against your protein of interest.
You can straight away inject your protein along with the adjuvant into the mammal organism of your choice. Hence, the antibody generated will be polyclonal in origin majorly against many antigenic region of protein and also some against His-tag too.
I hope this addressed your query.
Rajkumar
In the vast majority of cases, it is not necessary to remove the polyHis-Tag from recombinant proteins. The polyHis-tag is widely and preferably used for recombinant immunogen production because of its small size (0.84 KDa) and low immunogenicity. So, it doesn’t interfere with the immune response rised against the protein of interest. Although unusually, His-tag may affect the functionality of the protein. Then, take this into account if you will carried out functional assays and any problem arise. Anyway, my lab partners and I have produced many polyHis-tag recombinant antigens and they always resulted in both very good immunogenicity and functionality. Removing the tags can be a tedious labor and time compsuming, because depending on the proteins sequences, the protease treatments may result in protein degradation or losses. From my own experience, I usually recommend keeping the tag and not to remove it until interference evidences shows up during assays (which can not be solved by other strategy).
Your protein is very small, and you could link to a carrier protein (BSA, KLH, etc.) for do it immunogenic.
You can test the monoclonal antibodies obtained with the strategic of Thamsanqa, is a good method.
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