I am working with purification of protein by strep tag. In my protocol there is avidin in the binding buffer. Right now I do not have it. Can I proceed with the purification without adding avidin. What will happen if I do not avidin in my buffer?
Hi,
I am not an expert at this but avidin is binding biotion. Biotin is know to irreversibly block the binding sites of Strep-Tactin. So if you have biotin in your protein sample it will block all the binding sites and your strep tag will struggle to find an "open" binding site.
You could remove the biotin by dialysis, then you would not need to add the avidin.
I hope this is helping you with your question.
You can have a closer look at the Quiagen "Strep-tagged Protein Purification
Handbook" It has a troubleshoot section.
http://wolfson.huji.ac.il/purification/PDF/Tag_Protein_Purification/StrepTag/QIAGEN_StreptaggedProteinPurif.pdf
Hi,
I am not an expert at this but avidin is binding biotion. Biotin is know to irreversibly block the binding sites of Strep-Tactin. So if you have biotin in your protein sample it will block all the binding sites and your strep tag will struggle to find an "open" binding site.
You could remove the biotin by dialysis, then you would not need to add the avidin.
I hope this is helping you with your question.
You can have a closer look at the Quiagen "Strep-tagged Protein Purification
Handbook" It has a troubleshoot section.
http://wolfson.huji.ac.il/purification/PDF/Tag_Protein_Purification/StrepTag/QIAGEN_StreptaggedProteinPurif.pdf
Hello,
Nothing too drastic should happen if you don't add avidin in your binding buffer. Generally it is used to avoid non-specific binding of contaminants. The worst could happen that your elution might not be super clean. But just give it a try even without it. I recently purified a protein without avidin in the buffer and the elution was just fine!
Good luck
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