I am trying my luck in HEK293T cells, by transfection. I can express and detect Protein post transfection. But since my construct is untagged what methods can be employed tro prify it (keeping in mind that the yield after such transfections is low anyways)
thanks!
There are a lot of options depending on the size of your protein and what downstream applications you want to do. Could you provide more information on your current experiment and future needs?
Hi Jordan.
Well I have not puit any tags in the construct so that it doesntinterefere with the folding. however, now i plan to use the translated Protein for spectroscopy. I doubt i will have a large Background even if I go by Size exclusion chromatography.
what is your Suggestion?
Immunoprecipitation is a good suggestion. However, If you want to repeatedly purify the protein, you could go for immunoaffinity chromatography. These two methods are only possible when you have good affinity antibody. Otherwise, you may try several other separation methods (Ion exchange chromatography, hydrophobic chromatography, and finally size exclusion chromatography). But before I suggest any of these methods, the nature and the properties of the protein should be known to me. If you already know the properties of your protein, please go through these suggestions, you might figure it out the right method to purify the protein yourself.
Hope this information helps.
Hi Senthil
Thanks for the suggestions.. I have highly specific antibody to my Protein and yes I can try Immunoaffinity chromatography. I will appreciate if you also have some protocls to share!
Cheers
Dear all
Thanks fr great suggestions. I am trying pierce coIP kit (small scalebatch purification)
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