His-tag protein purification - contamination with other...
Posted 04 November 2004 - 12:25 AM
Posted 04 November 2004 - 04:36 AM
I would also use gel filtration as the final step in your purification method. You may need to concentrate your sample afterwards though, since the fractions are usually quite dilute.
Edited by gmcg, 04 November 2004 - 04:39 AM.
Posted 04 November 2004 - 05:59 AM
Posted 04 November 2004 - 06:13 AM
Posted 04 November 2004 - 11:41 PM
Posted 05 November 2004 - 05:39 AM
Posted 08 November 2004 - 07:57 PM
Posted 08 November 2004 - 08:11 PM
Posted 08 November 2004 - 10:22 PM
Posted 16 November 2004 - 12:26 PM
Are your His tag is on C-ter or N-ter of your protein. If your His-tag is on N-ter, I think your protein is troncated or the transcription was partially completed or you have a degradation problem. Did you have an antibody for your protein? (Not a antibody for 6xHis). Try a western blot on your eluate. If you get a lot of band.
In all case it is better to have His-tag at the end if your protein.
Posted 17 November 2004 - 12:00 AM
Posted 22 November 2004 - 12:57 AM
Posted 22 November 2004 - 07:58 AM
I have another question about his-tag protein. Do you know if is possible to purifie proteins witha His-tag in the middle of the protein? Any experiences?
I think that the principal problem is that the His-groups are not accesible to the column, and purificacion would be very poor.
Posted 25 November 2004 - 09:39 PM
I am having a similar problem with my recombinant protein. I was wondering what brand imidazole you all use
Thank you for any suggestions