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Re: [ccp4bb] Cannot running NTA to purify the protein having His-tag? |
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CCP4bb navigationCCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999Subject: Re: Cannot running NTA to purify the protein having His-tag? From: sabrina {- dot -} biarrotte-sorin {- at -} MAIL {- dot -} MCGILL {- dot -} CA Date: 2007-02-28 Hi, I would try to include up to 1 to 2 M NaCl in your lysis buffer and during purification .... you can then decrease your salt concentration in your elution buffer ... Good Luck Sabrina Biarrotte-Sorin Quoting Ngo Duc Tri > Dear CCP4 users, > > I'm purifying a kind of protease having His-tag. The protein is expressed in > insect cells and broken by sonication. > I used NTA resin to purify this protein. > Buffer A is 50mM phosphate buffer pH 7.5 and 300mM NaCl. Buffer B is 50mM > phosphate buffer pH 7.5, 300 mM NaCl and 300 mM Imidazole. > However, all proteins cannot bind to NTA resin. My protein is eluted in > Flow-through. I also check the NTA resin with the control His-tag. The > western blot also shows that my protein has His-tag. > > Do you have any ideas about my problem? I'm really appreciate all of your > advices how to solve this. Thank you very much! > > My best regards, > TriNgo > Sungkyunkwan University > -- Dr Sabrina Biarrotte-Sorin 740 Dr Penfield avenue Room 5400 MONTREAL (QC) H3A 1A4 Canada CCP4bb navigationCCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999 |
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