Hi everyone,
I have been looking on this bb and other websites as well but I could not find
a veredict. We are suspecting that when I elute my sample from my Ni-NTA
column, the imidazole concentration (250 mM) is making it to precipitate. Once
my sample has a cleavable TEV site, I was planning to incubate my loaded resin
overnight with TEV and get my sample back simply using my lysis buffer. And
here lies the problem. Most of the TEVs are kept in EDTA and DTT and I wonder
if they are essential for its protease activity or if I could use another
reducing agent more compatible with my resin (or maybe do not add both). I saw
that someone did not have EDTA and used b-mercap. instead of DTT. May I have
your comments if you guys already faced a similar situation?
Best wishes
______________________________________________________
Rafael Marques da Silva
PhD Student – Structural Biology
University of Leicester
Mestrando em Física Biomolecular
Universidade de São Paulo
Bacharel em Ciências Biológicas
Universidade Federal de São Carlos
phone: +55 16 99766-0021
"A sorte acompanha uma mente bem treinada"
________________________________________________
########################################################################
To unsubscribe from the CCP4BB list, click the following link:
https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1
This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list
hosted by www.jiscmail.ac.uk, terms & conditions are available at
https://www.jiscmail.ac.uk/policyandsecurity/
