Dear Amala,
1. Increase your buffer pH. As a rule of thumb, usually it is
recommended to work with pH 7.6 and above for Ni purification. Try pH 8.
2. Increase your buffer A [NaCl] to 500mM. Some proteins are more stable
in higher NaCl, and for many, 50mM is not enough. It is possible your
prote
I am trying to purify a protein using HIS-select Ni-affinity column
(washing with 50mM Tris pH7.5, 50mM NaCl, 30mM imidazole, 3mM BME,
5%glycerol and elution with the same buffer + 250mM imidazole, 500mM Nacl).
The protein (pI= 6.04) becomes cloudy/precipitated within 15minutes of
after elution. I
Dear all
yes my paalet is large enough and no my protein is not soluble ,and I am using
37degrees before and after induction with IPTG,and the vector that I am using
is pRSETA 3.2kd
Best Regards
Rana
From: Kelly Daughtry
To: rana ibd
Cc: CCP4BB@jiscmail.a
