Purification od insoluble protein (TIMP3)
Posted 20 December 2010 - 09:57 AM
I attempted to purify TIMP-3 protein, using protein-A sepharose and elutiing the purified protein with sodium citrate buffer pH3. Then, nutrilizing of eluted protein with either Glycine-OH or Tris-HCl buffer pH9. the protein that eluted and nutrilzed by this way provides cell toxixity. So, Could anyone help me in finding alternative buffers that be safe for cells?
Posted 20 December 2010 - 10:58 PM
Posted 21 December 2010 - 01:11 AM
This protein is highly basic, so it should be eluted with low pH buffer and consequently neutralized at high pH, otherwise TIMP-3 will stick into the tube. I tried to concentrate the protein but I lost most of the protein. Could you guide me to find safe alternatives to the elution and neutralizing buffers.