Removal of bacterial endotoxin in a protein solution by immobilized metal affinity chromatography

An isolated and purified Gram negative bacteria protein obtained by a method for removing bacterial endotoxin from a Gram negative bacterial protein solution, wherein the method uses immobilized metal ion affinity chromatography (IMAC) comprising the steps of:

• (a) applying the Gram negative bacteria protein solution to an IMAC resin, wherein the resin is equilibrated in a buffer;
• (b) eluting the endotoxin with the buffer of step (a), wherein the protein remains bound to the resin;
• (c) eluting the protein in an elution buffer comprising glycine;
• and
• (d) collecting the eluted protein,
• wherein the concentration of the endotoxin in the protein solution is effectively reduced by a factor of at least 200 and protein recovery is greater than at least 60%.