RAPID ONE-STEP PURIFICATION OF GOAT IMMUNOGLOBULINS BY IMMOBILIZED METAL-ION AFFINITY-CHROMATOGRAPHY

A rapid, single step purification of immunoglobulins from goat serum w as achieved using immobilized metal ion affinity chromatography (IMAC) on a new high capacity gel, Novarose, coupled to tris(2-aminoethyl)am ine (TREN) chelated with copper. When goat serum was adsorbed to this gel in buffer pH 7 at 11 cm/h (8.6 ml/h), the immunoglobulin fraction was recovered in a decreasing linear pH gradient at about pH 5.5. When the adsorption buffer was adjusted to pH 6.0 and the linear velocity increased to 110 cm/h (221 ml/h), an immunoglobulin fraction of greate r than 95% homogeneity was obtained. Protein purity was assessed by si lver-stained native and sodium dodecyl sulfate polyacrylamide gel elec trophoresis (SDS PAGE). The capacity of the gel for immunoglobulins wa s 17 mg immunoglobulin/ml at the low flow rate with adsorption at pH 7 and 15 mg immunoglobulin/ml at the high flow rate with adsorption at pH 6. No problems of back pressure or gel compression were observed at the higher linear velocity. The mild elution pH, high flow rate, and synthetic nature of the ligand support make this new metal-chelating g el a powerful alternative to the use of other currently available comm ercial gels commonly used for immunoglobulin purification.