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Protein G Purification of Antibodies

1. Reagent and Materials(1) Hi-Trap Protein G Column (Pharmacia Biotech #17-0404-01)(2) 20 mM Sodium Phosphate Buffer, pH 7.01.084 g NaH2PO4, anhydrous3.273 g Na2HPO4.7H2Oq.s. to 1 liter with di-H2O(3) 0.1M Glycine, pH 2.83.75 g Glycine1.4 ml HCl

Coupling Antibodies to Protein A or G

1. use 2 mg of antibody per ml wet beads (use appropriate antibody/protein A or G combination). 2. mix antibodies with beads and bind at room temperature for at least 1 hr (on roller). 3. wash the beads twice with 10 volumes borate buffer, spin

Purification of Monoclonal Antibodies Using Protein A/G

A major breakthrough in immunology came with the discovery that large amounts of relatively pure monoclonal antibodies (mAbs) could be prepared from the fusion of B cells (secreting the relevant mAb) with a nonsecreting myeloma cell line