Isolation of pure IgG1, IgG2a and IgG2b immunoglobulins from mouse serum using protein A-Sepharose

A simple and rapid method for isolating pure mouse IgG1, IgG2a and IgG2b immunoglobulins in nearly 100% yield is described. Mouse serum was fractionated on protein A-Sepharose 4B and the recovery of immunoglobulins was measured as a function of pH. When serum was applied to the column at pH 8.0, IgM, IgA and IgE were almost quantitatively recovered in the effluent together with non-immunoglobulin serum components. Providing the binding capacity of the column was not exceeded, essentially all IgG was retained at pH 8.0 and this could not be eluted by washing. Using buffers of decreasing pH, IgG1, IgG2a and IgG2b were sequentially eluted at pH 6.0–7.0, pH 4.5–5.0 and pH 3.5–4.0, respectively.