Isoelectric focusing of IgA and IgM in composite acrylamide-agarose gels

A convenient method for isoelectric focusing of intact polymeric IgA and IgM is described. This technique employed composite gels containing 1.0% acrylamide and 0.75% agarose which exhibited minimal electroendosmotic properties. The spectrotypes obtained with mouse IgA myeloma proteins, a human IgA myeloma and rabbit secretory IgA preparations were compared in three gel systems: 5% acrylamide, 0.8% agarose and the composite gel. With respect to resolution of component bands, the composite gel was superior to the other two systems. Hapten binding studies with MOPC-315 IgA and a rabbit secretory IgA anti-DNP antibody indicated that the focused IgA molecules retained their binding site integrity in the composite gel. The pI ranges obtained with microscale sucrose isoelectric focusing and composite gel system showed good correspondence, with the latter exhibiting enhanced resolution. Studies with MOPC-104E IgM revealed improved resolution in the composite gel when compared to the agarose system. Comparison of pI ranges for IgA and IgM immunoglobulins obtained in the present study with those reported previously suggest that IgA spectrotypes are confined to an acidic pI range (3.4--6.4), whereas IgM spectrotypes are not (4.3--8.8).