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Agarose isoelectric focusing of native human immunoglobulin M and alpha 2-macroglobulin.

The resolution of native 19S IgM and alpha 2-macroglobulin by agarose isoelectric focusing is described. The agarose used was practically charge free, thus avoiding disadvantages of electroendoosmosis, and gives a very large network gel, with minimal molecular sieving effects. The resolving power is comparable to that obtained in thin-layer isoelectric focusing in polyacrylamide gel. Clones of human IgM showed a microclonal heterogeneity, similar to IgG antibody heterogeneity, alpha 2-macroglobulin gave a pattern of seven bands in the pI range of 4.1 to 4.9. The flexibility of the agarose isoelectric focusing (IEF) system with regard to immunodetection techniques is illustrated by the use of immunofixation and two-dimensional crossed immunoelectrofocusing. The agarose IEF method has several advantages, viz., non-toxicity, simple handling, uniform and rapid gel formation, and considerably shortened fixing and staining times. The value of the new method is discussed, in particular its usefulness in detecting and isolating IgM antibodies of known specificity produced by cells in culture.