Purification of guinea pig pH 3 migration inhibitory factor.

Abstract

Macrophage migration inhibitory factor (MIF) from the guinea pig resides in 2 discrete and separable proteins referred to as pH 3 MIF and pH 5 MIF. One subfraction of pH 3 MIF was purified to apparent homogeneity from supernatants of stimulated lymph node cells. To monitor purification, biosynthetically radiolabeled MIF was prepared. Sensitized lymphocytes were stimulated in the presence of [3H]leucine by concanavalin A to produce radiolabeled mediators. MIF was purified 30,000-fold from the culture fluid by using gel filtration, sucrose density gradient electrophoresis, isoelectric focusing, and hydrophobic affinity chromatography. This procedure yielded a single 3H-labeled polypeptide with an apparent MW of 35,000 that coincides with MIF activity.