The Fractionation of Antigen‐Dependent Macrophage Migration Inhibition and Macrophage Activation Factors from Lymph Draining a Tuberculin Reaction

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Abstract
Efferent lymph, taken from BCG‐sensitized sheep 6 to 20 hr after inaction of PPD, was fractionated by chromatography on Sephadex G‐200 and on Sepharoseconcanavalin A and by rechromatography on Sephadex G‐200 The fractions were examined for their ability to inhibit the migration of normal guinea pig peritoneal exudate cells (MIF assay) and to stimulate the uptake of colloidal gold by these cells (GUS assay). Antigen‐dependent MIF activity and GUS activity eluted from Sephadex G 200 in the albumin fraction. No corresponding activities were found in this region when control lymph was fractionated in an identical manner. After chromatography on Sepharoseconcanavalin A, antigen‐dependent MIF activity was associated with a glycoprotein fraction containing IgG. After rechromatography on Sephadex G‐200, antigen‐dependent MIF and GUS activities eluted at a position corresponding with that of proteins of 60,000 to 70,000 molecular weight, and these fractions contained less than 2.5 μg/ml IgG. Soluble mediators produced in vivo by sheep with a delayed hypersensitivity response to PPD possessed antigen dependent MIF activity. The antigen‐dependent macrophage‐inhibiting and macro phage‐activating factors were closely associated in fractions that did not appear to contain significant amounts of IgG.