Agarose gel technique with human peripheral blood leukocytes as migrating indicator cells was used to demonstrate migration inhibition factor (MIF) in cell-free supernatants from cultures of tuberculin-stimulated human peripheral blood mononuclear leukocytes. In this two-step MIF agarose assay only a few hundred microliters of supernatant were used. MIF was produced if cells from Mantoux-positive persons were cultured with purified protein derivative of tuberculin (PPD), but also if the sensitive cells after preincubation with PPD were washed and cultured in medium without PPD. There was a good correlation between the migration inhibitory effect of supernatants from PPD-stimulated cultures and the Mantoux reaction. The two-step MIF agarose assay was more sensitive than the direct leukocyte migration agarose test and makes possible the testing of several pairs of supernatants and a standard of MIF activity on the same migrating indicator cell population.

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This study was supported by grants from the Danish Medical Research Council.

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