A novel method of screening for immunomodulating substances, establishment of an assay system and its application to culture broths of microorganisms.

A novel method of screening for immunomodulating substances is developed employing lymphocytes and three mitogens. Concanavalin A (Con A) and phytohemagglutinin (PHA) are applied as T cell specific stimulants and lipopolysaccharide (LPS) as a B cell specific stimulant respectively. The lymphocytes obtained from mouse spleen are cultured with an antibiotic or a sample extract in the presence or absence of mitogen for three days and pulsed with [3H]thymidine for five hours before harvest. Differential effects of a sample compound on [3H]thymidine incorporation by the activated and quiescent lymphocytes are scored. In this procedure most of the tested antibiotics or chemical compounds with different mode of actions show non-specific effects. Cyclosporin A, a potent immunosuppressive substance, suppresses both Con A and PHA responses more extensively than LPS response and quiescent cell growth, and two cytochrome bc1 complex inhibitors, funiculosin and antimycin A3, are less suppressive to PHA response than to the others. The present system was also applied to the methanol extracts of the culture broths prepared from the type strains of Actinomycetes and Penicillium.