A method for the production and quantitative assay of human lymphokine preparations

A procedure for the preparation and assay of a human lymphokine supernatant is described. Tonsillar lymphocytes at a concentration of 2 X 10(7)/ml in a serum-free medium are incubated for 2 h with PHA-P, washed free of unbound mitogen and incubated for a further 17 h. The supernatant is harvested and concentrated, and the active protein fraction is absorbed to and eluted from a column of hydroxylapatite. The eluate is desalted and sterile filtered. The preparations contain active material released from 10(8) cells in 1 ml and do not contain residual PHA-P. The most active preparations are obtained using PHA-P as the mitogen and at concentrations of 25-50 microgram/ml during the 2 h pulsing period. Activity is assessed by a quantitative assay based on the maintenance of mitogen activated blast cells in culture for 2 days.