Production and assay of ovine T cell growth factor by concanavalin A stimulated peripheral blood leukocytes

Conditions for the production and assay of ovine T cell growth factor (TCGF) activity were evaluated. Peripheral blood leukocytes were stimulated with concanavalin A (Con A) in the presence of 2% autologous serum or serum-free media. Supernatants were harvested after 30 hr and concentrated for further characterization. A 28 hr proliferation assay with 2.5 X 10(4) 24 hr Con A blasts per well was optimal for detection of TCGF. Peak TCGF activity occurred with a 30-37 KD molecular weight fraction. Production and assay of TCGF were performed under autologous conditions to reduce background stimulation which occurred when fetal bovine serum was present. This methodology required no cell lines or inbred animals and should be adaptable to the study of immunostimulatory molecules of other outbred species.