Preparation and properties of cloning inhibitory factor. II. Factors affecting its production and assay

Cloning Inhibition Factor (CIF), an activity present in PHA or antigen stimulated lymphocyte culture supernatants, inhibited the cloning of HeLa cells when diluted 1:9 in HeLa culture medium. CIF was not detectable at 8 hr, was maximal at 24–48 hr, and declined with longer periods of lymphocyte culture. CIF production increased with lymphocyte concentration up to 1–2 × 10⁶ lymphs/ml but plateaued at higher concentrations. At lower lymphocyte concentrations, more CIF activity was present when lymphocytes were cultured in 5% rather than 12% serum. PPD elicited similar CIF activity from either highly purified or unpurified lymphocytes. CIF activity was independent of HeLa medium serum concentration. It remained stable for 3–6 months at ?20 °C, but was inactivated by heating at 56 °C for 30 min. At a 1:9 dilution CIF was not cytocidal but produced cytopathic changes. CIF shares many properties with, and may be identical to, Proliferation Inhibitory Factor.