Serum specifically potentiates the mitogenic response of Nb2 lymphoma cells to rat prolactin

Previous observations in our laboratory indicated that rat serum samples being bioassayed for PRL with the use of Nb2 lymphoma cells produced mitogenic responses greater than maximally effective doses of purified rat PRL. The present studies were conducted to confirm these observations and to determine the possible serum factor or factors responsible for the potentiated response of serum. Twenty five rat serum pools prepared from blood samples obtained from lactating, ovariectomized, and steroid-treated rats were assayed in duplicate aliquots of 0.3-50 microliter against NIDDK-RP-1 rat PRL (11 IU/mg) by the Nb2 bioassay in Fischer's Medium containing 10% horse serum and by conventional double antibody RIA. The mitogenic response of 18 of the 25 pools were clearly greater than the response to standard rat PRL when included in the bioassay at 10-50 microliter/ml of cells. When less than 10 microliter serum pools were assayed, parallelism to the standard was observed; but the concentrations of PRL determined by bioassay were 82 +/- (SE) 4% of the levels determined by RIA in samples from ovariectomized, steroid-treated rats stored for two or four weeks and 62 +/- 2% in samples from lactating rats stored for 3-6 months. Horse serum and hypophysectomized rat serum also potentiated the mitogenic responses to 1 or 5 ng standard rat PRL when added at 10-150 microliter/ml of culture medium that already contained 100 microliter (10%) horse serum. No potentiation of the RIA was observed with rat or horse serum. Insulin, proinsulin, or C peptide of proinsulin (0.02 ng-2 micrograms/ml); fibroblast growth factor, nerve growth factor, epidermal growth factor, insulin-like growth factors I or II (0.002-200 ng/ml), T cell growth factor (TCGF) (0.0002-20 half-maximal units/ml) or platelet-derived growth factor (0.002-10 half-maximal units) did not potentiate the responses to 1 or 5 ng/ml standard rat PRL. Of the growth factors tested, only TCGF was mitogenic to Nb2 cells when placed alone in the medium, but this mitogenic effect of TCGF was not potentiated by horse serum. We conclude that serum can potentiate the mitogenic effect of PRL on Nb2 cells grown in Fischer's medium.