IL-3 production by stimulated 185-4 cells. (A) The 185-4 cells were cultured with medium alone, aggregated IgG (50 μg/mL), calcium ionophore A23187 (250 ng/mL), PMA (100 ng/mL), or a combination of A23187 and PMA. After 3 hours, culture supernatants were dialyzed. IL-3 was measured in each sample by a bioassay using IL-3–dependent cells as described in Materials and Methods. Data represent the means ± standard deviation (SD) of five replicates. (B) RT-PCR analysis of IL-3 production by 185-4 cells. The 185-4 cells were cultured with each stimulator for 3 hours. Total RNA (5 μg) from the cells was reverse transcribed and PCR amplified as described. PCR products were resolved in 2% agarose gel and visualized by staining with ethidium bromide.