Differences in IFNαproduction by pDCs from different lineages. (A) Total BM cells, CMPs, CDPs, or CLPs from C57BL/6 BM were cultured with FL and CD45.1 total BM feeder cells. After 5 days, pDCs were sorted as CD11c⁺BST-2⁺ and cultured with CpG2216. After 20 hours, supernatant was harvested and analyzed by ELISA for the presence of IFNα. (B) CLPs and CMPs were cultured and the resulting pDCs at day 5 were isolated as in panel A. pDCs were recultured in medium collected from day-7 FL BM cultures with CpG2216. After 20 hours, supernatant was harvested and analyzed by ELISA for the presence of IFNα and pDCs were analyzed by flow cytometry for CD69 expression and viability by propidium iodide exclusion. At the time of harvest, 67% of CMP-derived pDCs but only 28% of CLP-derived pDCs remained viable. (C) CLPs were isolated from C57BL/6 BM and transferred into irradiated CD45.1 recipients. After 14 days, pDCs were sorted as CD11c^intBST-2⁺ and cultured with CpG2216. After 20 hours, supernatant was harvested and analyzed by ELISA for the presence of IFNα. Results are pooled from 4 independent experiments (A) and from 3 independent samples from 1 experiment (B-C). Error bars show means ± SEM.