Expression of IFN-α and IFN-β mRNA in sDCs and BM-DCs.

(A) BM-DCs. DCs were generated by culture of BM cells from B6 mice for 6 days in the presence of GM-CSF. CD11c⁺ cells were purified by means of MACS beads (greater than 98% CD11c⁺), and mRNA was extracted and assayed for IFN-α and IFN-β mRNAs by RT-PCR. As a negative control, 2 μL sterile water was used instead of cDNA template. (B) sDCs. The sDCs were purified from the spleens of B6 mice as described in “Materials and methods” (greater than 98% CD11c⁺) and immediately placed in lysis buffer. After cell lysis, the sample was treated with DNAse. Total RNA was then extracted and assayed for IFN-α and IFN-β mRNA by RT-PCR (left lane). The negative control (right lane) corresponds to 2 μL total RNA subjected to direct PCR (ie, without reverse transcription) using the same IFN-α and IFN-β primers. Amplified products were separated by electrophoresis on a 1.2% agarose gel in the presence of molecular markers (not shown).