Immunosuppressive dendritic cells were isolated following leukapharesis by antisense oligonucleotides targeting CD40, CD80 and CD86.
Monocytes from Leukopack were separated by elutriation in a closed ELUTRATM system and then cultured in a closed Aastrom Replicell System in the presence of IL-4 and GM-CSF. QC-tested phosphorothioate-modified antisense oligonucleotides targeting the 5' end of the CD40, CD80 and CD86 gene primary transcripts were added to the cells in the Aastrom Replicell System at a final concentration of 3.3 micromolar. The sequences of each of the antisense oligonucleotides was: CD40: 5’ act ggg cgc ccg agc gag gcc tct gct gac 3’; CD80: 5’ ttg ctc acg tag aag acc ctc cca gtg atg 3’; CD86: 5’ aag gag tat ttg cga gct ccc cgt acc tcc 3’. On day 6, the cells were harvested and checked for viability and purity.