Lack of immune stimulation by immobilized CpG-oligodeoxynucleotide

Selected phosphorothioate oligodeoxynucleotides containing CpG (CpG-ODN) ac tivate immune responses, including B-cell proliferation and cytokine produc tion. The mechanism by which cells detect CpG-motifs is not known. There ar e conflicting reports in the literature concerning the ability of CpG-ODN l inked to solid supports to stimulate immunity. We prepared a fluorescent, b iotinylated CpG-ODN, a reagent that will support the growth of 7TD1 cells, a murine B-cell hybridoma line that requires CpG-ODN or interleukin-6 (HL-6 ) for survival. Stimulation of 7TD1 cell growth was not reduced by complexi ng biotinylated CpG-ODN to streptavidin, but cell growth was not supported by CpG-ODN coupled to streptavidin-coated latex, magnetic, gold, or agarose beads, A fluorescent CpG-ODN was also covalently attached to cyanogen brom ide-activated Sepharose beads via a 5'-amine group. These derivatized Sepha rose beads did support 7TD1 cell growth, but incubation of the beads with 7 TD1 cells resulted in the appearance of fluorescence within the cells, sugg esting that growth stimulation may be due to CpG-ODN leached from the beads . Our results are consistent with the need for CpG-ODN to be internalized i nto cells to be immunostimulatory.