Tumor antigen loading DC
SK-N-SH cells in logarithmic phase were resuspended and adjusted to
2×107/ml concentration. Cells were quickly frozen to
-80℃, then rewarmed in 37℃ water. After four cycles, cells suspension
was centrifuged with 10,000 r/min for 15 minutes and the supernatant was
collected and cryopreserved at -80℃. The lysate equivalent to
2×106 tumor cells was added into per millilite rmedium
of DCs and cultivated for 6 days. Four hours later, cells suspension was
added rmTNF-α and cultivated to 7th day. Then, the suspension cells were
collected and defined tumor antigen loaded DCs. The cell morphology of
DCs was observed by phase contrast microscope. Meanwhile, the expression
rate of CD11c, CD86 and MHC-II were detected by flow cytometry.