[Selection of storage time, temperature and anticoagulants of peripheral blood samples for culturing cytokine-induced killer cells].

OBJECTIVE

To investigate the effects of anticoagulants, preservation time and temperature of peripheral blood samples on the culture of cytokine-induced killer (CIK) cells so as to provide the experimental evidences for peripheral blood storage in autologous immunotherapy.

METHODS

Four samples of 60 mL peripheral blood were collected. After being added with heparin sodium, cell preservation liquid (sodium citrate) and EDTA solution, they were separately stored under 4°C, 22°C, and 30°C for 0, 4, 8, 24 hours. We divided the orthogonal experiments into 12 groups, and then separated mononuclear cells and induced them into CIK cells. The proliferation efficiency and IFN-γ secretion were compared in the 12 groups.

RESULTS

The proliferation of CIK cells was not obvious in EDTA group, but obvious in heparin sodium group and sodium citrate group, especially better in sodium citrate group. The storage time of blood did not have a significant impact on CIK cell culture, however the longer storage time, the lower cell proliferation efficiency. The proliferation efficiency decreased apparently after 16-day culture if preservation time exceeded 8 hours. After the culture period of 16 days, the efficiency of CIK cell proliferation was the highest at 22°C, followed by that at 4°C, and the lowest was at 30°C. It was not apparent that the temperature of blood storage affected CIK cell proliferation within 16-day culture period.

CONCLUSION

Both heparin sodium and sodium citrate can be used in blood sample anticoagulation for CIK cell culture. Blood samples are suitable for CIK cell culture which are stored within 24 hours between 4°C and 30°C.