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Soluble anti-TCRγδ stimulates expansion of human peripheral blood γδ T cells and its culture and storage
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Abstract:
Objective: To compare the different protocols for amplifying human peripheral blood γδ T cells and establish the suitable expansion and storage conditions for clinical application.Methods: Human peripheral blood mononuclear cells were isolated and stimulated by immobilized or soluble antiTCR γδ, and further cultured in RPMI 1640 supplemented with newborn bovine serum, RPMI 1640 supplemented with human AB plasma, or serum free AIM V medium separately. Cell numbers and survival rates were detected by trypan blue staining; purities and phenotypes of the expanded γδ T cells were measured by immunofluorescence staining; cytotoxicities of γδ T cells against different tumor cell lines were detected by lactate dehydrogenase (LDH) method. Results: RPMI 1640 supplemented with 15% newborn bovine serum or 5% human AB plasma had similar effects on the proliferation of γδ T cells but better than that of AIM V serum free medium. Soluble antibody stimulated the expansion of peripheral blood γδ T cells as effectively as immobilized antibody did. The expanded γδ T cells had complete T cell receptor (TCR) pool, containing TCR Vδ1 and Vδ2 subtypes, and displayed cytotoxic activities aganist lung cancer, hepatocellular carcinoma, and ovarian carcinoma cells in vitro. These γδ T cells, when suspended in normal saline supplemented with 0.25% human albumin or 1% human AB plasma, could maintain 95% survival for 12 hours at 4 ℃. Conclusion:Human peripheral blood γδ T cells can be effectively expanded in vitro by soluble antiTCR γδ in RPMI 1640 culture medium containing human AB plasma, and these expanded γδ T cells can be stably preserved in normal saline supplemented with plasma or albumin.
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Supported by the National High Technology Research and Development Program of China (No. 2007AA021109)
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