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树突状细胞 (dendriticcells,DCs)作为专职的抗原递呈细胞 ,已被广泛地应用于临床的肿瘤疫苗治疗。目前的临床方案大多为分次给病人注射 >10 6 个细胞 /次 ,不同批次培养的DCs,连续 4 - 6周。为了提高疗效 ,简化治疗程序及规范疗程 ,有必要将DCs低温保存 ,使患者在治疗过程中得到同批次的培养的DCs。本实验从患者外周血采取单个核细胞 ,经细胞淘洗分离单核细胞 ,在 80 0U/mlGM CSF + 10 0ng/mlIL 13的培养条件下 ,将单核细胞于Teflon疏水袋中诱导产生DCs。将DCs以 - 1℃梯度降温及不控温两种方式将DCs冻存于 - 80℃及液氮中。 1个月后 ,4 2℃快速复温。检测其免疫表型 (CD1a ,CD14,CD4 0 ,CD80 ,CD83,CD86,CD5 4 ,CD5 8,CD16,CD32 ,CD64,HLA DR)及其对自体及异体淋巴细胞的刺激作用。结果表明 ,2种方式的低温保存及复温过程不改变DCs的免疫表型及对淋巴细胞的刺激功能。结论 :应用Teflon袋 ,能够从外周血单核细胞中诱导出大批量的DCs ,这些DCs可被低温保存而不改变其功能 ,为其临床应用奠定了基础
Dendritic cells (DCs), as full-time antigen presenting cells, have been widely used in the clinical treatment of tumor vaccines. Most of the current clinical programs for the injection of patients with graded> 106 cells / time, different batches of DCs cultured for 4 to 6 weeks. In order to improve the curative effect, simplify the treatment procedure and standardize the course of treatment, it is necessary to keep the DCs cryogenic, so that the patients receive the same batch of cultured DCs during the treatment. In this experiment, mononuclear cells were taken from the peripheral blood of patients. Mononuclear cells were isolated by panning, and monocytes were induced to produce DCs in Teflon hydrophobic bags under the condition of 80 0U / ml GM CSF + 10 0 ng / ml IL-13. DCs were cryopreserved at -80 ℃ and liquid nitrogen in two ways: temperature gradient of -1 ℃ and no temperature control. After 1 month, 4 2 ℃ rapid rewarming. The immunophenotypes (CD1a, CD14, CD4o, CD80, CD83, CD86, CD54, CD5 8, CD16, CD32, CD64, HLA DR) and their stimulation on autologous and allogeneic lymphocytes were detected. The results showed that the two ways of cryopreservation and rewarming did not change the immunophenotype of DCs and the stimulation of lymphocytes. CONCLUSIONS: The use of Teflon bags induces high-volume DCs from peripheral blood mononuclear cells, which can be cryopreserved without changing their function, providing a basis for their clinical application