Development of a serum-free medium for lymphocyte cultures.

开发用于淋巴细胞培养的无血清培养基。

• 批准号: 63550728
• 负责人: OHMORI Hitoshi
• 金额: $ 0.96万
• 依托单位: Okayama University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1988
• 资助国家: 日本
• 起止时间: 1988 至 1989
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-63550728/
• 关键词: Murine lymphocytes; Primary culture; Antibody response; Serum-free medium; T cells; beta-Cyclodextrin; 無血清培地; リンパ球の無血清培養; 抗体応答

The regulatory mechanisms of immune responses have been investigated by using in vitro culture systems of murine lymphocytes. Fetal calf serum(FCS)-containing culture medium has been usually used in these cultures. But, it has often been pointed out that the magnitude and the properties of immune responses varied markedly depending on the lot of FCS used. In order to overcome these problems, we attempted to develop a serum-free medium that is able to induce a primary antibody responses as effectively as the serum-containing medium.1. Based on the discovery that beta-cyclodextrin is effective as a serum-substitute, we first devised a basal serum-free RPMI-1640 medium that was supplemented with beta-cyclodextrin, albumin, transferrin and insulin. The addition of ehter-soluble fraction of FCS resulted in a marked enhancement of the immune response. The active substance was purified and identified as cholesterol. In addition, low density lipoprotein (LDL) was found to be more effective than cholesterol itself.2. The screening was carried out to find substances that ard useful to further improve the efficiency of the above medium. We found that the simultaneous addition of L-alanine and putrescine was the most effective. By determining the optimal concentrations of these additives, we finally succeeded in establishing a serum-free medium that can induce a primary antibody response as effectively as FCS-containing culture medium.3. This serum-free medium was also applicable to investigate the proliferation and the differntiation of T cells, suggesting a wide applicability of this medium in various lymphocyte culture systems.

通过使用鼠淋巴细胞的体外培养系统研究了免疫反应的调节机制。胎牛血清（FCS）含有含有培养基的培养基通常在这些培养物中使用。但是，经常指出，免疫反应的幅度和特性明显取决于所用的FC。为了克服这些问题，我们试图开发一种能够像含血清的培养基一样有效地诱导主要抗体反应的无血清培养基1。基于发现β-环糊精作为血清替代物有效的发现，我们首先设计了一种无血清的RPMI-1640培养基，该培养基辅以β-蛋白丝蛋白，白蛋白，转移丝蛋白和胰岛素。 FCS的溶质分数的添加导致免疫反应的明显增强。将活性物质纯化并鉴定为胆固醇。另外，发现低密度脂蛋白（LDL）比胆固醇本身更有效2。进行筛选以找到对进一步提高上述培养基效率有用的物质。我们发现，同时添加L-丙氨酸和腐烂是最有效的。通过确定这些添加剂的最佳浓度，我们最终成功地建立了可以像含FCS的培养基一样有效地诱导原代抗体反应的无血清培养基。3。该无血清培养基也适用于研究T细胞的增殖和不同，这表明该培养基在各种淋巴细胞培养系统中的广泛适用性。

项目成果

Hitoshi Ohmori: "Development of a serum-free medium for in vitro immune response by using β-cyclodextrin.Demonstration of the requirements for polyamines." Journal of Immunological Methods. 112. 227-223 (1988)

Hitoshi Ohmori：“使用 β-环糊精开发用于体外免疫反应的无血清培养基。多胺的要求证明。”《免疫学方法杂志》112. 227-223 (1988)。

Hitoshi Ohmori: "Development of a serum-free medium for in vitro immune response by using beta-cyclodextrin. Demonstration of the requirements for polyamines." Journal of Immunological Methods. Vol.112. 227-233 (1988)

Hitoshi Ohmori：“使用 β-环糊精开发用于体外免疫反应的无血清培养基。证明对多胺的要求。”

Hitoshi Ohmori: "Demonstration of the requirements for cholesterol or low density lipoprotein in the antibody response of murine lymphocytes in serum-free medium." Agricultural and Biological Chemistry. 53. 209-213 (1989)

Hitoshi Ohmori：“演示无血清培养基中小鼠淋巴细胞的抗体反应对胆固醇或低密度脂蛋白的要求。”

大森斉: Journal of Immunological Methods. 112. 227-233 (1988)

大森仁：免疫学方法杂志 112. 227-233 (1988)

Hitoshi Ohmori: "Demonstration of the requirements for cholesterol or low density lipoprotein in the antibody response of murine lymphocytes in serum-free medium." Agricultural and Biological Chemistry Vol.53 pp.209-213 (1989).

Hitoshi Ohmori：“演示无血清培养基中小鼠淋巴细胞的抗体反应对胆固醇或低密度脂蛋白的要求。”