Birkeland S A
Cryobiology. 1976 Aug;13(4):442-7. doi: 10.1016/0011-2240(76)90100-0.
The influence of a series of factors on the frozen storage of lymphocytes was investigated. The cells were frozen using different freezing programmes, using the cryoprotectants dimethyl sulphoxide and glycerol in different concentrations in the freezing medium, and with variations in the period of exposure of cells to cryoprotectants before freezing and after thawing. Cell viability was evaluated by quantitative measurements of the cell-mediated immune response after stimulation with phytohemagglutinin, pokeweed mitogen, concanavalin A, and allogenic cells in mixed lymphocyte cultures. The factors investigated were found to have an important effect on the immune response, so that careful investigation and exact specification of the freezing system are necessary before frozen cells are used in blast-transformation tests. The best freezing programme had a duration of approximately 40 min with a smooth progression through the temperature range where phase transition takes place. The optimum dimethyl sulphoxide concentration in this programme was 8--10%. Dimethyl sulphoxide had no toxic effect on the cells, and no equilibration period was necessary prior to freezing. An equilibration period of 15 min with 10% glycerol was even better than the optimum programme with dimethyl sulphoxide.
研究了一系列因素对淋巴细胞冷冻保存的影响。使用不同的冷冻程序对细胞进行冷冻，在冷冻培养基中使用不同浓度的冷冻保护剂二甲基亚砜和甘油，并改变细胞在冷冻前和解冻后接触冷冻保护剂的时间。通过在混合淋巴细胞培养物中用植物血凝素、商陆有丝分裂原、刀豆球蛋白A和同种异体细胞刺激后对细胞介导的免疫反应进行定量测量来评估细胞活力。发现所研究的因素对免疫反应有重要影响，因此在将冷冻细胞用于 blast 转化试验之前，必须仔细研究并准确确定冷冻系统。最佳冷冻程序持续约40分钟，在发生相变的温度范围内平稳进行。该程序中二甲基亚砜的最佳浓度为8-10%。二甲基亚砜对细胞没有毒性作用，冷冻前无需平衡期。用10%甘油平衡15分钟甚至比二甲基亚砜的最佳程序更好。
