大鼠胚胎大脑组织的冷冻保存研究

Cryopreservation of cerebral tissue from 18-day fetal rats

用组织培养测定活性的方法研究了胎龄18d大鼠胚胎大脑组织的适宜冻存条件,较系统地观察了有关低温保护剂、冷冻和复温速率、保存时间等条件对胎鼠脑组织活性的影响。结果表明,以下条件能使冻存组织获得良好的培养结果:以1mol/L的二甲亚砜作为低温保护剂,以1℃/min的速率冷冻至-70℃,液氮保存,37℃水浴快速复温。在液氮中保存60d活性无明显变化。2mol/L甘油对胎脑组织也有一定的保护作用,但不如1mol/L二甲亚砜,而聚乙烯吡咯烷酮无明显保护作用。冷冻速率等于或超过5℃/min,则组织活性急剧下降,说明冷冻速率对胚胎大脑组织活性影响较大。

The optimal procedure for the cryopreservation of cerebral tissue from 18-dayfetal rats has been selected using tissue culture as a viability assay. We investigated the effects ofvarious cryopreserved conditions including cryoprotectants, freezing and thawing rates and stor-age time in liquid nitrogen on the tissue viability. The finding showed that the following conditionsyielded the best results: using DMEM with 20% horse serum and 1mol / L DMSO as the freezingmedium, freezing at a rate of 1℃ /min, storing in liquid nitrogen and rapidly thawing in 37℃water bath. We also found that 2mol / L glycerol had some protective effect although it was infe-rior to 1mol / L DMSO while PVP had no obvious effect. The cooling rates exceeding 5℃ / minresulted in sharp decrease of the viability of tissue which indicated that the embryonic cerebral tis-sue was very sensitive to cryoinjury. The tissue freeze-thawed in the optimal procedure showed onobvious viability loss during storage in liquid nitrogen for 1 to 60 days.