McColm A A, Latter V S
Trans R Soc Trop Med Hyg. 1986;80(1):29-33. doi: 10.1016/0035-9203(86)90188-4.
Several cryoprotectants were compared using two different cooling procedures for the cryopreservation of both normal and Plasmodium yoelii-infected mouse erythrocytes. Fast cooling to -196 degrees C by direct plunge into liquid nitrogen followed by rapid thawing in a 37 degree C water bath protected uninfected and parasitized erythrocytes against freeze-thaw damage significantly better than an interrupted slow cooling procedure in which cells were frozen to -70 degrees C at a rate of -1 degree C/min followed by storage in liquid nitrogen. Using the former procedure, the highest percentage erythrocyte recoveries (over 85%) and shortest pre-2% parasitaemia times (equivalent to unfrozen cells) in mice challenged with thawed parasitized blood were observed with the intracellular (penetrating) cryoprotectants glycerol or dimethylsulphoxide (Me2SO). At the concentrations used in this study, the extracellular (non-penetrating) cryoprotectants, hydroxyethylstarch, dextran or polyvinyl pyrrolidone were significantly less effective at protecting against freeze-thaw damage. Some evidence of freeze-thaw damage was obtained in cells fast frozen at low cell density (less than or equal to 10(4)/ml) in glycerol or Me2SO. This effect was masked when higher cell densities (10(7) to 10(8)/ml) were used. Addition of the anti-oxidant and membrane stabilizing molecules, vitamin E, sodium selenite or selenomethionine to infected erythrocytes just before and during cryopreservation, did not improve, and in one case inhibited, subsequent development in challenged mice.
使用两种不同的冷却程序对正常和感染约氏疟原虫的小鼠红细胞进行冷冻保存，比较了几种冷冻保护剂。通过直接投入液氮快速冷却至-196℃，然后在37℃水浴中快速解冻，对未感染和被寄生的红细胞的冻融损伤保护效果明显优于间断缓慢冷却程序，后者是将细胞以-1℃/分钟的速率冷冻至-70℃，然后储存在液氮中。使用前一种程序，在用解冻的被寄生血液攻击的小鼠中，观察到细胞内（穿透性）冷冻保护剂甘油或二甲基亚砜（Me2SO）的红细胞回收率最高（超过85%），2%寄生虫血症前时间最短（相当于未冷冻细胞）。在本研究中使用的浓度下，细胞外（非穿透性）冷冻保护剂羟乙基淀粉、右旋糖酐或聚乙烯吡咯烷酮在防止冻融损伤方面效果明显较差。在甘油或Me2SO中以低细胞密度（小于或等于10⁴/ml）快速冷冻的细胞中获得了一些冻融损伤的证据。当使用更高的细胞密度（10⁷至10⁸/ml）时，这种效应被掩盖。在冷冻保存前和保存期间向感染的红细胞中添加抗氧化剂和膜稳定分子维生素E、亚硒酸钠或硒代蛋氨酸，并没有改善，在一种情况下还抑制了攻击小鼠后的后续发育。
