Körber C, Englich S, Schwindke P, Scheiwe M W, Rau G, Hubel A, Cravalho E G
J Microsc. 1986 Mar;141(Pt 3):263-76. doi: 10.1111/j.1365-2818.1986.tb02721.x.
The freezing of biological cell suspensions can be understood in terms of ice formation in the external suspension medium and the cellular reactions to the changing environment. Cryomicroscopy allows a quantitative analysis of both categories of phenomena. Besides freezing stages of appropriate thermal design, the components used for that purpose include a microcomputer (PSI 80) based control system, an image analysis system (Intellect 100) and a spectrophotometer (MPV compact). The investigation of extracellular ice formation is focused on the following effects: The redistribution of solutes in the residual liquid and the resulting concentration profiles are determined photometrically or densitometrically. The transitions between various morphologies of the ice-liquid phase boundary (planar-cellular-dendritic) can be related to interface instability theories. With respect to solute segregation, the studies also involve the formation of bubbles from supersaturated gaseous solutes and freezing potentials resulting from the differential incorporation of cations and anions into the solid phase. The interaction between particles or cells and the advancing ice front is determined from critical interface velocities marking the transition between repulsion and entrapment. The effects of freezing on biological cells are studied mainly with blood cells, especially lymphocytes. The water efflux due to osmotical gradients across the membrane yields volume shrinkage curves which are recorded and analysed from video images for various cooling rates. Beyond a certain threshold cooling rate, intracellular ice starts to form, and different crystallization morphologies can be detected. The intracellular crystallization temperatures depend on cooling and warming rates as well as on the presence of penetrating cryoadditives. A fluorescence viability is used to determine the percentage of damaged cells immediately after thawing.
生物细胞悬液的冷冻可从外部悬浮介质中的冰形成以及细胞对不断变化的环境的反应来理解。低温显微镜术能够对这两类现象进行定量分析。除了具有适当热设计的冷冻阶段外，用于此目的的组件包括基于微型计算机（PSI 80）的控制系统、图像分析系统（Intellect 100）和分光光度计（MPV compact）。对细胞外冰形成的研究集中在以下影响方面：通过光度法或密度测定法确定残余液体中溶质的重新分布以及由此产生的浓度分布。冰 - 液相界面的各种形态（平面 - 胞状 - 树枝状）之间的转变可与界面不稳定性理论相关联。关于溶质偏析，研究还涉及过饱和气态溶质形成气泡以及阳离子和阴离子向固相的差异掺入所产生的冷冻电位。颗粒或细胞与前进的冰前沿之间的相互作用由标志着排斥和截留之间转变的临界界面速度来确定。冷冻对生物细胞的影响主要通过血细胞，特别是淋巴细胞进行研究。由于跨膜渗透梯度导致的水流出产生体积收缩曲线，这些曲线通过视频图像记录并针对各种冷却速率进行分析超过一定的阈值冷却速率后，细胞内开始形成冰，并且可以检测到不同的结晶形态。细胞内结晶温度取决于冷却和升温速率以及穿透性冷冻添加剂的存在。解冻后立即使用荧光活力来确定受损细胞的百分比。
