Ehrhart F, Schulz J C, Katsen-Globa A, Shirley S G, Reuter D, Bach F, Zimmermann U, Zimmermann H
Fraunhofer Institut für Biomedizinische Technik, Ensheimer Strasse 48, 66386 St. Ingbert, Germany.
Cryobiology. 2009 Apr;58(2):119-27. doi: 10.1016/j.cryobiol.2008.11.005. Epub 2008 Dec 3.
Cryopreservation of human tumour cells and tissue is a valuable tool for retrospective analysis and for the transport and handling of biopsy material. Tumour tissue consists of different cell types, which have different optimal freezing conditions, and extracellular matrix. A well-defined and authentic model system is required for developing new freezing protocols and media. This work describes the use of L929 and PC-3 spheroids as new model systems for freezing human tumours. Cell suspension and spheroids were frozen in different vessels (1 ml cryovials and a special, cryo-compatible 30 x 25 microl multi well plate) at slow rate (1 degrees C/min). Freezing media were combinations of culture or tumour transport medium (Liforlab) with the cryoprotective agents, Me(2)SO, trehalose and modified starch. We also present a new method of evaluating the viability of three dimensional multicellular systems to compare thawed spheroids objectively. Best viability (70%) of L929 spheroids occurred with a combination of Liforlab and starch hydrolysis product. The best cryopreservation results for spheroids were found with extracellular cryoprotectants, while optimum viability of single cells was achieved with Me(2)SO.
人类肿瘤细胞和组织的冷冻保存是回顾性分析以及活检材料运输和处理的一项重要工具。肿瘤组织由具有不同最佳冷冻条件的不同细胞类型以及细胞外基质组成。开发新的冷冻方案和培养基需要一个定义明确且真实的模型系统。这项工作描述了使用L929和PC - 3球体作为冷冻人类肿瘤的新模型系统。细胞悬液和球体在不同容器（1毫升冻存管和一种特殊的、与冷冻兼容的30×25微升多孔板）中以缓慢速率（1℃/分钟）冷冻。冷冻培养基是培养或肿瘤运输培养基（Liforlab）与冷冻保护剂二甲基亚砜（Me(2)SO）、海藻糖和变性淀粉的组合。我们还提出了一种评估三维多细胞系统活力的新方法，以便客观地比较解冻后的球体。L929球体在Liforlab和淀粉水解产物组合下具有最佳活力（70%）。对于球体，细胞外冷冻保护剂可获得最佳冷冻保存结果，而单细胞的最佳活力则通过二甲基亚砜实现。
