Gorfien S, Paul B, Walowitz J, Keem R, Biddle W, Jayme D
Life Technologies, Inc., 3175 Staley Road, Grand Island, New York, NY 14072, USA.
Biotechnol Prog. 2000 Sep-Oct;16(5):682-7. doi: 10.1021/bp000109a.
Many hybridoma and recombinant myeloma cell lines have been successfully adapted to growth in protein-free media. Compared with serum-supplemented media, use of protein-free media promotes superior cell growth and protein expression and facilitates downstream purification of the expressed product. Owing to its sterol auxotrophy, the NS0 myeloma is normally grown in either a serum-supplemented medium or a serum-free medium supplemented with an animal-derived lipoprotein. CD Hybridoma Medium (a protein-free, chemically defined formulation) grows many cell lines that do not exhibit lipid dependence, but this medium does not support growth of NS0 cells without further lipid supplementation. We tested several commercially available lipid supplements in CD Hybridoma Medium, including bovine EX-CYTE VLE. None of the tested supplements supported long-term growth of NS0 cells in CD Hybridoma Medium. Sustained long-term growth of NS0 cells was achieved in CD Hybridoma Medium supplemented with various animal- or plant-derived lipids complexed with cyclodextrin. NS0 cells adapted to CD Hybridoma Medium supplemented with cyclodextrin-lipid complex reached peak cell densities that were more than double those observed in serum-supplemented medium and were cultured for more than 15 passages. These cultures were also successfully cryopreserved and recovered in this defined medium. Through the use of cyclodextrin-based additives to CD Hybridoma Medium, it is possible to solubilize significant quantities of sterols and other lipids and to maintain a protein-free, chemically defined cultivation environment for NS0 cells. The culture system can be kept entirely free of animal-derived components if the supplement is made with plant-derived or synthetic lipids.
许多杂交瘤细胞系和重组骨髓瘤细胞系已成功适应无蛋白培养基中的生长。与补充血清的培养基相比，使用无蛋白培养基可促进细胞更好地生长和蛋白质表达，并便于对表达产物进行下游纯化。由于NS0骨髓瘤细胞存在固醇营养缺陷，通常在补充血清的培养基或添加动物源脂蛋白的无血清培养基中培养。CD杂交瘤培养基（一种无蛋白、化学成分明确的配方）可培养许多不依赖脂质的细胞系，但该培养基在不进一步添加脂质的情况下无法支持NS0细胞生长。我们在CD杂交瘤培养基中测试了几种市售脂质补充剂，包括牛源EX-CYTE VLE。所测试的补充剂均无法支持NS0细胞在CD杂交瘤培养基中长期生长。在添加了与环糊精复合的各种动物或植物源脂质的CD杂交瘤培养基中，NS0细胞实现了持续的长期生长。适应添加了环糊精-脂质复合物的CD杂交瘤培养基的NS0细胞达到的峰值细胞密度比在补充血清的培养基中观察到的密度高出一倍多，并且传代培养超过15次。这些培养物也成功地在这种限定培养基中冷冻保存和复苏。通过在CD杂交瘤培养基中使用基于环糊精的添加剂，可以溶解大量固醇和其他脂质，并为NS0细胞维持无蛋白、化学成分明确的培养环境。如果补充剂采用植物源或合成脂质制备，则培养系统可以完全不含动物源成分。
