公开(公告)号：US09163216B1

公开(公告)日：2015-10-20

申请号：US12765399

申请日：2010-04-22

申请人： James J. Hickman ,  Mainak Das ,  John W. Rumsey

发明人： James J. Hickman ,  Mainak Das ,  John W. Rumsey

IPC分类号： C12N5/077 ,  C12N5/0793

CPC分类号： C12N5/0652 ,  C12N5/0619 ,  C12N5/0658 ,  C12N2500/05 ,  C12N2500/46 ,  C12N2500/90 ,  C12N2500/99 ,  C12N2501/40 ,  C12N2502/1335 ,  C12N2533/30

摘要： The invention provides a nutrient medium composition and associated methods for lengthening the useful life of a culture of muscle cells. Disclosed is a method of culturing mammalian muscle cells, including preparing one or more carriers coated with a covalently bonded monolayer of trimethoxy-silylpropyl-diethylenetriamine (DETA); verifying DETA monolayer formation by one or more associated optical parameters; suspending isolated fetal rat skeletal muscle cells in serum-free medium according to medium composition 1; plating the suspended cells onto the prepared carriers at a predetermined density; leaving the carriers undisturbed for cells to adhere to the DETA monolayer; covering the carriers with a mixture of medium 1 and medium 2; and incubating. A cell nutrient medium composition includes Neurobasal, an antibiotic-antimycotic composition, cholesterol, human TNF-alpha, PDGF BB, vasoactive intestinal peptides, insulin-like growth factor 1, NAP, r-Apolipoprotein E2, purified mouse Laminin, beta amyloid, human tenascin-C protein, rr-Sonic hedgehog Shh N-terminal, and rr-Agrin C terminal.

摘要翻译： 本发明提供了一种营养培养基组合物和相关方法，用于延长肌肉细胞培养物的使用寿命。 公开了一种培养哺乳动物肌肉细胞的方法，包括制备一种或多种涂覆有共价键合的三甲氧基 - 甲硅烷基丙基 - 二亚乙基三胺（DETA）单层的载体; 通过一个或多个相关联的光学参数验证DETA单层形成; 根据培养基组成1，在无血清培养基中悬浮分离的胎鼠大鼠骨骼肌细胞; 以预定的密度将悬浮细胞电镀到制备的载体上; 使载体不受干扰以使细胞粘附到DETA单层上; 用介质1和介质2的混合物覆盖载体; 并孵化。 细胞营养培养基组合物包括Neurobasal，抗生素 - 抗真菌组合物，胆固醇，人TNF-α，PDGF BB，血管活性肠肽，胰岛素样生长因子1，NAP，r-载脂蛋白E2，纯化的小鼠层粘连蛋白，β淀粉样蛋白，人 腱生蛋白-C蛋白，rr-Sonic刺猬Shh N末端和rr-Agrin C端。

公开(公告)号：US08815584B1

公开(公告)日：2014-08-26

申请号：US12765996

申请日：2010-04-23

申请人： James J. Hickman ,  Mainak Das

发明人： James J. Hickman ,  Mainak Das

IPC分类号： C12N5/07 ,  C12N11/00 ,  C12N11/16

CPC分类号： C12N5/0619 ,  C12N5/0658 ,  C12N2500/90 ,  C12N2502/088 ,  C12N2502/1335

摘要： The invention provides a method of co-culturing mammalian muscle cells and mammalian motoneurons. The method comprises preparing one or more carriers coated with a covalently bonded monolayer of trimethoxysilylpropyl diethylenetriamine (DETA); suspending isolated fetal mammalian skeletal muscle cells in serum-free medium according to medium composition 1; suspending isolated fetal mammalian spinal motoneurons in serum-free medium according to medium composition 1; plating the suspended muscle cells onto the one or more carriers at a predetermined density and allowing the muscle cells to attach; plating the suspended motoneurons at a predetermined density onto the one or more carriers and allowing the motoneurons to attach; covering the one or more carriers with a mixture of medium composition 1 and medium composition 2; and incubating the carriers covered in the media mixture.

摘要翻译： 本发明提供了共培养哺乳动物肌肉细胞和哺乳动物运动神经元的方法。 该方法包括制备涂覆有共价键合的三甲氧基甲硅烷基丙基二亚乙基三胺（DETA）单层的一种或多种载体; 根据培养基组成1将分离的胎儿哺乳动物骨骼肌细胞悬浮于无血清培养基中; 根据培养基组成1，在无血清培养基中悬浮胎儿哺乳动物脊髓运动神经元; 以预定的密度将悬浮的肌肉细胞电镀到一个或多个载体上并允许肌肉细胞附着; 将预定密度的悬浮运动神经元电镀到一个或多个载体上并允许运动神经元附着; 用培养基组合物1和培养基组合物2的混合物覆盖一种或多种载体; 并培养介质混合物中覆盖的载体。