CN106190946A - 一种用于干细胞扩增的培养基 - Google Patents

一种用于干细胞扩增的培养基 Download PDF

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CN106190946A
CN106190946A CN201610570585.XA CN201610570585A CN106190946A CN 106190946 A CN106190946 A CN 106190946A CN 201610570585 A CN201610570585 A CN 201610570585A CN 106190946 A CN106190946 A CN 106190946A
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张正亮
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Anhui From Biological Technology Co Ltd
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Abstract

本发明公开一种用于干细胞扩增的培养基，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：营养因子：4‑6g/L转铁蛋白、0.1‑0.15g/L亚硒酸钠、10‑18g/L牛血清白蛋白、0.1‑0.2g/L谷氨酰胺、3‑4g/L糖蛋白、1‑2g/L载脂蛋白；细胞因子：2‑4g/L干细胞因子、3‑4g/L表皮生长因子、1‑2g/L成纤维细胞生长因子、1‑2g/L神经生长因子、2‑3g/L血小板衍化生长因子、4‑6g/L血小板因子；活性因子：8‑12g/L人胰岛素、0.5‑0.8g/L四甲烯二胺、0.2‑0.5g/L黄体激素、1‑2g/L维生素、0.5‑1g/L碳酸氢钠、2‑4g/L牛血清白蛋白、0.2‑0.4g/L抗生素、2‑4g/L寡肽‑1、2‑4g/L白介素6、1‑2g/L生物素。本发明提供的培养基能够快速扩增干细胞同时又不影响干细胞的潜能，提高了干细胞扩增速度，缩短了干细胞培养的时间，具有极佳的适用性。

Description

一种用于干细胞扩增的培养基
技术领域
本发明属于生物技术领域，涉及一种培养基，具体是一种用于干细胞扩增的培养基。
背景技术
干细胞是一类具有自我更新及多向分化潜能的细胞群体，即这些细胞可以通过细胞分裂维持自身细胞群的大小，同时又可以进一步分化成为各种不同的组织细胞，从而医学界称之为“万能细胞”。这种独一无二的特性使其成为再生医学和移植医学的一个不可缺少的细胞来源。也使它成为众多疾病，如血液疾病、帕金森病，糖尿病，脊髓损伤、癌症和心脏病等细胞治疗的最佳候选者，因此对于干细胞的研究在临床应用领域意义重大。
近年来，使用干细胞的技术被认为是治疗不可治愈疾病的新途径。干细胞是一种能够不断自我更新的细胞，并能增生、分裂、分化成其他各种成熟的细胞。干细胞可按不同的标准进行分类。按它们的来源可分为胚胎干细胞，骨髓干细胞和脐带血干细胞；按它们的增生和分化的能力可分为全潜能、多潜能和单潜能的干细胞；按它们的所能够分化成的细胞可分为神经干细胞、心肌干细胞、血液干细胞等；按它们的成熟的程度可分为胚胎干细胞和成熟组织干细胞。
诸多研究表明干细胞不仅可以自我更新，在条件合适的情况下能分化成其他功能细胞，因此干细胞有望成为治疗人类疑难疾病的有效手段。然而，干细胞在正常成体组织中含量甚微，在体外如何快速扩增与培养干细胞是研究干细胞的作用机制及探索其在人类疾病治疗方法的重要技术。
发明内容
本发明的目的在于提供一种用于干细胞扩增的培养基。
本发明的目的可以通过以下技术方案实现：
一种用于干细胞扩增的培养基，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：
营养因子：4-6g/L转铁蛋白、0.1-0.15g/L亚硒酸钠、10-18g/L牛血清白蛋白、0.1-0.2g/L谷氨酰胺、3-4g/L糖蛋白、1-2g/L载脂蛋白；
细胞因子：2-4g/L干细胞因子、3-4g/L表皮生长因子、1-2g/L成纤维细胞生长因子、1-2g/L神经生长因子、2-3g/L血小板衍化生长因子、4-6g/L血小板因子；
活性因子：8-12g/L人胰岛素、0.5-0.8g/L四甲烯二胺、0.2-0.5g/L黄体激素、1-2g/L维生素、0.5-1g/L碳酸氢钠、2-4g/L牛血清白蛋白、0.2-0.4g/L抗生素、2-4g/L寡肽-1、2-4g/L白介素6、1-2g/L生物素。
本发明的有益效果：
本发明提供的培养基能够快速扩增干细胞同时又不影响干细胞的潜能，干细胞的扩增速度较常规培养基提高3-5倍，而且能用于培养多种组织的干细胞，具有极佳的适用性，所培养的干细胞分化能力强，可分化成多种功能细胞，具有很高的科研和医学应用价值；
本发明培养基培养的干细胞保持了原有生物学特性，并且维持了干细胞的多向分化潜能；
本发明培养基成分确定，因此重现性和可控性都要比血清培养基好，且培养的干细胞安全，可用于干细胞临床治疗，其生产比血清培养基更适合规模化；
本发明基于对临床上用途较广的干细胞进行研究，能快速扩增干细胞而又 不影响其分化潜能的细胞因子，提高了干细胞扩增速度，缩短了干细胞培养的时间。
具体实施方式
下面将结合本发明实施例对本发明的技术方案进行清楚、完整地描述，显然，所描述的实施例仅仅是本发明一部分实施例，而不是全部的实施例。基于本发明中的实施例，本领域普通技术人员在没有作出创造性劳动前提下所获得的所有其它实施例，都属于本发明保护的范围。
实施例1
一种用于干细胞扩增的培养基，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：
营养因子：5g/L转铁蛋白、0.12g/L浓度为25％的亚硒酸钠、15g/L牛血清白蛋白、0.15g/L谷氨酰胺、3.5g/L糖蛋白、1.5g/L载脂蛋白；
细胞因子：3g/L干细胞因子、3.5g/L表皮生长因子、1.5g/L成纤维细胞生长因子、1.5g/L神经生长因子、2.5g/L血小板衍化生长因子、5g/L血小板因子；
活性因子：10g/L人胰岛素、0.6g/L四甲烯二胺、0.4g/L黄体激素、1.5g/L维生素、0.8g/L浓度为55％的碳酸氢钠、3g/L牛血清白蛋白、0.3g/L抗生素、3g/L寡肽-1、3g/L白介素6、1.5g/L生物素。
将营养因子、细胞因子、活性因子分别混合后，再依次混合，最后加入到基础培养基DMEM/F12中，调节ph至中性。
实施例2
一种用于干细胞扩增的培养基，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：
营养因子：6g/L转铁蛋白、0.1g/L浓度为28％的亚硒酸钠、18g/L牛血清白 蛋白、0.1g/L谷氨酰胺、4g/L糖蛋白、1g/L载脂蛋白；
细胞因子：4g/L干细胞因子、3g/L表皮生长因子、2g/L成纤维细胞生长因子、1g/L神经生长因子、3g/L血小板衍化生长因子、4g/L血小板因子；
活性因子：12g/L人胰岛素、0.5g/L四甲烯二胺、0.5g/L黄体激素、1g/L维生素、1g/L浓度为59％的碳酸氢钠、2g/L牛血清白蛋白、0.4g/L抗生素、2g/L寡肽-1、4g/L白介素6、1g/L生物素。
实施例3
一种用于干细胞扩增的培养基，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：
营养因子：4g/L转铁蛋白、0.15g/L浓度为22％的亚硒酸钠、10g/L牛血清白蛋白、0.2g/L谷氨酰胺、3g/L糖蛋白、2g/L载脂蛋白；
细胞因子：2g/L干细胞因子、4g/L表皮生长因子、1g/L成纤维细胞生长因子、2g/L神经生长因子、2g/L血小板衍化生长因子、6g/L血小板因子；
活性因子：8g/L人胰岛素、0.8g/L四甲烯二胺、0.2g/L黄体激素、2g/L维生素、0.5g/L浓度为56％的碳酸氢钠、4g/L牛血清白蛋白、0.2g/L抗生素、4g/L寡肽-1、2g/L白介素6、2g/L生物素。
本发明添加糖蛋白是为了促进干细胞或细胞表面蛋白的分子识别，本发明培养基培养的干细胞通过细胞迁移及生物识别，相同的干细胞在一定部位相互交联，聚集成片，实现集落式增殖；载脂蛋白可刺激干细胞贴壁生长，并且载脂蛋白本身具有促细胞分裂作用；维生素参与细胞的正常发育，并维持机体多种生理活动，并且在体外干细胞的培养中对干细胞增殖和分化有重要影响；人胰岛素对干细胞中糖代谢、脂肪代谢和蛋白代谢起着关键调节作用，对干细胞的增殖和存活起到促进作用。
本发明提供的干细胞扩增培养基，能够使干细胞保持原有的生物学特性，且维持了多向分化的潜能；本发明培养基组成是确定的，因此重现性、可控性、且培养的干细胞是安全的；培养基细胞增殖速率较快，得到的细胞数量较多，且代次间较为稳定，提高了干细胞扩增速度，大大缩短了干细胞培养的时间。
在本说明书的描述中，参考术语“一个实施例”、“示例”、“具体示例”等的描述意指结合该实施例或示例描述的具体特征、结构、材料或者特点包含于本发明的至少一个实施例或示例中。在本说明书中，对上述术语的示意性表述不一定指的是相同的实施例或示例。而且，描述的具体特征、结构、材料或者特点可以在任何的一个或多个实施例或示例中以合适的方式结合。
以上公开的本发明优选实施例只是用于帮助阐述本发明。优选实施例并没有详尽叙述所有的细节，也不限制该发明仅为所述的具体实施方式。显然，根据本说明书的内容，可作很多的修改和变化。本说明书选取并具体描述这些实施例，是为了更好地解释本发明的原理和实际应用，从而使所属技术领域技术人员能很好地理解和利用本发明。本发明仅受权利要求书及其全部范围和等效物的限制。

Claims (1)

1.一种用于干细胞扩增的培养基，其特征在于，该培养基由1L基础培养基DMEM/F12、以及下述因子制成：
营养因子：4-6g/L转铁蛋白、0.1-0.15g/L亚硒酸钠、10-18g/L牛血清白蛋白、0.1-0.2g/L谷氨酰胺、3-4g/L糖蛋白、1-2g/L载脂蛋白；
细胞因子：2-4g/L干细胞因子、3-4g/L表皮生长因子、1-2g/L成纤维细胞生长因子、1-2g/L神经生长因子、2-3g/L血小板衍化生长因子、4-6g/L血小板因子；
活性因子：8-12g/L人胰岛素、0.5-0.8g/L四甲烯二胺、0.2-0.5g/L黄体激素、1-2g/L维生素、0.5-1g/L碳酸氢钠、2-4g/L牛血清白蛋白、0.2-0.4g/L抗生素、2-4g/L寡肽-1、2-4g/L白介素6、1-2g/L生物素。
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