CN108220232A - 一种基于胎牛血清的培养基的培养方法 - Google Patents

一种基于胎牛血清的培养基的培养方法 Download PDF

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CN108220232A
CN108220232A CN201810233420.2A CN201810233420A CN108220232A CN 108220232 A CN108220232 A CN 108220232A CN 201810233420 A CN201810233420 A CN 201810233420A CN 108220232 A CN108220232 A CN 108220232A
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王翔
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Tianjin Booko Biotechnology Co Ltd
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Abstract

本发明公开了一种基于胎牛血清的培养基的培养方法，包括以下步骤：1)将下列成分及含量依次加入器皿中：胎牛血清：20ul/ml；重组人胰岛素：5ug/ml；碱性成纤维细胞生长因子：4ng/ml；人转铁蛋白：5ug/ml；亚硒酸：6.84ug/ml；丝氨酸：105ug/ml；氯化胆碱：89.3ug/ml；腺嘌呤：12.2ug/ml；单乙酰胺：14ug/ml；磷酰乙醇胺：0.1mmol/ml；三碘甲状腺氨酸：13.46pg/ml，每加入一种成分搅拌2～3分钟；2)采用吸管吸取步骤1)中的液体3mL～5mL到15mLd离心管中，离心管以200g/min的转速离心3min～5min，弃上清后置于细胞培养箱中培养2小时；3)吸取步骤2)中培养2小时后的细胞悬液，按体积比1:20～1:25接种量接种于DMEM培养液中，培养3天～4天后即得。本发明使间充质干细胞在细胞形态和增殖能力方面都得到了加强。

Description

一种基于胎牛血清的培养基的培养方法
技术领域
本发明属于血清培养液技术领域，特别涉及一种基于胎牛血清的培养基的培养方法，用于间充质干细胞。
背景技术
间充质干细胞【mesenchymal stem cells,MSC】是干细胞家族的重要成员，来源于发育早期的中胚层和外胚层，属于多能干细胞，MSC最初在骨髓中发现，因其具有多向分化潜能、造血支持和促进干细胞植入、免疫调控和自我复制等特点而日益受到人们的关注。如间充质干细胞在体内或体外特定的诱导条件下，可分化为脂肪、骨、软骨、肌肉、肌腱、韧带、神经、肝、心肌、内皮等多种组织细胞，连续传代培养和冷冻保存后仍具有多向分化潜能，可作为理想的种子细胞用于衰老和病变引起的组织器官损伤修复。
细胞培养在干细胞冻存中广泛应用，目前用于培养间充质干细胞的培养基存在有影响细胞存活率的问题，目前尚未得到很好地解决。
发明内容
针对现有技术中存在的技术问题，本发明提供一种基于胎牛血清的培养基的培养方法。
为解决上述技术问题，本发明采用的技术方案是：一种基于胎牛血清的培养基的培养方法，包括以下步骤：
1)将下列成分及含量依次加入器皿中：胎牛血清：20ul/ml；重组人胰岛素：5ug/ml；碱性成纤维细胞生长因子：4ng/ml；人转铁蛋白：5ug/ml；亚硒酸：6.84ug/ml；丝氨酸：105ug/ml；氯化胆碱：89.3ug/ml；腺嘌呤：12.2ug/ml；单乙酰胺：14ug/ml；磷酰乙醇胺：0.1mmol/ml；三碘甲状腺氨酸：13.46pg/ml，每加入一种成分搅拌2～3分钟；
2)采用吸管吸取步骤1)中的液体3mL～5mL到15mLd离心管中，所述离心管以200g/min的转速离心3min～5min，弃上清后置于细胞培养箱中培养2小时；
3)吸取步骤2)中培养2小时后的细胞悬液，按体积比1:20～1:25接种量接种于DMEM培养液中，培养3天～4天后，并定期观察，即得。
作为优选，所述DMEM为高糖DMEM。
与现有技术相比，本发明所具有的有益效果是：本发明在通用型DMEM培养基的基础上，添加包括胎牛血清在内的多种促进细胞生长的物质，使间充质干细胞在细胞形态和增殖能力方面都得到了加强。
具体实施方式
为使本领域技术人员更好的理解本发明的技术方案，下面结合具体实施例对本发明作详细说明。
一种基于胎牛血清的培养基的培养方法，包括以下步骤：
1)将下列成分及含量依次加入器皿中：胎牛血清：20ul/ml；重组人胰岛素：5ug/ml；碱性成纤维细胞生长因子：4ng/ml；人转铁蛋白：5ug/ml；亚硒酸：6.84ug/ml；丝氨酸：105ug/ml；氯化胆碱：89.3ug/ml；腺嘌呤：12.2ug/ml；单乙酰胺：14ug/ml；磷酰乙醇胺：0.1mmol/ml；三碘甲状腺氨酸：13.46pg/ml，每加入一种成分搅拌2～3分钟；
2)采用吸管吸取步骤1)中的液体3mL～5mL到15mLd离心管中，所述离心管以200g/min的转速离心3min～5min，弃上清后置于细胞培养箱中培养2小时；
3)吸取步骤2)中培养2小时后的细胞悬液，按体积比1:20～1:25接种量接种于DMEM培养液中，培养3天～4天后，并定期观察，即得。
本实施例中，DMEM为高糖DMEM。
以上实施例仅为本发明的示例性实施例，不用于限制本发明，本发明的保护范围由权利要求书限定。本领域技术人员可以在本发明的实质和保护范围内，对本发明做出各种修改或等同替换，这种修改或等同替换也应视为落在本发明的保护范围内。

Claims (2)

1.一种基于胎牛血清的培养基的培养方法，用于培养人间充质干细胞，其特征在于，包括以下步骤：
1)将下列成分及含量依次加入器皿中：胎牛血清：20ul/ml；重组人胰岛素：5ug/ml；碱性成纤维细胞生长因子：4ng/ml；人转铁蛋白：5ug/ml；亚硒酸：6.84ug/ml；丝氨酸：105ug/ml；氯化胆碱：89.3ug/ml；腺嘌呤：12.2ug/ml；单乙酰胺：14ug/ml；磷酰乙醇胺：0.1mmol/ml；三碘甲状腺氨酸：13.46pg/ml，每加入一种成分搅拌2～3分钟；
2)采用吸管吸取步骤1)中的液体3mL～5mL到15mLd离心管中，所述离心管以200g/min的转速离心3min～5min，弃上清后置于细胞培养箱中培养2小时；
3)吸取步骤2)中培养2小时后的细胞悬液，按体积比1:20～1:25接种量接种于DMEM培养液中，培养3天～4天后，并定期观察，即得。
2.根据权利要求1所述的一种基于胎牛血清的培养基的培养方法，其特征在于，所述DMEM为高糖DMEM。
CN201810233420.2A 2018-03-21 2018-03-21 一种基于胎牛血清的培养基的培养方法 Pending CN108220232A (zh)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116004521A (zh) * 2023-01-06 2023-04-25 超技良食(深圳)生物科技有限公司 一种用于鸡肌源性干细胞体外增殖的培养基及制备方法和应用

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN116004521A (zh) * 2023-01-06 2023-04-25 超技良食(深圳)生物科技有限公司 一种用于鸡肌源性干细胞体外增殖的培养基及制备方法和应用
CN116004521B (zh) * 2023-01-06 2023-11-03 汕头得宝投资有限公司 一种用于鸡肌源性干细胞体外增殖的培养基及制备方法和应用

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