CN114075540A - 全化学成分hek293细胞培养基及其应用 - Google Patents

全化学成分hek293细胞培养基及其应用 Download PDF

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CN114075540A
CN114075540A CN202010852123.3A CN202010852123A CN114075540A CN 114075540 A CN114075540 A CN 114075540A CN 202010852123 A CN202010852123 A CN 202010852123A CN 114075540 A CN114075540 A CN 114075540A
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陈宜顶
孟祥春
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Suzhou Xinweixi Biopharmaceutical Co ltd
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Abstract

本发明提供一种全化学成分HEK293细胞培养基及其应用。所述培养基包括氨基酸、无机盐、维生素、微量元素、碳水化合物和有机分子。本发明的细胞培养基不含任何生长因子及其它蛋白添加物，无非化学成分界定组分；通过引入更多包括微量元素在内的组分，替代一般培养基设计中所必需的生长因子，支持HEK293细胞的高密度和高活率生长。

Description

全化学成分HEK293细胞培养基及其应用
技术领域
本发明涉及细胞培养技术领域，具体地说，涉及一种全化学成分HEK293细胞培养基及其应用。
背景技术
近年来，随着基因工程技术的问世，培养基已被广泛用于多种细胞和微生物培养，包括各种动物细胞、植物细胞、细菌、真菌和病毒等。生物制药工业中的目标产物蛋白可以通过在液体培养基中培养细胞而大量生产。这些细胞为天然的或工程化的细胞，利用发酵罐等设备在液体培养基中快速成倍增长，并生产目标产物。这些产物有的直接从培养基中收集，有的从收获的细胞中提取获得。培养基提供细胞培养过程中所需的营养物质，在早期的细胞培养过程中，培养基配方以血清的化学组成和物理化学性质(如渗透压、pH值等)为基础。但不同类型的细胞在不同的培养环境中对于氧气/二氧化碳分压、营养物的需求有所不同，因此，培养基的配方取决于特定的细胞要求。一般而言，培养基的主要成分包括氨基酸、有机盐、无机盐、维生素、微量元素、碳源和其它有机分子等，各组分的浓度根据给定细胞的特定要求而相应调整。
自1990年开始出现无蛋白培养基PFM(含水解物)和完全化学成分限定培养基。从2000年以来，无动物源完全化学成分培养基持续优化，用以支持细胞高密度培养和高产物表达。全化学成分限定培养基具有批间差异小、可重复性高，容易进行纯化和下游加工，便于细胞功能的精确评估，减少抗生素使用量等优势。
发明内容
本发明的目的是提供一种全化学成分(无血清无蛋白无动物源性)HEK293细胞培养基及其应用。
为了实现本发明目的，第一方面，本发明提供一种全化学成分HEK293细胞培养基，所述培养基包括氨基酸、无机盐、维生素、微量元素、碳水化合物和有机分子；其中，氨基酸的组成及用量如下：
Figure BDA0002645087430000011
Figure BDA0002645087430000021
无机盐的组成及用量如下：
Figure BDA0002645087430000022
微量元素的组成及用量如下：
Figure BDA0002645087430000023
维生素的组成及用量如下：
Figure BDA0002645087430000024
碳水化合物和有机分子的组成及用量如下：
Figure BDA0002645087430000025
Figure BDA0002645087430000031
优选地，所述细胞培养基中：氨基酸的组成及用量如下：
Figure BDA0002645087430000032
无机盐的组成及用量如下：
Figure BDA0002645087430000033
微量元素的组成及用量如下：
Figure BDA0002645087430000034
Figure BDA0002645087430000041
维生素的组成及用量如下：
Figure BDA0002645087430000042
碳水化合物和有机分子的组成及用量如下：
Figure BDA0002645087430000043
进一步地，所述全化学成分细胞培养基的pH值为6.9-7.1。
第二方面，本发明提供所述培养基的配制方法，包括：按比例称取各组分，用水溶解，然后用水定容，并调pH至6.9-7.1。
第三方面，本发明提供所述培养基在哺乳动物细胞，特别是HEK293细胞或重组HEK293细胞体外培养中的应用。
第四方面，本发明提供利用所述全化学成分细胞培养基培养HEK293细胞的方法，包括：在装有全化学成分细胞培养基的生物反应器中悬浮培养HEK293细胞，悬浮培养的条件为：温度36.5-37.0℃，pH 6.9-7.1，溶氧浓度40～45％；接种的起始细胞浓度为0.5×106～0.7×106个细胞/mL。
借由上述技术方案，本发明至少具有下列优点及有益效果：
(一)本发明利用统计学实验设计并确定各组分的最佳用量，无任何生长因子及其它蛋白添加物，无非化学成分界定组分，所有组分化学结构已知；
(二)通过在培养基中引入更多包括微量元素在内的组分，替代一般培养基设计中所必需的生长因子(如胰岛素、转铁蛋白等)，支持细胞(特别是HEK293细胞)高密度和高活率生长；
(三)通过在培养基中引入脂类物质，亚油酸甲酯0.1～10.0mg/L(优选1.5mg/L)，油酸甲酯1.0～20.0mg/L(优选4.2mg/L)，花生四烯酸甲基酯0.1～10.0mg/L(优选1.3mg/L)，亚麻酸甲酯0.1～10.0mg/L(优选2.2mg/L)，棕榈油酸甲酯1.0～20.0mg/L(优选3.6mg/L)，棕榈酸甲酯0.1～20.0mg/L(优选1.0mg/L)，HEK293细胞可产生重要的脂肪酸和脂肪酸衍生物；
(四)由于无任何生长因子及蛋白，所有组分化学结构已知，均为小分子物质，成本低，便于配制、储存、批间差异小，且易于下游纯化和加工。
附图说明
图1为本发明较佳实施例中3组平行全化学成分细胞培养基培养HEK293细胞的生长曲线。
图2为本发明较佳实施例中3L灌流培养体系中使用全化学成分细胞培养基培养HEK293细胞(293F细胞)的密度及活率曲线。其中，A1、A2为本发明培养基灌流时的细胞生长数据，B为采用CN104450607A中的培养基在相同实验条件下测定的数据。
具体实施方式
以下实施例用于说明本发明，但不用来限制本发明的范围。若未特别指明，实施例中所用的技术手段为本领域技术人员所熟知的常规手段，所用原料均为市售商品。
实施例1用于HEK293悬浮培养的全化学成分细胞培养基及培养方法
一、培养基的配制
1、准备配制培养基所需药品/试剂。
2、玻璃瓶的灭菌。
3、滤器的灭菌
囊式滤器：冲洗囊式滤器，正反冲洗各15min，按顺序安装，进出气口螺丝拧松，包好锡箔纸，将滤器包好锡箔纸，管子连接处用扎带扎好，进液管口用锡纸包裹，出液口安装转换瓶，于高压灭菌锅中灭菌。
4、配制过程
洗净1L烧杯，加入约700mL的超纯水。加入如下药品：氨基酸的组成及用量如下：
Figure BDA0002645087430000061
无机盐的组成及用量如下：
Figure BDA0002645087430000062
微量元素的组成及用量如下：
Figure BDA0002645087430000063
维生素的组成及用量如下：
Figure BDA0002645087430000064
Figure BDA0002645087430000071
碳水化合物和有机分子的组成及用量如下：
Figure BDA0002645087430000072
5、混合定容
将溶解后的试剂用超纯水定容，并将最终混合液的pH调至6.9-7.1之间，得到全化学成分HEK293细胞培养基。
6、无菌过滤
将灭菌好已贴上标签的1L蓝盖瓶喷洒酒精后放入超净工作台中，然后将囊式滤器的转换瓶瓶盖和1L蓝盖瓶瓶盖进行无菌对换，打开蠕动泵，将转速调节到300r/min，待培养基达到1L蓝盖瓶刻度线时关闭蠕动泵，过滤结束。放入冰箱2～8℃避光储存。
二、HEK293细胞的悬浮培养
1、试剂及设备
125mL三角摇瓶、超净工作台、37℃恒温摇床、全自动细胞计数仪、HEK293细胞、待检测培养基、台式离心机。
2、实验步骤
2.1准备实验细胞
复苏并活化HEK293细胞，待细胞密度达到2.0×106cells/mL～2.5×106cells/mL，活率≥96％，按密度0.5×106cells/mL离心换液(800rpm离心5min)，换为步骤一配制的全化学成分细胞培养基，将培养基装入250mL玻璃三角摇瓶中。装液量为50mL，恒温摇床设置37℃，135rpm转速培养。
2.2记录细胞生长数据
培养期间不添加任何试剂，每隔24h取样，用台盼蓝染色细胞并记录细胞生长密度和活率。绘制细胞生长曲线和活率变化曲线。如图1所示，3组平行实验(A1、A2、A3)在不补充任何试剂且所用摇床为非二氧化碳供给的普通摇床中所养细胞密度峰值均在9×106cells/mL以上，增长到峰值以前细胞活率均在90％以上。
三、培养HEK293细胞在灌流培养体系中的生长情况
全化学成分细胞培养基可用于高密度的细胞生长及活力(图2)，为了检验存活细胞的数量，将培养基装入3L的灌流培养反应器内，装液量为2L，接种HEK293，接种的起始细胞浓度为0.5×106cells/mL，悬浮培养条件为：温度36.5-37.0℃，pH 6.9-7.1，溶氧浓度40～45％。连续培养522h，期间监控活性细胞的细胞密度。活性细胞计数通过标准的台盼蓝染色分析并计数，氧气和二氧化碳通过管道连续供应到反应器中。
在相同条件下进一步测试了采用CN104450607A说明书实施例1公开的方法对HEK293细胞进行悬浮培养，结果显示最高细胞密度为19.1×106cells/mL(图2)，在培养长达500h内，细胞活率都在85％以下。从图2可以看出，在相同培养条件下，采用本发明改良的全化学成分细胞培养基培养HEK293，细胞倍增时间更短，细胞密度最高比CN104450607A方法高出50％左右，整个培养期间细胞活率明显更高。
通过以上比较可知，采用本发明提供的全化学成分HEK293细胞培养基，可以在10天左右使细胞由接种密度0.5×106cells/mL增加到25×106～30×106cells/mL，并在较长时间内(约10天)维持高密度高活率生长，细胞快速增长，在高密度维持较长时间可收获更多的蛋白。相比之下，采用CN104450607A方法细胞倍增所需时间更长，最高细胞密度更低，整个培养过程细胞活率较差。
虽然，上文中已经用一般性说明及具体实施方案对本发明作了详尽的描述，但在本发明基础上，可以对之做一些修改或改进，这对本领域技术人员而言是显而易见的。因此，在不偏离本发明精神的基础上所做的这些修改或改进，均属于本发明要求保护的范围。

Claims (6)

1.全化学成分HEK293细胞培养基，其特征在于，所述培养基包括氨基酸、无机盐、维生素、微量元素、碳水化合物和有机分子；
其中，氨基酸的组成及用量如下：
Figure FDA0002645087420000011
无机盐的组成及用量如下：
Figure FDA0002645087420000012
微量元素的组成及用量如下：
Figure FDA0002645087420000013
维生素的组成及用量如下：
Figure FDA0002645087420000014
Figure FDA0002645087420000021
碳水化合物和有机分子的组成及用量如下：
Figure FDA0002645087420000022
2.根据权利要求1所述的培养基，其特征在于，
其中，氨基酸的组成及用量如下：
Figure FDA0002645087420000023
无机盐的组成及用量如下：
Figure FDA0002645087420000031
微量元素的组成及用量如下：
Figure FDA0002645087420000032
维生素的组成及用量如下：
Figure FDA0002645087420000033
碳水化合物和有机分子的组成及用量如下：
Figure FDA0002645087420000034
Figure FDA0002645087420000041
3.根据权利要求1或2所述的培养基，其特征在于，pH值为6.9-7.1。
4.权利要求3所述培养基的配制方法，其特征在于，包括：按比例称取各组分，用水溶解，然后用水定容，并调pH至6.9-7.1。
5.权利要求1-3任一项所述培养基在HEK293细胞或重组HEK293细胞体外培养中的应用。
6.利用权利要求1-3任一项所述培养基培养HEK293细胞的方法，其特征在于，包括：在装有所述培养基的生物反应器中悬浮培养HEK293细胞，悬浮培养的条件为：温度36.5-37.0℃，pH 6.9-7.1，溶氧浓度40～45％；接种的起始细胞浓度为0.5×106～0.7×106个细胞/mL。
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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114836367A (zh) * 2022-04-28 2022-08-02 上海东富龙生物试剂有限公司 用于hek293细胞培养及腺病毒、腺相关病毒复制扩增的化学成分限定培养基

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101052708A (zh) * 2004-11-02 2007-10-10 阿雷斯贸易股份有限公司 哺乳动物细胞的无血清细胞培养基
CN102876626A (zh) * 2011-07-14 2013-01-16 宁波安柯普顿生物技术有限公司 支持cho高密度悬浮生长的无血清无蛋白全化学成分界定的培养基
CN104450607A (zh) * 2014-11-14 2015-03-25 武汉金开瑞生物工程有限公司 用于hek293细胞悬浮生长的全化学成分培养基及培养方法
CN106190950A (zh) * 2016-07-01 2016-12-07 北京双鹭药业股份有限公司 一种cho细胞无血清无蛋白培养基及其制备方法
CN111304149A (zh) * 2020-02-21 2020-06-19 新乡医学院 一种支持hek293细胞悬浮培养的无血清无蛋白培养基及其制备方法和应用

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101052708A (zh) * 2004-11-02 2007-10-10 阿雷斯贸易股份有限公司 哺乳动物细胞的无血清细胞培养基
CN102876626A (zh) * 2011-07-14 2013-01-16 宁波安柯普顿生物技术有限公司 支持cho高密度悬浮生长的无血清无蛋白全化学成分界定的培养基
CN104450607A (zh) * 2014-11-14 2015-03-25 武汉金开瑞生物工程有限公司 用于hek293细胞悬浮生长的全化学成分培养基及培养方法
CN106190950A (zh) * 2016-07-01 2016-12-07 北京双鹭药业股份有限公司 一种cho细胞无血清无蛋白培养基及其制备方法
CN111304149A (zh) * 2020-02-21 2020-06-19 新乡医学院 一种支持hek293细胞悬浮培养的无血清无蛋白培养基及其制备方法和应用

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
MA SK ET AL.: "Overexpression of G-Protein-Coupled Receptor 40 Enhances the Mitogenic Response to Epoxyeicosatrienoic Acids", 《PLOS ONE》, pages 0113130 *
朱晓宇 石军: "锌对人骨髓间充质干细胞体外增殖和成脂分化的影响", 《广西医学》, pages 928 - 933 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN114836367A (zh) * 2022-04-28 2022-08-02 上海东富龙生物试剂有限公司 用于hek293细胞培养及腺病毒、腺相关病毒复制扩增的化学成分限定培养基

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