Mahboudi Fereidoun, Abolhassan Mohammad Reza, Azarpanah Armita, Aghajani-Lazarjani Hamideh, Sadeghi-Haskoo Mohammad Amin, Maleknia Shaian, Vaziri Behrouz
Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.
Avicenna J Med Biotechnol. 2013 Jul;5(3):140-7.
Recombinant monoclonal antibodies have been marketed in last three decades as the major therapeutic proteins against different cancers. However choosing a proper medium and supplements to reach the high expression is a challenging step. Despite of commercial serum free and chemically defined media, there are still numerous researches seeking the optimum media to gain higher expression titer. Selecting the best basal media followed by proper supplementation to increase the cell density and expression titer needs proper and accurate investigation.
In this study, we have determined the expression titer of monoclonal antibody against human CD20 using soy extract, Essential Amino Acid, Non-Essential Amino Acid, Panexin NTS, Peptone, Yeast extract, Insulin-transferrin selenite, Human Serum Albumin, Bovine Serum Albumin, Lipid, and two commercially available supplements, Power and Xtreme feed. In each experiment, the expression level was compared with a well defined media, ProCHO5, RPMI 1640 and DMEM-F12.
It has been shown that supplementing the ProCHO5 basal medium with 10% power feed or combination of 5% PanexinNTS,1.5 g/L yeast and 1.5g/L peptone results in the best production levels with 450 and 425 mg/L of anti CD20 mAb expression level, respectively.
Panexin NTS, yeast and peptone cane be proper supplement for fed-batch cell culture instead of commercial Power feed supplement which is a cost effective way to increase expression level. And thereby ProCHO5 may be replaced with common media such as RPMI 1640 and DMEM-F12.
在过去三十年中,重组单克隆抗体作为针对不同癌症的主要治疗性蛋白质已投放市场。然而,选择合适的培养基和添加剂以实现高表达是具有挑战性的一步。尽管有商业化的无血清和化学成分明确的培养基,但仍有大量研究在寻找能获得更高表达滴度的最佳培养基。选择最佳基础培养基并进行适当补充以提高细胞密度和表达滴度需要进行恰当且准确的研究。
在本研究中,我们使用大豆提取物、必需氨基酸、非必需氨基酸、人参皂苷NTS、蛋白胨、酵母提取物、胰岛素 - 转铁蛋白亚硒酸盐、人血清白蛋白、牛血清白蛋白、脂质以及两种市售添加剂Power和Xtreme feed,测定了抗人CD20单克隆抗体的表达滴度。在每个实验中,将表达水平与一种明确的培养基ProCHO5、RPMI 1640和DMEM - F12进行比较。
结果表明,用10%的Power feed补充ProCHO5基础培养基,或用5%的人参皂苷NTS、1.5 g/L酵母和1.5 g/L蛋白胨的组合进行补充,可分别获得最佳产量水平,抗CD20单克隆抗体表达水平分别为450和425 mg/L。
人参皂苷NTS、酵母和蛋白胨可作为补料分批细胞培养的合适添加剂,取代商业化的Power feed添加剂,这是提高表达水平的一种经济有效的方法。因此,ProCHO5可用RPMI 1640和DMEM - F12等常用培养基替代。