Zeng Q S, Takeyama H, Kanda S, Irie R F
John Wayne Cancer Institute, Santa Monica, CA 90404.
Hum Antibodies Hybridomas. 1994;5(1-2):75-80.
In this study we demonstrated that fetal calf serum (FCS) depleted of IgG by protein G affinity chromatography (G-FCS) is superior to whole FCS or serum-free culture media as a culture supplement for the production of purified IgG monoclonal antibodies (MAb). One hundred ml FCS was applied to a 25 ml protein G Sepharose 4 Fast Flow Column, which was shaken gently for 2 days at 4 degrees C. The procedure was repeated using a protein G column for an additional day. G-FCS was used at a concentration of 5% in RPMI 1640 medium to grow the mouse myeloma cell line P3X63.Ag8.653, which secretes an IgG-1 mouse-human chimeric monoclonal antibody (TVE-1). Cell density, viability, doubling time, and antibody production were used as indices to compare the efficacy of this medium with that of whole FCS medium, AIM-V (Gibco, USA) and other serum-free media. The results demonstrate that cell growth and antibody production in -GFCS medium did not differ significantly from that in FCS medium, but were significantly better than in the serum-free media (p < 0.001). TVE-1 antibody in the spent tissue culture media was purified by 50% ammonium sulfate precipitation and Protein A affinity chromatography. An antibody that was more than 99% pure was obtained. Endotoxin analysis revealed that the IgG depletion process does not generate a significant level of endotoxin in FCS (< 0.06 EU/ml).(ABSTRACT TRUNCATED AT 250 WORDS)
在本研究中，我们证明了通过蛋白G亲和层析去除IgG的胎牛血清（G-FCS）作为纯化IgG单克隆抗体（MAb）生产的培养补充剂优于全胎牛血清或无血清培养基。将100 ml胎牛血清应用于25 ml蛋白G Sepharose 4 Fast Flow柱，在4℃下轻轻摇晃2天。使用蛋白G柱再重复该过程一天。G-FCS以5%的浓度用于RPMI 1640培养基中培养分泌IgG-1小鼠-人嵌合单克隆抗体（TVE-1）的小鼠骨髓瘤细胞系P3X63.Ag8.653。细胞密度、活力、倍增时间和抗体产量用作指标，比较该培养基与全胎牛血清培养基、AIM-V（美国Gibco公司）和其他无血清培养基的效果。结果表明，G-FCS培养基中的细胞生长和抗体产生与胎牛血清培养基中的无显著差异，但明显优于无血清培养基（p < 0.001）。用过的组织培养基中的TVE-1抗体通过50%硫酸铵沉淀和蛋白A亲和层析进行纯化。获得了纯度超过99%的抗体。内毒素分析表明，IgG去除过程不会在胎牛血清中产生显著水平的内毒素（< 0.06 EU/ml）。（摘要截短至250字）
