Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, 110 8th Street, Troy, New York 12180, USA.
Biotechnol Bioeng. 2013 Oct;110(10):2664-76. doi: 10.1002/bit.24945. Epub 2013 May 16.
This study provides a detailed investigation into the performance of a stimuli responsive ELP-Z based process for monoclonal antibody (mAb) affinity precipitation. A multidimensional high-throughput screening (HTS) protocol was developed and employed to investigate the effects of a variety of operating conditions on mAb yield and aggregation during the process. Precipitation efficiency of ELP-Z in the absence of mAb was first determined as a function of temperature and sodium sulfate concentration and conditions producing high yields were identified. HTS was then employed to determine appropriate conditions for the initial capture and co-precipitation of mAbs at high yields using ELP-Z. mAb elution from ELP-Z was then examined using HTS and the mAb yields and aggregate content of the overall process were determined. It was observed that mAb aggregation was sensitive primarily to the elution conditions and that this behavior was antibody specific and a strong function of operating temperature and elution pH. Importantly, for both mAbs examined in this study, the results indicated that room temperature operation and appropriate elution pH could be readily employed to produce both high mAb yields and low aggregate content using this approach. This study demonstrates the ability of ELP-Z based affinity precipitation for mAb purification and shows that HTS can be successfully employed to rapidly develop a robust and high yield process.
本研究深入探讨了基于响应性 ELP-Z 的单克隆抗体 (mAb) 亲和沉淀工艺的性能。开发并采用多维高通量筛选 (HTS) 方案来研究各种操作条件对该过程中 mAb 产率和聚集的影响。首先确定了在不存在 mAb 的情况下 ELP-Z 的沉淀效率随温度和硫酸钠浓度的变化，并确定了产生高产率的条件。然后采用 HTS 确定使用 ELP-Z 以高产率进行 mAb 初始捕获和共沉淀的合适条件。然后通过 HTS 检查 ELP-Z 从 mAb 的洗脱情况，并确定整个过程中 mAb 的产率和聚集含量。结果表明，mAb 的聚集主要对洗脱条件敏感，并且这种行为具有抗体特异性，强烈依赖于操作温度和洗脱 pH。重要的是，对于本研究中检查的两种 mAb，结果表明，使用这种方法，室温操作和适当的洗脱 pH 可以很容易地用于生产高 mAb 产率和低聚集含量。本研究证明了基于 ELP-Z 的亲和沉淀在 mAb 纯化方面的能力，并表明 HTS 可成功用于快速开发稳健且高产率的工艺。
