Abstract：
Aggregation of monoclonal antibodies (mAbs) is the driving force for their undesirable immunogenic effects. There are multiple factors responsible for aggregation in therapeutic proteins. One significant cause is the process-related shear and interfacial stress generated due to impellers and stirrers. This investigation focuses on understanding the possible aggregation arising upon stirring mAb formulations using stirrers made of different materials. We used quantitative laser diffraction (qLD) to monitor and quantify the stirring induced formation of submicron and subvisible aggregates in the size range from 100 nm to 10 µm. We analysed various aspects of aggregate generation, such as onset of aggregation, particle size distribution, and concentration of aggregates generated using stirrers of different materials. We observed that mixing with stainless steel stirrers resulted in a quicker onset of aggregation and led to significantly higher concentrations of aggregates. Analysis of the stirred samples using dynamic light scattering (DLS) and background imaging technique (BMI) were conducted to complement the qLD analysis. All the three techniques resulted in a similar trend, showing presence of larger and higher quantities of aggregates in steel stirred samples, as compared to those stirred using PEEK and glass. Additionally, we performed SEC-HPLC to quantify the soluble fraction of monomer and recorded that the least amount was present in the steel stirred samples. This work highlights the need for optimizing the materials used for fabricating the stirrers/impellers.
摘要：
单克隆抗体(mAb)的聚集是其不期望的免疫原性作用的驱动力。有多种因素导致治疗性蛋白质的聚集。一个重要原因是由于叶轮和搅拌器产生的与过程相关的剪切和界面应力。该研究集中于理解在使用由不同材料制成的搅拌器搅拌mAb制剂时产生的可能聚集。我们使用定量激光衍射（qLD）来监测和量化搅拌诱导的亚微米和亚可见聚集体的形成，其尺寸范围为100nm至10µm。我们分析了总量生成的各个方面，例如聚集的开始，粒度分布，以及使用不同材料的搅拌器产生的骨料浓度。我们观察到，与不锈钢搅拌器混合导致聚集的更快开始，并导致聚集体的浓度显着升高。使用动态光散射(DLS)和背景成像技术(BMI)进行搅拌样品的分析以补充qLD分析。所有这三种技术都导致了类似的趋势，显示在钢搅拌样品中存在越来越多的骨料，与使用PEEK和玻璃搅拌的那些相比。此外,我们进行SEC-HPLC以定量单体的可溶性部分,并记录在钢搅拌样品中存在最少量。这项工作突出了优化用于制造搅拌器/叶轮的材料的需要。
