An analysis of the relationship between the number of plates measured with a small molecule tracer and the breakthrough curve of a strongly bound protein in short laboratory chromatography columns (1–5 cm) considering flow nonuniformity is presented. For practical conditions, while axial dispersion has only a small impact on the breakthrough curve, radial flow nonuniformity has a profound effect. Radial parabolic velocity profiles lead to tailing tracer peaks and broader breakthrough curves. Profiles where the velocity varies radially only in a thin region near the column wall lead to fronting tracer peaks and early breakthrough when the velocity at the wall is higher than the average and to tailing peaks and tailing breakthrough curves when the velocity at the wall is lower than the average. Experiments conducted in laboratory minicolumns (0.5–1 cm diameter, 0.5–1 ml volume) show tracer peaks and protein breakthrough curves that are consistent with higher velocities at the wall. The model presented in this work provides a tool to model experimental breakthrough data and to assess the degree of flow uniformity required to obtain meaningful dynamic binding capacity measurements using minicolumns in a high‐throughput lab setting.

中文翻译：

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短色谱柱中 HETP 测量值与穿透曲线之间的关系

分析了使用小分子示踪剂测量的板数与考虑流动不均匀性的短实验室色谱柱（1-5 cm）中强结合蛋白的穿透曲线之间的关系。对于实际情况，虽然轴向扩散对突破曲线的影响很小，但径向流动不均匀性具有深远的影响。径向抛物线速度剖面导致拖尾示踪峰和更宽的突破曲线。速度仅在靠近柱壁的薄区域呈径向变化的剖面，当壁处速度高于平均值时，会导致示踪剂峰前伸和早期突破，而当壁处速度较低时，会导致拖尾峰和拖尾突破曲线高于平均水平。在实验室微型柱中进行的实验 (0. 5-1 cm 直径，0.5-1 ml 体积）显示示踪峰和蛋白质穿透曲线，这些曲线与壁处的较高速度一致。本工作中提出的模型提供了一种工具来模拟实验突破数据并评估在高通量实验室环境中使用微型柱获得有意义的动态结合容量测量所需的流动均匀度。