Asymmetric-flow field-flow fractionation (AF4) has been recognized as an invaluable tool for the characterisation of particle size, polydispersity, drug loading and stability of nanopharmaceuticals. However, the application of robust and high quality standard operating procedures (SOPs) is critical for accurate measurements, especially as these complex drug nanoformulations are most often inherently polydisperse. In this review we describe a unique international collaboration that lead to the development of a robust SOP for the measurement of physical-chemical properties of nanopharmaceuticals by multi-detector AF4 (MD-AF4) involving two state of the art infrastructures in the field of nanomedicine, the European Union Nanomedicine Characterization Laboratory (EUNCL) and the National Cancer Institute-Nanotechnology Characterisation Laboratory (NCI-NCL). We present examples of how MD-AF4 has been used for the analysis of key quality attributes, such as particle size, shape, drug loading and stability of complex nanomedicine formulations. The results highlight that MD-AF4 is a very versatile analytical technique to obtain critical information on a material particle size distribution, polydispersity and qualitative information on drug loading. The ability to conduct analysis in complex physiological matrices is an additional very important advantage of MD-AF4 over many other analytical techniques used in the field for stability studies. Overall, the joint NCI-NCL/EUNCL experience demonstrates the ability to implement a powerful and highly complex analytical technique such as MD-AF4 to the demanding quality standards set by the regulatory authorities for the pre-clinical safety characterization of nanomedicines.

非对称流场流分级分离（AF4）已被认为是表征纳米药物的粒径，多分散性，载药量和稳定性的宝贵工具。但是，可靠而高质量的标准操作程序（SOP）的应用对于精确测量至关重要，尤其是因为这些复杂的药物纳米制剂通常固有地是多分散的。在这篇综述中，我们描述了一次独特的国际合作，该合作导致了通过多检测器AF4（MD-AF4）来测量纳米药物的物理化学性质的稳健SOP的开发，涉及纳米医学领域的两个最先进的基础设施，欧盟纳米医学表征实验室（EUNCL）和美国国家癌症研究所纳米技术表征实验室（NCI-NCL）。我们提供了一些示例，说明了如何将MD-AF4用于关键质量属性的分析，例如复杂的纳米药物配方的粒径，形状，载药量和稳定性。结果表明，MD-AF4是一种非常通用的分析技术，可获取有关材料粒径分布，多分散性和载药量的定性信息的关键信息。相对于在稳定性研究领域中使用的许多其他分析技术，在复杂的生理基质中进行分析的能力是MD-AF4的另一个非常重要的优势。总体，