G protein-coupled receptors (GPCRs) are organised within the cell membrane into highly ordered macromolecular complexes along with other receptors and signalling proteins. Understanding how heterogeneity in these complexes affects the pharmacology and functional response of these receptors is crucial for developing new and more selective ligands. Fluorescence correlation spectroscopy (FCS) and related techniques such as photon counting histogram (PCH) analysis and image-based FCS can be used to interrogate the properties of GPCRs in these membrane microdomains, as well as their interaction with fluorescent ligands. FCS analyses fluorescence fluctuations within a small-defined excitation volume to yield information about their movement, concentration and molecular brightness (aggregation). These techniques can be used on live cells with single-molecule sensitivity and high spatial resolution. Once the preserve of specialist equipment, FCS techniques can now be applied using standard confocal microscopes. This review describes how FCS and related techniques have revealed novel insights into GPCR biology.

G 蛋白偶联受体 (GPCR) 在细胞膜内与其他受体和信号蛋白一起组织成高度有序的大分子复合物。了解这些复合物中的异质性如何影响这些受体的药理学和功能反应对于开发新的和更具选择性的配体至关重要。荧光相关光谱 (FCS) 和相关技术，如光子计数直方图 (PCH) 分析和基于图像的 FCS，可用于询问这些膜微域中 GPCR 的特性，以及它们与荧光配体的相互作用。FCS 分析小定义激发体积内的荧光波动，以产生有关其运动、浓度和分子亮度（聚集）的信息。这些技术可用于具有单分子灵敏度和高空间分辨率的活细胞。FCS 技术曾经是专业设备的保留地，现在可以使用标准共聚焦显微镜来应用。这篇综述描述了 FCS 和相关技术如何揭示 GPCR 生物学的新见解。