Super-resolution microscopy (SRM) has transformed our understanding of proteins' subcellular organization and revealed cellular details down to nanometers, far beyond conventional microscopy. While localization precision is independent of the number of fluorophores attached to a biomolecule, labeling density is a decisive factor for resolving complex biological structures. The average distance between adjacent fluorophores should be less than half the desired spatial resolution for optimal clarity. While this was not a major limitation in recent decades, the success of modern microscopy approaching molecular resolution down to the single-digit nanometer range will depend heavily on advancements in fluorescence labeling. This review highlights recent advances and challenges in labeling strategies for SRM, focusing on site-specific labeling technologies. These advancements are crucial for improving SRM precision and expanding our understanding of molecular interactions.

中文翻译：

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超分辨率显微镜的位点特异性蛋白质标记策略

超分辨率显微镜 (SRM) 改变了我们对蛋白质亚细胞组织的理解，并揭示了小至纳米的细胞细节，远远超出了传统显微镜的范围。虽然定位精度与生物分子上附着的荧光团数量无关，但标记密度是解析复杂生物结构的决定性因素。相邻荧光团之间的平均距离应小于所需空间分辨率的一半，以获得最佳清晰度。虽然这并不是近几十年来的主要限制，但现代显微镜能否成功将分子分辨率降低到个位数纳米范围将在很大程度上取决于荧光标记的进步。本综述重点介绍了 SRM 标签策略的最新进展和挑战，重点关注特定位点的标签技术。这些进步对于提高 SRM 精度和扩大我们对分子相互作用的理解至关重要。