西湖大学章永登小组的研究显示，带有一个物镜的镜面增强型4Pi-SMLM可实现各向同性纳米级成像。该项研究成果发表在2026年3月31日出版的《自然—生物技术》上。

在这里，该课题组介绍了镜面增强型4Pi-SMLM (me4Pi-SMLM)，这是一种单物镜配置，其主题是基于镜子的照明光束的反向反射，以产生相位可调的干涉条纹。该设计将基于像散的方法的轴向分辨率提高了约5倍，其性能可与传统的4Pi-SMLM相媲美，同时大大降低了系统的复杂性和维护。在生物样品中，me4Pi-SMLM实现近各向同性定位精度2-3 nm，使各种超微结构特征的清晰和明确的可视化。

此外，它达到了低于15脑切片的纳米各向同性分辨率，促进高保真双色成像，纳米级全细胞重建和活细胞成像。me4Pi-SMLM可以无缝集成到现有的3D-SMLM系统中，以最小的成本和努力提高性能。

据悉，双物镜4Pi单分子定位显微镜（4Pi- smlm）提供各向同性纳米级分辨率；然而，它的广泛采用受到仪器复杂性和严格的校准要求的限制。

附：英文原文

Title: Mirror-enhanced 4Pi-SMLM with one objective enables isotropic nanoscale imaging

Author: Yu, Zijing, Zheng, Bei, Zhan, Yajing, Li, Shuxin, Wang, Xulong, Dai, Qiuyang, Chen, Yanqin, Wei, Zongfang, Zhao, Wenxuan, Chen, Linlin, Tang, Jiaxin, Xia, Tianchang, He, Lingjuan, Liu, Changliang, Wu, Xudong, Yu, Xiaochun, Zhang, Yongdeng

Issue&Volume: 2026-03-31

Abstract: Dual-objective 4Pi single-molecule localization microscopy (4Pi-SMLM) offers isotropic nanoscale resolution; however, its broader adoption is limited by instrumental complexity and stringent alignment requirements. Here we introduce mirror-enhanced 4Pi-SMLM (me4Pi-SMLM), a single-objective configuration that uses mirror-based retroreflection of the illumination beam to generate phase-tunable interference fringes. This design improves the axial resolution of astigmatism-based methods by approximately fivefold, delivering performance comparable to conventional 4Pi-SMLM while greatly reducing system complexity and maintenance. me4Pi-SMLM achieves near-isotropic localization precision of 2–3nm in biological samples, enabling clear and unambiguous visualization of diverse ultrastructural features. Furthermore, it achieves sub-15nm isotropic resolution in brain slices and facilitates high-fidelity two-colour imaging, nanoscale whole-cell reconstruction and live-cell imaging. me4Pi-SMLM can be seamlessly integrated into existing 3D-SMLM systems, enhancing performance with minimal cost and effort.

DOI: 10.1038/s41587-026-03083-7

Source: https://www.nature.com/articles/s41587-026-03083-7