Masullo Luciano A, Szalai Alan M, Lopez Lucía F, Pilo-Pais Mauricio, Acuna Guillermo P, Stefani Fernando D
Centro de Investigaciones en Bionanociencias (CIBION), Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Godoy Cruz 2390, C1425FQD Ciudad Autónoma de Buenos Aires, Buenos Aires, Argentina.
Departamento de Física, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Güiraldes 2620, C1428EHA Ciudad Autónoma de Buenos Aires, Buenos Aires, Argentina.
Light Sci Appl. 2022 Jun 30;11(1):199. doi: 10.1038/s41377-022-00896-4.
Localization of single fluorescent emitters is key for physicochemical and biophysical measurements at the nanoscale and beyond ensemble averaging. Examples include single-molecule tracking and super-resolution imaging by single-molecule localization microscopy. Among the numerous localization methods available, MINFLUX outstands for achieving a ~10-fold improvement in resolution over wide-field camera-based approaches, reaching the molecular scale at moderate photon counts. Widespread application of MINFLUX and related methods has been hindered by the technical complexity of the setups. Here, we present RASTMIN, a single-molecule localization method based on raster scanning a light pattern comprising a minimum of intensity. RASTMIN delivers ~1-2 nm localization precision with usual fluorophores and is easily implementable on a standard confocal microscope with few modifications. We demonstrate the performance of RASTMIN in localization of single molecules and super-resolution imaging of DNA origami structures.
单个荧光发射体的定位是纳米级及超总体平均的物理化学和生物物理测量的关键。示例包括单分子追踪和通过单分子定位显微镜进行的超分辨率成像。在众多可用的定位方法中，MINFLUX脱颖而出，与基于宽场相机的方法相比，其分辨率提高了约10倍，在中等光子计数下即可达到分子尺度。MINFLUX及相关方法的广泛应用受到了装置技术复杂性的阻碍。在此，我们介绍RASTMIN，一种基于光栅扫描包含最低强度的光图案的单分子定位方法。RASTMIN使用普通荧光团可实现约1-2纳米的定位精度，并且只需进行少量修改就能轻松在标准共聚焦显微镜上实现。我们展示了RASTMIN在单分子定位和DNA折纸结构超分辨率成像方面的性能。
