Single-molecule localization microscopy(SMLM)enables three-dimensional(3D)investigation of nanoscale structures in biological samples,offering unique insights into their organization.However,traditional 3D super-resol...Single-molecule localization microscopy(SMLM)enables three-dimensional(3D)investigation of nanoscale structures in biological samples,offering unique insights into their organization.However,traditional 3D super-resolution microscopy using high numerical aperture(NA)objectives is limited by imaging depth of field(DOF),restricting their practical application to relatively thin biological samples.Here,we developed a unified solution for thick sample super-resolution imaging using a deformable mirror(DM)which served for fast remote focusing,optimized point spread function(PSF)engineering,and accurate aberration correction.By effectively correcting the system aberrations introduced during remote focusing and sample aberrations at different imaging depths,we achieved high-accuracy,large DOF imaging(~8μm)of the whole-cell organelles[i.e.,nuclear pore complex(NPC),microtubules,and mitochondria]with a nearly uniform resolution of approximately 35 nm across the entire cellular volume.展开更多
基金National Natural Science Foundation of China(62375116)Shenzhen Medical Research Fund(B2302038)+2 种基金Key Technology Research and Development Program of Shandong(2021CXGC010212)Science,Technology and Innovation Commission of Shenzhen Municipality(JCYJ20220818100416036,KQTD20200820113012029)Startup Grant from Southern University of Science and Technology。
文摘Single-molecule localization microscopy(SMLM)enables three-dimensional(3D)investigation of nanoscale structures in biological samples,offering unique insights into their organization.However,traditional 3D super-resolution microscopy using high numerical aperture(NA)objectives is limited by imaging depth of field(DOF),restricting their practical application to relatively thin biological samples.Here,we developed a unified solution for thick sample super-resolution imaging using a deformable mirror(DM)which served for fast remote focusing,optimized point spread function(PSF)engineering,and accurate aberration correction.By effectively correcting the system aberrations introduced during remote focusing and sample aberrations at different imaging depths,we achieved high-accuracy,large DOF imaging(~8μm)of the whole-cell organelles[i.e.,nuclear pore complex(NPC),microtubules,and mitochondria]with a nearly uniform resolution of approximately 35 nm across the entire cellular volume.
