Iterative immunostaining combined with expansion microscopy and image processing reveals nanoscopic network organization of nuclear lamina.

ABSTRACT: Investigation of nuclear lamina architecture relies on superresolved microscopy. However, epitope accessibility, labeling density, and detection precision of individual molecules pose challenges within the molecularly crowded nucleus. We developed iterative indirect immunofluorescence (IT-IF) staining approach combined with expansion microscopy (ExM) and structured illumination microscopy to improve superresolution microscopy of subnuclear nanostructures like lamins. We prove that ExM is applicable in analyzing highly compacted nuclear multiprotein complexes such as viral capsids and provide technical improvements to ExM method including three-dimensional-printed gel casting equipment. We show that in comparison with conventional immunostaining, IT-IF results in a higher signal-to-backgrou

SUBMITTER: Mantyla E 

PROVIDER: S-EPMC10398900 | biostudies-literature | 2023 Aug

REPOSITORIES: biostudies-literature