Whole-cell, multicolor superresolution imaging using volumetric multifocus microscopy

Significance
A major challenge in modern biological studies is in the determination of the 3D molecular architecture of cellular organelles. In recent years, much progress in nanoscale imaging has been made because of the advent of superresolution optical microscopy. However, many superresolution techniques are still limited to 2D acquisition. Here, we show a volumetric approach for superresolution imaging based on the simultaneous imaging of multiple sample planes using multifocal microscopy. The depth over which structures can be reconstructed reaches 4 µm, comparable with the thickness of many cellular organelles or even whole cells.