Acquisition speed in high-resolution imaging in three dimensions (3D) remainsa major challenge in modern microscopy. When 3D information is collected sequentially,
acquisition speed is limited and increases with the sample volume size
imaged. Aberration-corrected multifocus microscopy (MFM) employs diffractive
Fourier optics to multiplex and refocus the microscope image to enable truly simultaneous
3D imaging, without loss of resolution. This thesis describes a new
type of ultra-fast 25-camera-array multifocus microscope (M25) which employs
an array of small, fast CMOS cameras sensitive enough for live 3D fluorescence
imaging. The use of multiple sensors allows M25 to significantly push acquisition
speed, depth or field, and field of view to capture 130×130×50 μm3 volumes
at ~100 Hz. This new optical design also employs a radically simplified chromatic
correction module from classical MFM systems, consisting of simple pairs
of diffractive elements. The prototype instrument here described is customized for
functional neural circuit imaging and locomotion in small model organisms and is
demonstrated on Caenorhabditis elegans, fruit fly (Drosophila melanogaster), and
lamprey (Petromyzon marinus).