Fast and sensitive multi-color 3D imaging using aberration-corrected multi-focus microscopy

Conventional acquisition of three-dimensional (3D) microscopy data requires sequential z-scanning and is often too slow to capture biological events. We report a new aberration-corrected multi-focus microscopy method capable of producing an instant focal stack of nine 2D images. Appended to an epifluorescence microscope, the multi-focus system enables high-resolution 3D imaging in multiple colors with single molecule sensitivity, at speeds limited by the camera readout time of a single image.