In vivo Three-Dimensional Superresolution Fluorescence Tracking using a Double-Helix Point Spread Function.
The point spread function (PSF) of a widefield fluorescence microscope is not suitable for three-dimensional super-resolution imaging. We characterize the localization precision of a unique method for 3D superresolution imaging featuring a double-helix point spread function (DH-PSF). The DH-PSF is designed to have two lobes that rotate about their midpoint in any transverse plane as a function of the axial position of the emitter. In effect, the PSF appears as a double helix in three dimensions. By comparing the Cramer-Rao bound of the DH-PSF with the standard PSF as a function of the axial position, we show that the DH-PSF has a higher and more uniform localization precision than the standard PSF throughout a 2 μm depth of field. Comparisons between the DH-PSF and other methods for 3D super-resolution are briefly discussed. We also illustrate the applicability of the DH-PSF for imaging weak emitters in biological systems by tracking the movement of quantum dots in glycerol and in live cells.
Lew
Matthew D
MD
Department of Electrical Engineering, Stanford University, Stanford, CA 94305.
Thompson
Michael A
MA
Badieirostami
Majid
M
Moerner
W E
WE