How do non-ideal conditions affect imaging? A mismatch of immersion medium with a homogenous sample will cause depth-dependent loss of contrast and resolution, focal position shifts between varying wavelengths and axial focus errors for all forms of fluorescence microscopy. These effects result in observed aberrations in the expected point spread function(PSF -- the blurring of a point source recorded by a detector over a specified focal range) for conventional microscopy and depth-dependent attenuation for confocal microscopy, as well. Additionally, properties of the sample, such as refractive index differences between different subregions within a cell will also affect imaging quality.
The goal of my project is to develop means to quantify and computationally compensate for these imaging aberrations for conventional (widefield) microscopy during the deconvolution process, where data collection lens' PSF is used to remove the out-of-focus information. I will take a series of steps to move from the current, spatially invariant deconvolution to a process which will allow measured sample properties to direct a spatially variant deconvolution. I also hope to develop evaluation tools and a modeling system with which one can determine the best method for accomplishing this end.
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