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Widefield Fluorescence Microscope point-spread function

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Widefield Fluorescence Microscope point-spread function

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Generate the point-spread function for a widefield fluorescence microscope

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Description

This function generate the point-spread function for a widefield microscope using a scalar diffraction-limited model (Stokseth refer to [1] and [3] below)
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Input parameters:
lambdaEx: Excitation Wavelength (in nm)
lambdaEm: Emission wavelegnth (in nm)
numAper: Numerical aperture of the objective
magObj: Objective total magnification
rindexObj: Refractive index of the objective immersion medium
ccdSize: Pixel dimension of the CCD (in the plane of the camera)
rindex_sp: Refractive index of the specimen medium
xysize: Size of the desired image (specimen view size/pixel dimension)
nslices: Number of slices desired (Depth view/Z axis sampling)
depth: depth of the specimen under the cover-slip in nm
dxy: CCD pixel size (in nm)
dz: Optical axis Z sampling or defocusing (in nm)
nor: Normalization on the PSF (default: no normalization)
0: L-infinity normalization
1: L-1 normalization
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Example usage:
Note that for large z sections, the algorithms speed is dependent on the 2D fFTs
Without spherical aberration, and normalized to l-infinity:
Microscope objective: 63X/1.4, oil immersion
CCD camera pixel size: 6.45 microns
Fluorophore is GFP and excited in blue
>> psf = wfmpsf(488, 520, 1.4, 63, 1.518, 6450, 300, 128, 64, 1.518, 0, 0);
With spherical aberration:
Microscope objective: 63X/1.4, oil immersion
CCD camera pixel size: 6.45 microns
Specimen immersed has a buffer index of 1.45
and imaged at 15 microns in depth
Fluorophore is GFP and excited in blue
>> psf = wfmpsf(488, 520, 1.2, 40, 1.33, 6450, 300, 256, 128, 1.45, 15000, 0);
To plot the PSFs:
>> load cmap_fire;
Remove any errors due to the Fourier transform
>> psf(psf<5.7e-06) = 5.7e-06;
>> figure, imagesc(maxintensityproj(log(psfl), 1)');
>> colormap(cmap_fire); colorbar;
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References:
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[1] P. A. Stokseth (1969). `Properties of a defocused optical system'. J. Opt. Soc. Am. A 59:1314?1321.
[2] P. Pankajakshan, et al. (2010). `Point-spread function model for fluorescence macroscopy imaging'. In Proc. of Asilomar Conference on Signals, Systems and Computers.
[3] P. Pankajakshan (2009). Blind Deconvolution for Confocal Laser Scanning Microscopy. Ph.D. thesis, Universite de Nice Sophia-Antipolis.
Author: Praveen Pankajakshan
Copyright (C) 2011 Praveen Pankajakshan
All rights reserved.

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MATLAB release MATLAB 7.12 (R2011a)
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Comments and Ratings (2)
09 Apr 2012 Praveen Pankajakshan

Dear Linda,
Please send me your email and I can send you the updated function with the coverglass thickness added as well.

08 Apr 2012 Linda

Very helpful function. Could you please update to accommodate very thick cover glass? In my case, we are going through the bottom of a glass slide (1mm).

Thanks!

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