Methods And System For Compensating For Spatial Cross-Talk
An embodiment generally relates to a method of processing signals. The method includes providing for a plurality of filters, where each filter is configured to process an associated dye. The method also includes determining a residual error for at least one filter during dye amplification and modifying the at least one filter based on the residual error. The method further includes filtering subsequent signals associated with the modified at least one filter.
1 . An apparatus for calibrating for spatial cross-talk correction in an imaging system, the apparatus comprising:
a plate comprising:
an array of pinholes, each pinhole substantially smaller than the resolution of the imaging system, wherein the plate is configured to be illuminated and imaged by the imaging system to correct for spatial cross-talk.
2 . The apparatus of claim 1 , wherein each pinhole of the array of pinholes is spaced at least three times the separation of features being measured on the imaging system.
3 . A method of using the apparatus of claim 1 , the method comprising:
imaging the apparatus illuminated to form an initial image;
smoothing the initial image; and
subtracting the initial image from the smoothed initial image to form a calibrated image.
4 . The method of claim 3 , the method further comprising:
partitioning the calibrated image into a plurality of regions, each region comprising at least one pinhole;
creating at least one subimage from a selected region of the plurality of regions; and normalizing any features located in the at least one subimage.
5 . The method of claim 4 , further comprising:
determining an intensity weighted centroid associated for each pinhole located in the at least one subimage; and
shifting the intensity weighted centroid to a middle of the subimage.
6 . The method of claim 5 , further comprising averaging the at least one subimage to provide a final signal-to-noise point spread function (“PSF”) image.
7 . The method of claim 5 , further comprising using median aggregation on the at least one subimage to provide a signal-to-noise PSF image.
8 . The method of claim 3 , the method further comprising:
convolving the calibrated image with a feature profile to form a convolved image; and quantifying the convolved image using a user's algorithm at a selected pinhole and neighboring pinholes to the selected pinhole.
9 . The method of claim 8 , the method further comprising determining a cross-talk coefficient for the selected pinhole in each neighbor direction divided by a convolved pinhole flux.
10 . The method of claim 9 , further comprising estimating a signal-to-noise ratio for the selected pinhole by aggregating the values associated with the neighboring pinholes for each direction.
11 . An apparatus comprising of means for performing the method of claim 3 .
12 . A computer-readable medium comprising computer-executable instructions for performing the method of claim 3 .
13 . A system comprising of:
a calibration plate;
a light source configured to illuminate the calibration plate; and
a processor configured to receive digitize image of an illuminated calibration plate, wherein the processor is configured to image the illuminated calibration plate to form an initial image, smoothing the initial image, and subtract the initial image from the smoothed initial image to form a calibrated image.
14 . The system of claim 13 , wherein the processor is further configured to partition the calibrated image into a plurality of regions, each region comprising at least one pinhole and create at least one subimage from a selected region of the plurality of regions.
15 . The system of claim 14 , wherein the processor is further configured to normalize any features located in the at least one subimage.
16 . The system of claim 15 , wherein the processor is further configured to determine an intensity weighted centroid associated for each pinhole located in the at least one subimage and shift the intensity weighted centroid to a middle of the subimage.
17 . The system of claim 14 , wherein the processor is further configured to average the at least one subimage to provide a final signal-to-noise point spread function (“PSF”) image.
18 . The system of claim 17 , wherein the processor is further configured to use median aggregation on the at least one subimage to provide a signal-to-noise PSF image.
19 . The system of claim 18 , wherein the processor is further configured to convolve the calibrated image with a feature profile to form a convolved image and quantify the convolved image using a user's algorithm at a selected pinhole and neighboring pinholes to the selected pinhole.
20 . The system of claim 19 , wherein the processor is further configured to determine a cross-talk coefficient for the selected pinhole in each neighbor direction divided by a convolved pinhole flux.