Image processing and analysis of array data
View Patent ↗Disclosed are methods of processing images, and in particular aligning and orienting grids from a fluorescent array of signal sources, by linking nearest neighbor sources to form a hexagon, and then using the hexagon lines for alignment and orientation of the grid. Also disclosed are methods and algorithms for aligning image with grid and correcting for signal beads which are smaller than a grid field and the shift in a grid field. These methods can be used where the signal sources are fluorescent images from a microarray. Also disclosed are methods of automated watershed clustering following transformation.
1. A method of finding a grid from a fluorescent image of an array of signal sources, comprising:
finding the signal source centers;
building lines connecting neighboring signal sources into hexagons;
partitioning hexagon lines;
finding the orientation angle of the array;
finding row boundaries of the array;
finding column boundaries of the array;
transforming the entire array align at zero orientation angle with an image frame;
finding row grid lines on the transformed array;
finding column grid lines on the transformed array;
if the rows of the array are vertical, placing all X coordinates of transformed signal source centers into a set of row positions;
if rows of the array are horizontal, collecting all Y coordinates of transformed signal source centers into the set row positions;
partitioning the set of row positions into groups, where adjacent groups are within a first threshold distance;
calculating and sorting groups in order of group means;
removing groups where The distance to a neighbor group is within a second threshold distance;
determining if gaps between groups exceed the first threshold distance, and, if so, inserting an appropriate number of groups so that groups are within the first threshold distance from each other;
determining the respective relative positions of rows based on the mean of the member beads in the respective rows;
generating a set of grid lines such that member grid lines in the set are midway between adjacent rows; and
adding one top line, parallel to the end lines and adjacent to the first row of the array, and adding one bottom line, parallel to the grid lines and adjacent to the last row of the array, said top and bottom lines being at a distance from the respective nearest grid line which is the same as the distance between adjacent grid lines.
2. The method of claim 1 further including
separating transformed signal sources into two groups, where one of the groups contains signal sources from odd numbered rows, while the other group contains signal sources from even numbered rows.
3. The method of claim 2 further including finding two sets of column grid lines by separating transformed signal sources into two sets, where one set contains signal sources from odd numbered rows, and the other set contains signal sources from even numbered rows.
4. The method of claim 1 wherein individual signal sources are fluorescent beads.