Aligning grids derived from fluorescent images of an array of signal sources

Abstract

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.

Description

BACKGROUND[0001]Recent rapid advances in molecular biology have created more demand for high volume testing based on the need to screen ever larger compound libraries, validate ever increasing numbers of genetic markers and test ever more diversified patient populations. This has led to the development of new array formats, particularly for nucleic acid and protein-protein interaction analysis, which permit parallel processing by performing requisite assays in a “multiplexed” format.[0002]Conventionally, such assays are performed by producing arrays of nucleic acids and antibodies by way of “spotting” or “printing” of aliquot solutions on filter paper, blotting paper or other substrates. However, notwithstanding their widespread current use in academic research for gene expression analysis and protein profiling, arrays produced by spotting have shortcomings, particularly in applications placing high demands on accuracy and reliability and calling for the analysis of a large volume o...

AI Technical Summary

Problems solved by technology

However, this technology is limited in practice to producing short oligonucleotide probes—as a result, alternative technologies are required for the production of cDNA and protein arrays.

In addition, in-situ synthersis precludes rapid probe array customization given the time and cost involved in the requisite redesign of the photochemical synthesis process.

In addition to these inherent difficulties relating to assay performance, spatially encoded arrays produced by spotting or in-situ synthesis conventional methods generally require specialized optical scanning instrumentation to extract data of useable quality.

However, several problems must be overcome.

First, beads displaying receptors which do not produce an assay signal will not be visible in the fluorescence image, and this generally will lead to the corruption of grid edges, thereby affecting the step of image rotation which relies on the accurate determination of the original image orientation by measuring the orientation of edges.

Second, signal sources such as fluorescent beads may be randomly displaced from the center of each grid field, a fact which can affect the intensity recorded from that field.

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