Method and system for measuring a molecular array background signal from a continuous background region of specified size

• US 7,221,785 B2
• Filed: 05/21/2002
• Issued: 05/22/2007
• Est. Priority Date: 05/21/2002
• Status: Expired due to Fees

• Current Assignee: Agilent Technologies, Inc.
• Original Assignee: Agilent Technologies, Inc.
• Inventors: Curry, Bo U., Ghosh, Srinka, Cattell, Herbert F.
• Primary Examiner(s): Sherau; Ishrat
• Assistant Examiner(s): SHAH, UTPAL D
• Application Number:
  US10/153,394
• Publication Number:
  US 20030219151A1
• Time in Patent Office: 1,827 Days
• Field of Search: 382/162, 345/591, 435/6
• US Class Current: 382/129
• CPC Class Codes: G06T 2207/10056 Microscopic image
• CPC Class Codes: G06T 2207/10064 Fluorescence image
• CPC Class Codes: G06T 2207/30072 Microarray; Biochip, DNA array; Well plate
• CPC Class Codes: G06T 7/11 Region-based segmentation
• CPC Class Codes: G06T 7/194 involving foreground-background segmentation

Abstract

A method and system for estimating the background signal over an arbitrarily-sized region of a scanned image of a molecular array, including a background region surrounding the ROI corresponding to the feature. A bit mask is generated, based on a molecular-array, feature-based data set that includes pixel-based intensities and a list of features, including feature coordinates and feature ROI radii, to indicate those pixels in the scanned image of the molecular array corresponding to background, and those pixels in the scanned image of the molecular array corresponding to features and ROIs. An integrated intensity for a background region of arbitrary size and shape can be efficiently determined by selecting pixels within the background region that are indicated to be background pixels in the bit mask. By selecting background anuli of sufficient size to overlap with the background anuli of neighboring features, a relatively continuous function of background-signal-verusus-position can be obtained across the surface of a molecular array in order to examine non-local, background-signal-related phenomena.

Claims

1. A method for determining a background signal intensity for a particular feature from a representation of scanned intensity values of a molecular array comprising data points and containing discrete features at specified locations, each feature having a region of interest containing a number of data points, the method comprising:

specifying a size for a background region corresponding to the feature;

selecting as background data points those data points within the specified background region;

filtering the selected data points to remove all data points that lie within specified locations of features and at least one region of interest of features in the region of specified size for the background region; and

determining the background signal intensity for the particular feature from the filtered, selected data points, wherein background-subtraction of data points from the particular feature using the determined background signal intensity facilitates calculation of accurate and reproducible log ratios between at least one of multi-channel and inter-array data.

2. The method of claim 1 further including:

preparing a background mask with elements, corresponding to data points scanned from the molecular array, containing indications of whether or not corresponding data points are background data points.

3. The method of claim 2 wherein preparing a background mask with elements, corresponding to data points scanned from the molecular array, indicating whether or not corresponding data points are background data points further includes:

allocating a number of background-mask elements;

storing a background-data-point indication in each of the number of elements; and

for each feature of the molecular array,

for each data point within a region of interest for the feature,

storing a non-background-data-point indication in a corresponding background-mask element.

4. The method of claim 2 wherein filtering the selected data points to remove data points that lie within regions of interest of features of the molecular array near the feature further includes:

identifying as non-background data points those selected data points for which corresponding background-mask elements contain non-background-data-point indications.

5. The method of claim 4 wherein identifying as non-background data points those data points for which corresponding background-mask elements contain non-background-data-point indications further includes:

for each selected data point in the set of potential background data points,

identifying the data point as a non-background data point when the corresponding background mask element contains a non-background-data-point indication.

6. The method of claim 2 wherein background-mask elements are bits.

7. The method of claim 2 wherein background-mask elements are memory-storage units suitable for storing Boolean values.

8. The method of claim 1 wherein specifying a corresponding background region further includes specifying a radius for a background-region annulus and a feature having a region of interest surrounded by the background-region annulus.

9. The method of claim 1 wherein specifying a corresponding background region further includes specifying a size and a shape for the background region and a feature surrounded by the background region.

10. The method of claim 1 wherein specifying a corresponding background region further includes specifying a size, a shape, and a position for the background region.

11. The method of claim 1 wherein specifying a corresponding background region further includes specifying a size for the background region.

12. The method of claim 1 wherein specifying a corresponding background region further includes specifying a shape and a position for the background region.

13. The method of claim 1 wherein data points are pixel intensity values associated with pixels in the scanned image of a molecular array.

14. A method of analyzing a feature from a representation of scanned intensity values of a molecular array comprising data points and containing discrete features, said method comprising:

accessing a representation of a background-corrected data set produced by subtracting from feature intensities background signal intensities using the method of claim 1, that has been maintained for subsequent analysis by one of;

storing the representation of the background-corrected data set in a computer-readable medium, and transferring the representation of the background-corrected data set to an intercommunicating entity via electronic signals; and

analyzing signal intensity of the feature based at least in part on the background signal intensity determined for the particular feature from the filtered, selected data points.

15. The method of claim 1, wherein said data points that lie within said regions of interest of features of the molecular array near the feature are removed regardless of signal intensity levels of said data points that lie within said regions of interest.

16. The method of claim 1, wherein all data points that lie with said regions of interest of features of the molecular array near the feature are removed.

18. The method of claim 1, wherein background signal intensities calculated are used to:

evaluate operation of a molecular array scanner;

calibrate a molecular array scanner;

evaluate the quality of a molecular array; and

/or

evaluate the reproducibility of a molecular-array-based experiment.

19. A method according to claim 1 wherein the background signal intensity is communicated to a remote location.

17. A computer readable medium carrying one or more sequences of instructions for determining a background signal intensity for a particular feature from a representation of scanned intensity values of a molecular array comprising data points and containing discrete features at specified locations, each feature having a region of interest containing a number of data points, wherein execution of one or more sequences of instructions by one or more processors causes the one or more processors to perform the steps of:

selecting as background data points those data points within a specified background region;

filtering the selected data points to remove all data points that lie within specified locations of features and at least one region of interest of features in the region of specified size for the background region; and

determining the background signal intensity for the particular feature from the filtered, selected data points, wherein background-subtraction of data points from the particular feature using the determined background signal intensity facilitates calculation of accurate and reproducible log ratios between at least one of multi-channel and inter-array data.

20. A molecular-array-data processing system comprising:

a computer processor;

a communications medium by which molecular array data points are received by the molecular-array-data processing system;

one or more memory components that store molecular array data points; and

a program, stored in the one or more memory components and executed by the computer processor to perform the following steps receiving specified location of features;

specifying a size for a background region corresponding to the feature;

selecting as background data points those data points within the specified background region;

filtering the selected data points to remove all data points that lie within specified locations of features and at least one region of interest of features in the region of specified size for the background region; and

determining the background signal intensity for the particular feature from the filtered, selected data points, wherein background-subtraction of data points from the particular feature using the detennined background signal intensity facilitates calculation of accurate and reproducible log ratios between at least one of multi-channel and inter-array data.

21. The molecular-array-data processing system of claim 20 wherein the program prepares a background mask with elements, corresponding to data points scanned from the molecular array, containing indications of whether or not corresponding data points are background data points and identifies as non-background data points those selected data points for which corresponding background-mask elements contain non-background-data-point indications.

22. A method for determining a background signal intensity for a particular feature from a representation of scanned intensity values of a molecular array comprising data points and containing discrete features at specified locations, each feature having a region of interest containing a number of data points represented by pixel intensity values associated with pixels in a scanned image of the molecular array, the method comprising:

specifying size for a background region corresponding to the feature;

selecting as background pixels those pixels within the specified background region;

filtering the selected pixels to remove all pixels that lie within specified location of features and at least one region of interest of features in the region of specified size for the background region; and

determining the background signal intensity for the particular feature from the filtered, selected pixels, wherein background-subtraction of pixels from the particular feature using the determined background signal intensity facilitates calculation of accurate and reproducible log ratios between at least one of multi-channel and inter-array data.

23. The method of claim 22, wherein the size of the background region specified includes regions of interest of more than one feature.