System and method for visualization and digital analysis of protein and other macromolecule microarrays

• US 20040072274A1
• Filed: 05/08/2003
• Published: 04/15/2004
• Est. Priority Date: 05/09/2002
• Status: Abandoned Application

• Current Assignee: Miragene Incorporated
• Original Assignee: Miragene Incorporated
• Inventors: Lebrun, Stewart J.
• Application Number:
  US10/431,686
• Publication Number:
  US 20040072274A1
• Time in Patent Office: Days
• US Class Current: 435/21
• CPC Class Codes: G01N 21/253 for batch operation, i.e. multisample apparatus analytical automats G01N35/00

Abstract

A system and method are disclosed for the rapid, reproducible and inexpensive imaging and digital analysis of molecular interactions between ligands and proteins and/or nucleic acids immobilized on an addressable two-dimensional microarray.

Claims

1. A method for visualization and digital analysis of sample interactions with a biological array, comprising:

allowing the sample to interact with the array, which comprises biological molecules immobilized on a solid substrate in a two-dimensional and addressable pattern;

contacting the array with a secondary detector molecule comprising an enzyme;

incubating the array with a developing agent comprising a substrate of the enzyme, such that the enzyme catalyzes a reaction wherein the substrate is converted to a detectable product;

digitizing an array image created by the detectable product by scanning the array on a digital scanner; and

analyzing the digital image.

2. The method of claim 1, wherein the detectable product is selected from the group consisting of a colorometric precipitate, a colorometric enzyme-analyte, a colorometric dye-analyte, a colorometric intermediate-analyte, and a radioactive-analyte.

3. The method of claim 1, wherein prior to contacting the array with the secondary detector molecule, the array is washed to remove unreacted sample.

4. The method of claim 1, wherein prior to digitizing the array image, the array is washed to terminate the reaction.

5. The method of claim 1, wherein prior to digitizing the array image, the array is dried.

6. The method of claim 1, wherein the enzyme is Alkaline Phosphatase.

7. The method of claim 1, wherein the substrate is BCIP/NBT.

8. The method of claim 1, wherein the scanner is selected from the group consisting of Epson PERFECTION 1650, Canon CANOSCAN N1240U, and Hewlett-Packard SCANJET 5300C.

9. The method of claim 1, wherein prior to scanning the array, the array is placed together with one or more additional arrays in a template having from about 1 to 20 array slots.

10. The method of claim 1, wherein the array is labeled with a barcode.

11. The method of claim 1, further comprising correcting for user error and slide variations using an imaging program.

12. The method of claim 1, further comprising quantifying markers to construct a standard curve, such that visual intensity can be converted into molecular mass.

13. The method of claim 12, wherein a clinical index is calculated by dividing the molecular mass by the dilution factor.

14. A method for analysis of sample interactions with a protein microarray, comprising:

allowing the sample to interact with the microarray, comprising a plurality of proteins immobilized on a solid substrate in a two-dimensional and addressable pattern, the solid substrate comprising a barcode for sample identification and a PVDF membrane adhered to a rigid support;

contacting the microarray with a secondary detector molecule comprising a selective binding moiety and an enzyme conjugated thereto;

washing the microarray to remove unbound secondary detector molecules;

incubating the array with a developing agent comprising a substrate of the enzyme, such that the enzyme catalyzes a reaction wherein the substrate is converted to a detectable product;

washing the microarray to terminate the reaction and remove unreacted developing agent;

scanning the microarray using a digital scanner to create a digital image of the microarray; and

analyzing the digital image which corresponds to the sample interactions with the protein microarray.

15. A system for analysis of autoimmune diseases in humans, comprising:

a microarray, comprising a plurality of autoimmune markers immobilized on a solid substrate in a two-dimensional and addressable pattern, the solid substrate comprising a PVDF membrane adhered to a rigid support;

a first reagent comprising anti-human IgG conjugated to an enzyme;

a second reagent comprising a developing agent comprising a substrate of the enzyme, wherein the first and second reagents react when combined to yield a colorometric change; and

a flatbed digital scanner.

16. A system for analysis of sample interactions with a biological array, comprising:

an array, comprising a plurality of biological molecules immobilized on substrate in a two-dimensional and addressable pattern, the substrate comprising a rigid support which has an opaque surface or has been modified to have an opaque surface, said surface being adapted to bind the plurality of biological molecules;

a flatbed scanner adapted to produce a digital image of the array; and

a software program adapted to quantify and analyze the digital image.