Methods for concurrently processing multiple biological chip assays

• US 6,720,149 B1
• Filed: 05/28/2002
• Issued: 04/13/2004
• Est. Priority Date: 06/07/1995
• Status: Expired due to Fees

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Trulson, Mark, Norviel, Vernon A., Fodor, Stephen P. A., Rava, Richard P.
• Primary Examiner(s): Horlick, Kenneth R.
• Assistant Examiner(s): WILDER, CYNTHIA B
• Application Number:
  US10/157,252
• Time in Patent Office: 686 Days
• Field of Search: 435/6, 435/91.2, 435/288.4, 435/DIG.49, 435/24.33, 435/5, 435/7.1, 435/7.9, 536/23.1, 536/24.3, 536/24.37, 536/33
• US Class Current: 506/4
• CPC Class Codes: B01J 19/0046 Sequential or parallel reactions, e.g. for the synthesis of polypeptides or polynucleotides; Apparatus and devices for combinatorial chemistry or for making molecular arrays synthesis methods per se C40B50/00
• CPC Class Codes: B01J 2219/00286 Reactor vessels with top and bottom openings
• CPC Class Codes: B01J 2219/00315 Microtiter plates
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00576 fluorophore
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• CPC Class Codes: B01J 2219/00608 DNA chips
• CPC Class Codes: B01J 2219/0061 The surface being organic
• CPC Class Codes: B01J 2219/00612 the surface being inorganic
• CPC Class Codes: B01J 2219/00617 by chemical means
• CPC Class Codes: B01J 2219/00619 using hydrophilic or hydrophobic regions
• CPC Class Codes: B01J 2219/00621 by physical means, e.g. trenches, raised areas
• CPC Class Codes: B01J 2219/00626 Covalent
• CPC Class Codes: B01J 2219/00637 by coating it with another layer
• CPC Class Codes: B01J 2219/00639 the compounds being trapped in or bound to a porous medium
• CPC Class Codes: B01J 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00662 Two-dimensional arrays within two-dimensional arrays
• CPC Class Codes: B01J 2219/00702 : Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00707 : separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00722 : Nucleotides
• CPC Class Codes: B01L 2300/041 : Connecting closures to device or container
• CPC Class Codes: B01L 2300/044 : pierceable, e.g. films, membranes
• CPC Class Codes: B01L 2300/046 : Function or devices integrated in the closure
• CPC Class Codes: B01L 2300/0636 : Integrated biosensor, microarrays
• CPC Class Codes: B01L 2300/0819 : Microarrays; Biochips
• CPC Class Codes: B01L 2300/0829 : Multi-well plates; Microtitration plates
• CPC Class Codes: B01L 2400/0683 : mechanically breaking a wall or membrane within a channel or chamber
• CPC Class Codes: B01L 3/5027 : by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip B01L3/5023 takes precedence; micromixers B01F33/30; microreactors for synthesis B01J19/0093; microcapillary devices in general B81B1/00
• CPC Class Codes: B01L 3/5085 : for multiple samples, e.g. microtitration plates
• CPC Class Codes: B01L 3/50853 : with covers or lids
• CPC Class Codes: C12Q 1/6837 : using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G01N 2035/00158 : Elements containing microarrays, i.e. "biochip"
• CPC Class Codes: G01N 21/64 : Fluorescence; Phosphorescence
• CPC Class Codes: G01N 35/028 : having reaction cells in the form of microtitration plates

Abstract

Methods for concurrently processing multiple biological chip assays by providing a biological chip plate comprising a plurality of test wells, each test well having a biological chip having a molecular probe array; introducing samples into the test wells; subjecting the biological chip plate to manipulation by a fluid handling device that automatically performs steps to carry out reactions between target molecules in the samples and probes; and subjecting the biological chip plate to a biological chip plate reader that interrogates the probe arrays to detect any reactions between target molecules and probes.

Claims

1. A method for concurrently processing multiple biological chip assays comprising the steps of:

(a) providing a biological chip plate comprising a plurality of probe arrays and, surrounding the probe arrays, material resistant to the flow of liquid, thereby forming a plurality of test wells, each test well defining a space for the introduction of a sample;

(b) introducing into each test well test samples from a plurality ofdifferent patients, wherein each test sample contains target molecules;

(c) manipulating the biological chip plate with a fluid handling device that automatically performs steps to carry out reactions between target molecules in the test samples and probes in a plurality of the test wells, wherein the fluid handling device controls temperature, sample handling, substrate handling and washing of the test wells; and

(d) interrogating the probe arrays of the biological chip plate with a biological chip plate reader to detect reactions between target molecules and probes in a plurality of the test wells to generate assay results.

2. The method of claim 1 further comprising the step of processing the results with a computer.

3. The method of claim 2 wherein the results are processed into an image with the intensity in each region of the image varying according to the binding affinity between targets and probes.

4. The method of claim 1 wherein the probes are DNA or RNA molecules.

5. The method of claim 4 wherein

(a) the reactions are hybridization of target moleculcs to probes; and

(b) the results provide information regarding the nucleotide sequence of the target molecule.

6. The method of claim 5 wherein the assay is the detection or identification of a pathogenic organism.

7. The method of claim 6 wherein the pathogenic organism is HIV.

8. The method of claim 5 wherein the assay is the detection or identification of a human nucleic acid.

9. The method of claim 8 wherein the assay is the detection of a human gene variant.

10. The method of claim 9 wherein the human gene variant indicates the existence of, or predisposition to, cystic fibrosis, diabetes, muscular dystrophy or cancer.

11. The method of claim 1 further comprising controlling the temperature of the samples in the test wells and adding or removing fluids from the lest wells at predetermined times.

12. The method, of claim 1 wherein the target molecule is tagged with a fluorescent marker, a chemiluminescent marker, a light scattering marker or a radioactive marker.

13. The method of claim 12 wherein the marker is a fluorescent marker selected from the group consisting of fluorcscein, rhodamine, or cyaninc.

14. The method of claim 12 wherein the biological chip plate reader comprises:

(a) a confocal detection device having a monochromatic or polychroniatic light source;

(b) optics for dirccting ail excitation light from the light source at the substrate;

(c) a temperature controller for conrolling temperature of the substrate during a reaction; and

(d) a detector for detecting fluorescence emitted by the targets in response to the excitation light.

15. The method of claim 1 wherein the manipulating step comprises robotically controlling pipets for adding or removing fluids from the test wells at predetermined times.

16. The method of claim 14 wherein the detector comprises a photomultiplier tube.

17. The method of claim 14 wherein the interrogating step comprises focusing the excitation light to a point on the plate and determining the region from which the fluorescence originated.

18. The method of claim 14 wherein the target is tagged with a fluorescent marker and the biological chip plate reader comprises a CCD array.

19. The method of claim 1 wherein the probe arrays each have at least about 2500 features.

20. The method of claim 1 wherein the probe arrays each have at least about 50,000 features.

21. The method of claim 1 wherein said chip plate is made by the steps of mating a wafer comprising a plurality of probe arrays to an array of open ended wells.

23. The method of claim 1 wherein the interrogating step comprises determining the on-or off-rates of binding between target molecules in a sample and probes.

24. The method of claim 1 wherein the probe arrays are produced by light-directed probe synthesis.

25. The method as recited in claim 24 wherein said arrays of probes are made by repeating the steps of:

(a) selectively directing light at said surface to remove photoremovable protecting groups; and

(b) selectively coupling monomers to said surface.

26. The method of claim 24 wherein the chip plate comprises a contiguous substrate that comprises a contiguous surface and the plurality of probe arrays are attached to the contiguous surface.

27. The method of claim 1 wherein each probe array is about 0.25 cm².

28. The method of claim 1 wherein each probe array is about 1 cm².

29. The method of claim 1 wherein the probe arrays comprise leaturcs of about 100 microns on a side.

30. The method of claim 1 wherein the probe arrays comprise features of about 20 microns on a side.

31. The method of claim 1 wherein the chip plate comprises a contiguous substrate that comprises a contiguous surface and the plurality of probe arrays are attached to the contiguous surface.

22. A method for concurrently processing multiple biological chip assays, comprising the steps of:

(a) providing a biological chip plate comprising a plurality of probe arrays and, surrounding the probe arrays, material resistant to the flow of liquid, thereby forming a plurality of test wells, each test well defining a space for the introduction of a sample;

(b) introducing into each test well test samples from a plurality of diffcrcnt patients, wherein each test sample contains target molecules;

(c) manipulating the biological chip plate with a fluid handling device that automatically performs steps to carry out reactions between target molecules in the test samples and probes in a plurality of the test wells, wherein the fluid handling device controls temperature, sample handling, substrate handling and washing of the test walls; and

(d) interrogating the probe arrays of the biological chip plate with a biological chip plate reader to detect reactions between target molecules and probes in a plurality of the test wells to generate assay results, wherein the interrogating step comprises the steps of;

(i) directing excitation light through a bottom surface of said plate; and

(ii) detecting where targets bound on said probe arrays.