Multiplexed molecular analysis apparatus and method

• US 6,083,763 A
• Filed: 12/31/1997
• Issued: 07/04/2000
• Est. Priority Date: 12/31/1996
• Status: Expired due to Term

• Current Assignee: High Throughput Genomics, Inc.
• Original Assignee: Genometrix Inc.
• Inventors: Balch, William J.
• Primary Examiner(s): Chin, Christopher L.
• Application Number:
  US09/002,170
• Time in Patent Office: 916 Days
• Field of Search: 422/62, 422/63, 422/67, 422/68.1, 422/81, 422/105, 422/112, 435/286.1, 435/286.6, 435/286.5, 435/6, 436/518, 436/524, 436/525, 436/527, 436/531, 436/43, 436/50
• US Class Current: 506/9
• 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 2219/00315 Microtiter plates
• CPC Class Codes: B01J 2219/00317 Microwell devices, i.e. having large numbers of wells
• CPC Class Codes: B01J 2219/00369 in multiple or parallel arrangements
• CPC Class Codes: B01J 2219/00378 Piezoelectric or ink jet dispensers
• CPC Class Codes: B01J 2219/00511 Walls of reactor vessels
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/00596 Solid-phase processes
• CPC Class Codes: B01J 2219/00605 the compounds being directly bound or immobilised to solid supports
• 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 2219/00621 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 2219/00659 Two-dimensional arrays
• CPC Class Codes: B01J 2219/00707 separated from the reactor apparatus
• CPC Class Codes: B01J 2219/00722 Nucleotides
• CPC Class Codes: B01L 2300/0893 having a very large number of wells, microfabricated wells
• CPC Class Codes: B01L 3/0262 using touch-off at substrate or container
• CPC Class Codes: B01L 3/0268 : using pulse dispensing or spraying, eg. inkjet type, piezo actuated ejection of droplets from capillaries
• CPC Class Codes: B01L 3/5085 : for multiple samples, e.g. microtitration plates
• 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 2035/00237 : Handling microquantities of analyte, e.g. microvalves, capillary networks
• CPC Class Codes: G01N 2035/1034 : Transferring microquantities of liquid
• CPC Class Codes: G01N 2035/1039 : Micropipettes, e.g. microcapillary tubes
• CPC Class Codes: G01N 2035/1041 : Ink-jet like dispensers
• CPC Class Codes: G01N 33/54366 : Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 35/0099 : comprising robots or similar manipulators robots per se B25J
• CPC Class Codes: G01N 35/028 : having reaction cells in the form of microtitration plates
• CPC Class Codes: G01N 35/1065 : Multiple transfer devices
• CPC Class Codes: G01N 35/1074 : arranged in a two-dimensional array
• CPC Class Codes: Y10S 435/808 : Optical sensing apparatus
• CPC Class Codes: Y10S 436/80 : Fluorescent dyes, e.g. rhodamine
• CPC Class Codes: Y10S 436/804 : Radioisotope, e.g. radioimmunoassay
• CPC Class Codes: Y10S 436/805 : Optical property
• CPC Class Codes: Y10S 436/809 : Multifield plates or multicontainer arrays
• CPC Class Codes: Y10T 436/11 : Automated chemical analysis
• CPC Class Codes: Y10T 436/115831 : Condition or time responsive

Abstract

A method and apparatus for analyzing molecular structures within a sample substance using an array having a plurality of test sites upon which the sample substance is applied. The invention is also directed to a method and apparatus for constructing molecular arrays having a plurality of test sites. The invention allows for definitive high throughput analysis of multiple analytes in complex mixtures of sample substances. A combinatorial analysis process is described that results in the creation of an array of integrated chemical devices. These devices operate in parallel, each unit providing specific sets of data that, when taken as a whole, give a complete answer for a defined experiment. This approach is uniquely capable of rapidly providing a high density of information from limited amounts of sample in a cost-effective manner.

Claims

1. A device for preparing a reaction substrate having biosites for conducting multiplexed microassays to determine binding between a target analyte and a capture probe, including:

a plurality of capillary tubes, each tube having a proximal end and a distal end;

an attachment site for holding said capillary tubes at a point spaced from the distal ends of such capillary tubes;

an array template for slidably holding each capillary tube near its distal end, and for allowing the distal end of each capillary tube to move with respect to the attachment site;

at least one manifold for positioning the proximal end of each capillary tube within a corresponding supply chamber, wherein each supply chamber is capable of supplying a liquid reagent to at least one corresponding capillary tube; and

a positioning device for precisely positioning the array template and said capillary tubes with respect to said reaction substrate and depositing liquid reagents from said distal end of said capillary tubes onto said reaction substrate as biosites.

2. The device of claim 1, wherein said plurality of capillary tubes comprises about 2 to about 10,000 tubes, having a center to center spacing of about 80 μ

m to about 5 mm.

3. The device of claim 1, wherein each supply chamber supplies only one capillary tube.

4. The device of claim 1, wherein a supply chamber supplies a plurality of capillary tubes.

5. The device of claim 1, wherein said capillary tubes comprise stainless steel, plastic, rubber, glass, or fused silica coated with polyimide.

6. The device of claim 1, wherein said device includes a plurality of array templates.

7. The device of claim 1, further comprising a positioning device for precisely positioning said reaction substrate.

8. The device of claim 1, wherein said capillaries have an inside diameter of about 10 to about 200 μ

m, and an outside diameter of about 80 to about 500 μ

m.

9. The device of claim 1, wherein said array template comprises an array of sleeves, each sleeve having an inside diameter sufficient to permit a capillary to slide through, and a length sufficient to allow a precise pattern to be maintained while depositing fluids onto said reaction substrate.

10. The device of claim 1, wherein said array template comprises a plurality of holes formed in a rigid material.

11. The device of claim 1, wherein said array template comprises a rigidly formed or held mesh.

12. The device of claim 1, further comprising a housing for containing said supply chambers.

13. The device of claim 12, wherein said housing is capable of maintaining an inert atmosphere.

14. The device of claim 12, wherein said housing is capable of maintaining an elevated or reduced temperature.

15. The device of claim 12, wherein said housing may be pressurized to a predetermined pressure.

16. The device of claim 15, wherein said pressure is modulated to control the flow of liquid reagents through said capillary tubes.

17. The device of claim 1, wherein said supply chambers are positioned higher than the distal end of said capillary tubes to provide a pressure head.

18. The device of claim 1, wherein deposit of liquid reagent is controlled by a means for electrophoresis.

19. The device of claim 1, wherein deposit of liquid reagent is controlled by a means for electro-osmosis.

20. The device of claim 1, wherein said device includes multiple sets of supply chambers, each for supplying liquid reagent to a subset of said capillary tubes.

21. The device of claim 1, wherein said capillary tubes are free to flex between said attachment site and said array template.

22. The device of claim 1, wherein the reaction substrate is pre-etched to define a pattern of reactive areas matching the geometry of the deposited biosites.

23. The device of claim 1, wherein said biosites are deposited substantially simultaneously.

24. The device of claim 1, wherein each biosite is fluidically isolated from each other biosite.

25. The device of claim 1, wherein the liquid reagent comprises at least one capture probe.

26. An apparatus for preparing a reaction substrate for use as an assay device, including:

an attachment site for holding a plurality of flexible capillary tubes at a point spaced from the distal ends of such capillary tubes;

a structure for positioning the proximal end of each capillary tube within a corresponding supply chamber, each supply chamber capable of supplying a liquid reagent to at least one corresponding capillary tube;

a print head for holding said plurality of capillary tubes near their distal ends and for precisely positioning the distal ends of said capillary tubes with respect to said reaction substrate to deposit liquid reagents onto said reaction substrate as biosites.

27. The apparatus of claim 26, where the print head includes an ink-jet deposition device.