Combinatorial strategies for polymer synthesis

• US 6,864,101 B1
• Filed: 05/26/2000
• Issued: 03/08/2005
• Est. Priority Date: 11/22/1991
• Status: Expired due to Fees

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: Modlin, Douglas N., Winkler, James L., Ross, Debra A., Aldwin, Lois, Fodor, Stephen P. A., Buchko, Christopher J.
• Primary Examiner(s): Ludlow, Jan M.
• Application Number:
  US09/579,949
• Time in Patent Office: 1,747 Days
• Field of Search: 436/518, 436/809, 436/180, 436/89, 436/94, 738/63, 738/64, 738/648.1, 422/100, 141/31
• US Class Current: 506/32
• 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 2219/00364 Pipettes
• CPC Class Codes: B01J 2219/00367 capillary
• 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 2219/00389 Feeding through valves
• CPC Class Codes: B01J 2219/00416 Vacuum
• CPC Class Codes: B01J 2219/0043 for direct application of reagents, e.g. through openings in a shutter
• CPC Class Codes: B01J 2219/00432 Photolithographic masks
• CPC Class Codes: B01J 2219/00475 Sheets
• CPC Class Codes: B01J 2219/005 Beads
• CPC Class Codes: B01J 2219/00527 Sheets
• CPC Class Codes: B01J 2219/00529 DNA chips
• CPC Class Codes: B01J 2219/00531 essentially square
• CPC Class Codes: B01J 2219/00536 in the shape of disks
• CPC Class Codes: B01J 2219/00585 Parallel processes
• CPC Class Codes: B01J 2219/0059 Sequential 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 2219/00608 DNA chips
• 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 2219/00659 : Two-dimensional arrays
• CPC Class Codes: B01J 2219/00686 : Automatic
• CPC Class Codes: B01J 2219/00702 : Processes involving means for analysing and characterising the products
• CPC Class Codes: B01J 2219/00711 : Light-directed synthesis
• CPC Class Codes: B01J 2219/00722 : Nucleotides
• CPC Class Codes: B01J 2219/00725 : Peptides
• CPC Class Codes: B82Y 30/00 : Nanotechnology for materials or surface science, e.g. nanocomposites
• CPC Class Codes: C07B 2200/11 : Compounds covalently bound to a solid support
• CPC Class Codes: C07H 21/00 : Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07K 1/045 : using devices to improve synthesis, e.g. reactors, special vessels
• CPC Class Codes: C07K 1/047 : Simultaneous synthesis of different peptide species; Peptide libraries
• CPC Class Codes: C40B 40/06 : Libraries containing nucleotides or polynucleotides, or derivatives thereof
• CPC Class Codes: C40B 40/10 : Libraries containing peptides or polypeptides, or derivatives thereof
• CPC Class Codes: C40B 60/14 : for creating libraries
• CPC Class Codes: G01N 35/1072 : with provision for selective pipetting of individual channels
• CPC Class Codes: Y10S 436/809 : Multifield plates or multicontainer arrays
• CPC Class Codes: Y10T 436/143333 : Saccharide [e.g., DNA, etc.]
• CPC Class Codes: Y10T 436/2575 : Volumetric liquid transfer

Abstract

A method for producing polymer arrays by spacing a dispenser a distance from a surface of a support, dispensing a volume containing a monomer in a single coupling step of less than 5 nl to occupy a localized area of less than 1 cm² of the surface of the support, allowing the monomer to bind directly or indirectly to the support and repeating the steps to produce an array of at least 100 polymer ligands at a density of 1000 per cm² or greater.

Claims

1. An automated method of forming an array of ligands on a support having localized areas comprising

(a) locating a dispenser to dispense a solution a distance away from a surface of the support;

(b) dispensing a first monomer in a volume of solution in a single coupling step of less than 5 nl onto the surface of the support to occupy a localized area smaller than 1 cm²;

(c) allowing the first monomer to attach directly or indirectly to the surface of the support at the localized area;

(d) repeatedly dispensing an additional group of added monomers in a volume of solution in a single coupling step of less than 5 nl at a same or different localized area in a manner to couple with a compound at one or more localized areas until an array of at least 100 different ligands, each at an individual localized area is formed and wherein density of localized areas on the support is at least about 1000 localized areas per cm² of surface of the support.

2. The method of claim 1 wherein the monomer is dissolved in solution.

3. The method of claim 1 wherein the monomer is in the form of a pellet.

4. The method of claim 1 wherein the support further comprises a cover plate.

5. The method of claim 1 wherein the step of dispensing includes a dispenser positioned between about 5 microns and about 50 microns away from the support.

6. The method of claim 1 wherein the step of dispensing includes a dispenser positioned about 10 microns away from the support.

7. The method of claim 1 wherein the volume fits within a region having a diameter of less than about 300 microns.

8. The method of claim 1 wherein the monomer comprises a nucleotide or an amino acid.

9. The method of claim 1 wherein the ligand comprises a nucleic acid, oligonucleotide, polynucleotide, peptide, or polypeptide.

10. The method of claim 1 wherein the ligand comprises at least 2 monomers.

11. The method of claim 1 wherein the ligand comprises greater than 100 monomers.

12. The method of claim 1 wherein the ligand comprises 2, 3, 4, 5, 6, 10, 15, 20, 30, 40, 50, 75, or 100 monomers.

13. The method of claim 1 wherein the support is selected from the group consisting of substantially flat substrates, substrates having raised or depressed regions, beads, gels, sheets, particles, strands, precipitates, spheres, containers, capillaries, pads, slices, films, plates, and slides.

14. The method of claim 1 wherein the support comprises a gel.

15. The method of claim 1 wherein the support comprises biological materials, nonbiological materials, organic materials or inorganic materials.

16. The method of claim 1 wherein the support is a disc, square, or circle.

17. The method of claim 1 wherein the one or more localized areas are smaller than 1 mm².

18. The method of claim 1 wherein the one or more localized areas are smaller than 0.5 mm².

19. The method of claim 1 wherein the one or more localized areas are smaller than 10,000 μ

m².

20. The method of claim 1 wherein the one or more localized areas are smaller than 100 μ

m².

21. The method of claim 1 wherein an array of at least 1000 different ligands at different localized areas is formed.

22. The method of claim 1 wherein an array of at least 10,000 different ligands at different localized areas is formed.

23. The method of claim 1 wherein an array of at least 100,000 different ligands at different localized areas is formed.

24. The method of claim 1 wherein an array of at least 1,000,000 different ligands at different localized areas is formed.

25. The method of claim 1, wherein an array of at least 1000 different polymers occupying localized areas within 1 cm² of the surface of the support.

26. The method of claim 1, wherein the support comprises glass, derivatized glass, pyrex, quartz, a polymeric material, polystyrene, polycarbonate, silicon or a gel.

27. The method of claim 2, wherein the solution comprises an aqueous solution.

28. The method of claim 1 wherein the step of dispensing includes a dispenser comprising a plurality of dispensing units, wherein the plurality of dispensing units is in fluid communication with a solution comprising a monomer and wherein steps (a) and (c) comprise dispensing a volume of solution in a single coupling step of less than 5 nl from one or more of the plurality of dispensing units.

29. The method of claim 1, wherein the support bears at least two reference points for positioning the dispenser over at least one of said localized areas for dispensing of the volume of solution.

30. The method of claim 29, wherein the reference points comprise global reference points for positioning the dispenser over a local region of the surface of the support, and local reference points within the local region for positioning the dispenser over a localized area within the local region.

31. The method of claim 29, wherein the dispenser further comprises a camera for identifying the reference points.

32. The method of claim 29 further comprising the step of sensing changes in capacitance to identify the reference points.

33. The method of claim 29 further comprising the step of sensing changes in light intensity to identify the reference points.

34. The method of claim 29 further comprising the step of sensing changes in resistivity to identify the reference points.

35. The method of claim 29 further comprising the step of sensing changes in optical properties to identify the reference points.

36. The method of claim 29 further comprising the step of sensing changes in magnetic properties to identify the reference points.

37. The method of claim 28 wherein the plurality of dispensing units comprises a manifold of delivery lines.

38. The method of claim 28 wherein the plurality of dispensing units comprises an array of pipettes.

39. The method of claim 28 wherein the plurality of dispensing units comprises a series of tubes.

40. The method of claim 28 wherein the plurality of dispensing units includes control valves.

41. The method of claim 1 wherein the monomer is bound indirectly to the surface of the support via a linker molecule.

42. The method of claim 1 wherein the step of dispensing includes a dispenser that is moved relative to the support.

43. The method of claim 1 wherein the support is moved relative to a dispenser.

44. The method of claim 1 wherein the one or more localized areas are spaced less than about 3 mm apart.

45. The method of claim 1 wherein the one or more localized areas are spaced less than between about 5 microns and 100 microns apart.

46. The method of claim 1 wherein the one or more localized areas has an angular relation between each localized area of about 1 degree.

47. The method of claim 1 wherein the one or more localized areas has an angular relation between each localized area of about 0.1 degree.

48. The method of claim 1 wherein the support comprises at least about 100 localized areas.

49. The method of claim 1 wherein the support comprises at least about 1000 localized areas.

50. The method of claim 1 wherein the support comprises at least about 10,000 localized areas.

51. The method of claim 1 wherein the support comprises at least about 10,000 localized areas per cm² of surface of substrate.

52. The method of claim 1 wherein the support comprises a strand including one or more of glass, derivatized glass, quartz, or a polymeric material.

53. The method of claim 1 wherein the step of dispensing includes a dispenser comprising a dispenser tip and a sheath encircling the dispenser tip and rigidly extending a fixed distance beyond the dispenser tip.

54. The method of claim 1 wherein the surface of the support comprises a hydrophilic substance.

55. The method of claim 1 wherein the surface of the support comprises a hydrophobic substance.

56. The method of claim 1 wherein the surface of the support comprises a hydrophilic substance and a hydrophobic substance.

57. The method of claim 1 wherein the surface of the support comprises a hydrophilic group.

58. The method of claim 1 wherein the surface of the support comprises a hydrophobic group.

59. The method of claim 1 wherein the surface of the support comprises a hydrophilic group and a hydrophobic group.

60. The method of claim 1 wherein the surface of the support comprises a photoresist.

61. The method of claim 1 wherein the surface of the support is cleaned prior to the step of dispensing the volume of solution.

62. The method of claim 1 wherein the step of dispensing includes a dispenser comprising a pipette.

63. The method of claim 1 wherein the step of dispensing includes a dispenser comprising a capillary tube.

64. The method of claim 1 wherein the step of dispensing includes a dispenser comprising an electrophoretic pump.

65. The method of claim 1 wherein the step of dispensing includes a dispenser comprising an osmotic pump.

66. The method of claim 1 wherein the step of dispensing includes a dispenser comprising a cell sorter.

67. A method of forming an array of ligands on a support having localized areas comprising

(a) locating a dispenser comprising a plurality of dispensing units a distance away from a surface of the support, wherein the plurality of dispensing units is in fluid communication with a solution comprising a monomer;

(b) dispensing a volume of solution in a single coupling step of less than 5 nl from the dispenser, with the volume contacting the surface at a localized area smaller than 1 cm²;

(c) allowing the monomer to attach directly or indirectly to the surface of the support at the localized area;

(d) repeating steps a through c to attach a same or different monomer at a same or different localized area until an array of at least 100 different ligands, each at an individual localized area is formed and wherein density of localized areas on the support is at least about 1000 localized areas per cm² of surface of the support.

68. The method of claim 67 wherein an array of at least 1000 different ligands at different localized areas is formed.

69. The method of claim 67 wherein an array of at least 10,000 different ligands at different localized areas is formed.

70. The method of claim 67 wherein an array of at least 100,000 different ligands at different localized areas is formed.

71. The method of claim 67 wherein an array of at least 1,000,000 different ligands at different localized areas is formed.

72. The method of claim 67 wherein the monomer is an amino acid or a nucleotide.

73. The method of claim 67 wherein the ligand is a polypeptide or a nucleic acid.