Fluorometric detection using visible light

• US 6,512,236 B2
• Filed: 01/02/2001
• Issued: 01/28/2003
• Est. Priority Date: 03/07/1997
• Status: Expired due to Term

• Current Assignee: Clare Chemical Research Incorporated
• Original Assignee: Clare Chemical Research Incorporated
• Inventors: Seville, Mark
• Primary Examiner(s): Hannaher, Constantine
• Application Number:
  US09/753,783
• Publication Number:
  US 20020089658A1
• Time in Patent Office: 756 Days
• Field of Search: 250/458.1, 250/459.1
• US Class Current: 250/458.1
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6447 by visual observation
• CPC Class Codes: G01N 21/645 Specially adapted constructive features of fluorimeters

Abstract

Systems, devices and methods are provided for viewing a pattern of fluorophors capable of fluorescing when exposed to visible light, e.g., fluorescently stained DNA, protein or other biological material. The system includes a light source emitting light in the visible spectrum, such as a fluorescent lamp used in domestic lighting, a first optical filter capable of transmitting light from the source at wavelengths capable of exciting the fluorophors and of absorbing light of other wavelengths, and a second optical filter capable of blocking substantially all the light from the source not blocked by the first filter, so that the only light reaching the viewer is light produced by fluorescence of the fluorophors.

Claims

1. A visible light transilluminator system for viewing one or more patterns of fluorescence emitted by fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser colored light producing element that produces light, at least a portion of which is capable of exciting said fluorophors;

b) a first colored optical filter positioned between said light producing element and said fluorophors, wherein said first optical filter is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type; and

c) a second colored optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light from said fluorophors of said emitted type and of preventing transmission of light of said excitation type;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

2. The transilluminator of claim 1 comprising a light detector.

3. The transilluminator of claim 1 wherein said first optical filter is blue or green and said light producing element appears blue to the eye.

4. The transilluminator of claim 3 wherein said second optical filter is yellow.

5. The transilluminator of claim 3 wherein said second optical filter is amber.

6. The transilluminator of claim 3 wherein said second optical filter is red.

7. The transilluminator of claim 1 wherein said first optical filter is blue or green and said light producing element appears green to the eye.

8. The transilluminator of claim 7 wherein said second optical filter is yellow.

9. The transilluminator of claim 7 wherein said second optical filter is amber.

10. The transilluminator of claim 7 wherein said second optical filter is red.

11. The transilluminator of claim 1 wherein said light producing element appears red to the eye.

12. The transilluminator of claim 11 wherein said second optical filter is red.

13. The transilluminator of claim 1 wherein said light producing element produces light with a maximal output over a wavelength range of approximately 380 nm to approximately 440 nm and said first filter is violet-blue.

14. The transilluminator of claim 13 wherein said second optical filter is yellow.

15. The transilluminator of claim 13 wherein said second optical filter is red.

16. The transilluminator of claim 13 wherein said second optical filter is amber.

17. The transilluminator of claim 1 wherein said light producing element produces light with a maximal output over a wavelength range of approximately 440 nm to approximately 470 nm and said first optical filter is blue.

18. The transilluminator of claim 17 wherein said second optical filter is yellow.

19. The transilluminator of claim 17 wherein said second optical filter is red.

20. The transilluminator of claim 17 wherein said second optical filter is amber.

21. The transilluminator of claim 1 wherein said light producing element produces light with maximal output over a wavelength range of approximately 460 nm to approximately 510 nm and said first optical filter is blue or blue-green.

22. The transilluminator of claim 21 wherein said second optical filter is yellow.

23. The transilluminator of claim 21 wherein said second optical filter is red.

24. The transilluminator of claim 21 wherein said second optical filter is amber.

25. The transilluminator of claim 1 wherein said light producing element produces light with maximal output over a wavelength range of approximately 500 nm to approximately 550 nm and said first optical filter is green or blue-green.

26. The transilluminator of claim 25 wherein said second optical filter is yellow.

27. The transilluminator of claim 25 wherein said second optical filter is red.

28. The transilluminator of claim 25 wherein said second optical filter is amber.

29. The transilluminator of claim 1 wherein said light producing element produces light with a maximal output over a wavelength range of approximately 550 nm to approximately 650 nm.

30. The transilluminator of claim 29 wherein said second optical filter is red.

31. The transilluminator of claim 29 wherein said second optical filter is amber.

32. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors or a material containing fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser colored light source comprising a light producing element that produces light, at least a portion of which is capable of exciting said fluorophors, and a first colored optical filter positioned between said light producing element and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type, wherein the non-laser light source produces light substantially free of light in the ultraviolet region; and

b) a second colored optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

33. The photoluminescent imaging system of claim 32 comprising a light detector positioned along an optical illumination axis defined by a notional line connecting said light source and said fluorophors.

34. The photoluminescent imaging system of claim 32 comprising a light detector positioned along a point not on an optical illumination axis defined by a notional line connecting said light source and said fluorophors.

35. The photoluminescent imaging system of claim 32 comprising a light detector in optical communication with said second filter and positioned sufficiently far from said fluorophors to provide spacial resolution of individual emitting fluorophors within said material containing fluorophors.

36. The photoluminescent imaging system of claim 32 wherein said light producing element comprises one or more lamps.

37. The photoluminescent imaging system of claim 32 wherein said light source and said second optical filter are comprised within an integrated horizontal gel electrophoresis unit.

38. The photoluminescent imaging system of claim 32 wherein said light source and said second colored optical filter are comprised within an integrated vertical gel electrophoresis unit.

39. The photoluminescent imaging system of claim 32 wherein said light source and said second colored optical filter are comprised within an integrated thin layer chromatography unit.

40. The photoluminescent imaging system of claim 32 wherein said light source and said second colored optical filter are comprised within an integrated fish tank.

41. The visible light photoluminescent imaging system of claim 32 wherein the colored light source produces light comprising substantially of light capable of exciting said fluorophors.

42. The visible light photoluminescent imaging system of claim 32 wherein the light producing element contains phosphors.

43. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser light source comprising a light producing element that produces light, at least a portion of which is capable of exciting said fluorophors, and a first optical filter positioned between said light producing element and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type, wherein the non-laser light source produces light substantially free of light in the ultraviolet region; and

b) a second optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

44. The transilluminator of claim 43 comprising one or more additional optical filters positioned in optical communication with said second optical filter.

45. The photoluminescent imaging system of claim 43 wherein said light producing element has a total power per square area less than approximately 0.072 W cm⁻

2.

46. The photoluminescent imaging system of claim 43 wherein said light producing element has a luminous flux less than approximately 2670 lumens.

47. The photoluminescent imaging system of claim 43 wherein said first optical filter and said second optical filter have a transmission area ranging from approximately 9 cm² to approximately 1180 cm².

48. The photoluminescent imaging system of claim 43 wherein said second optical filter is a short wavelength cutoff filter in the visible region.

49. The photoluminescent imaging system of claim 43 wherein said first optical filter is a long wavelength cutoff filter in the visible region.

50. The photoluminescent imaging system of claim 43 wherein said first optical filter and said second optical filter are cutoff filters in the visible region.

51. The visible light photoluminescent imaging system of claim 43 wherein the non-laser light source produces less ultraviolet light than a UV transilluminator.

52. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser light source comprising a light producing element that produces light, a portion of which is capable of exciting said fluorophors, and a first optical filter positioned between said light producing element and said fluorophors, wherein said first optical filter is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type;

b) fluorophors having a Stokes shift of less than 100 nm; and

c) a second optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of light of said excitation type;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

53. A method of viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said method comprising:

a) passing visible light substantially free of light in the ultraviolet region from a non-laser colored light source on to said fluorophors or a material containing said fluorophors, whereby said fluorophors emit light of said emitted type, wherein said colored light source comprises a light producing element that produces light, at least a portion of which is said excitation type, and a first colored optical filter positioned between said light producing element and said fluorophors, wherein said first optical filter is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type;

b) passing said emitted light through a second colored optical filter, which is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of light of said excitation type, whereby an image of said patterns of fluorescence is formed; and

c) viewing said image with a human eye and/or other light detector.

54. The method of claim 53 wherein said fluorophors are comprised in fluorescently stained DNA.

55. The method of claim 53 wherein said material containing fluorophors comprises at least one fluorescently labeled protein.

56. The method of claim 53 wherein said material containing fluorophors comprises at least one fluorescent protein.

57. The method of claim 53 wherein said fluorophors are comprised in a gel.

58. The method of claim 53 wherein said fluorophors are comprised in a living organism.

59. The method of claim 53 wherein said fluorophors are viewed in an array of test tubes.

60. The method of claim 53 wherein said fluorophors are comprised in fluorescent fish in a fish tank.

61. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength and polarization different from said excitation type, said system comprising:

a) a non-laser light producing element that produces light, at least a portion of which is light of said excitation type;

b) a first optical polarization filter positioned between said light producing element and said fluorophors, wherein said first filter is capable of transmitting polarized light of said excitation type having a first polarization orientation and capable of preventing transmission of light from the light-producing element having a polarization orientation different than said first polarization orientation; and

c) a second optical polarization filter positioned in optical communication with said fluorophors, wherein said second filter is capable of transmitting polarized light of said emitted type having a second polarization orientation and preventing transmission of light of said excitation type and wherein said second polarization orientation is different than that of said first orientation and is different from the polarization orientation of light from said light producing element scattered by said fluorophors;

said system being constructed and arranged such that patterns of fluorescence, emitted by said fluorophors, of wavelengths different than that of light of said excitation type, are viewable.

62. The photoluminescent imaging system of claim 61 comprising a light detector.

63. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) colored non-laser exciting light substantially free of light in the ultraviolet region, at least a portion of which is of a wavelength in the range of said excitation type;

b) a first colored optical filter positioned in optical communication with said fluorophors or a material containing said fluorophors, through which light of said excitation type is transmitted, wherein said first optical filter is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type; and

c) a second colored optical filter, positioned in optical communication with said fluorophors, wherein said second colored filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of light of said excitation type;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

64. The photoluminescent imaging system of claim 63 comprising a light detector.

65. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength and polarization different from said excitation type, said system comprising:

a) a non-laser light producing element that produces light, at least a portion of which is light of said excitation type;

b) a first optical polarization filter positioned between said light producing element and said fluorophors, wherein said first filter is capable of transmitting polarized light of said excitation type having a first polarization orientation and capable of preventing transmission of light from the light-producing element having a polarization orientation different than said first polarization orientation;

c) a second optical polarization filter positioned in optical communication with said fluorophors, wherein said second filter is capable of transmitting polarized light of said emitted type having a second polarization orientation and preventing transmission of light of said excitation type and wherein said second polarization orientation is different than that of said first orientation; and

d) a third optical filter positioned in optical communication with said fluorophors, wherein the third optical filter is capable of transmitting light of wavelength of said emitted type from said fluorophors and of preventing transmission of light of wavelength of said excitation type.

said system being constructed and arranged such that patterns of fluorescence, emitted by said fluorophors, of wavelengths different than that of light of said excitation type, are viewable.

66. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors capable of being excited by light of an excitation type and of producing light of an emitted type, wherein at least a portion of said emitted type is of a wavelength and polarization different from said excitation type, said system comprising:

a) a non-laser light producing element that produces light, at least a portion of which is light of said excitation type;

b) a first optical polarization filter positioned between said light producing element and said fluorophors, wherein said first filter is capable of transmitting polarized light of said excitation type having a first polarization orientation and capable of preventing transmission of light from the light-producing element having a polarization orientation different than said first polarization orientation;

c) a second optical polarization filter positioned in optical communication with said fluorophors, wherein said second filter is capable of transmitting polarized light of said emitted type having a second polarization orientation and preventing transmission of light of said excitation type and wherein said second polarization orientation is different than that of said first orientation; and

d) a third optical filter positioned between said light producing element and said fluorophors, wherein the third optical filter is capable of transmitting light having a wavelength of said excitation type from said light producing element and of preventing transmission of light having a wavelength of said emitted type;

said system being constructed and arranged such that patterns of fluorescence, emitted by said fluorophors, of wavelengths different than that of light of said excitation type, are viewable.

67. A visible light fluorescence imaging system for viewing one or more patterns of fluorescence emitted by fluorophors or a material containing fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) at least one blue, fluorescent lamp that produces light, at least a portion of which is capable of exciting said fluorophors, wherein said fluorescent lamp has a maximal output at about 450 nm;

b) a blue optical filter positioned between said fluorescent lamp and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type, wherein the blue optical filter has an absorbance greater than about 3 for light having a wavelength greater than about 550 nm; and

c) an amber optical filter positioned in optical communication with said fluorophors, that is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light, wherein said amber optical filter has an absorbance greater than about 3 for light having a wavelength less than about 540 nm;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

68. The visible light fluorescence imaging system of claim 67 wherein each fluorescent lamp has a nominal power consumption of about 9 W.

69. The visible light fluorescence imaging system of claim 67 wherein each fluorescent lamp has a nominal power consumption of about 28 W.

70. The visible light fluorescence imaging system of claim 67 wherein the optical filters have a transmission surface area ranging from about 19 cm to about 14,800 cm².

71. A visible light fluorescence imaging system for viewing one or more patterns of fluorescence emitted by fluorophors or a material containing fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) at least one blue light emitting diode that produces light, at least a portion of which is capable of exciting said fluorophors, wherein said light emitting has a maximal output at about 470 nm,

b) a blue optical filter positioned between said light emitting diode and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type, wherein the blue optical filter has an absorbance greater than about 3 for light having a wavelength greater than about 550 nm; and

c) an amber optical filter positioned in optical communication with said fluorophors, that is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light, wherein said amber optical filter has an absorbance greater than about 3 for light having a wavelength less than about 540 nm.

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

72. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser light source comprising a light producing element that produces light, at least a portion of which is capable of exciting said fluorophors, and a first optical filter positioned between said light producing element and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type;

b) a second optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light; and

c) at least one additional optical filter positioned in optical communication with said second optical filter, wherein said additional optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of infrared light;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

73. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser light source comprising a light producing element that produces light, at least a portion of which is capable of exciting said fluorophors, and a first optical filter positioned between said light producing element and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type;

b) a second optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light; and

c) at least one additional optical filter positioned in optical communication with said second optical filter, wherein said additional optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of light generated by fluorescence of said second optical filter;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.

74. A visible light photoluminescent imaging system for viewing one or more patterns of fluorescence emitted by fluorophors, capable of being excited by light of an excitation type and capable of emitting light of an emitted type, wherein at least a portion of said emitted type is of a wavelength different from said excitation type, said system comprising:

a) a non-laser light source comprising a light producing element that produces light, at least a portion of which is capable of exciting said fluorophors, and a first optical filter positioned between said light producing element and said fluorophors that is capable of transmitting light of said excitation type and of preventing transmission of light of said emitted type, wherein said first filter serves as a direct support for said material containing said fluorophors; and

b) a second optical filter positioned in optical communication with said fluorophors, wherein said second optical filter is capable of transmitting light of said emitted type from said fluorophors and of preventing transmission of said excitation light;

said system being constructed and arranged such that patterns of fluorescence emitted by said fluorophors are viewable.