Depth of Field Extension for Optical Tomography

• US 20090103792A1
• Filed: 10/22/2007
• Published: 04/23/2009
• Est. Priority Date: 10/22/2007
• Status: Active Grant

• Current Assignee: VisionGate, Inc.
• Original Assignee: VisionGate, Inc.
• Inventors: Rahn, J. Richard, Hayenga, Jon W.
• Granted Patent: US 7,787,112 B2
• Time in Patent Office: Days
• US Class Current: 382/131
• CPC Class Codes: A61B 5/0073 by tomography, i.e. reconstruction of 3D images from 2D projections A61B5/0066 takes precedence
• CPC Class Codes: G01N 15/1468 with spatial resolution of the texture or inner structure of the particle
• CPC Class Codes: G01N 21/4795 spatially resolved investigating of object in scattering medium in vivo A61B

Abstract

An optical projection tomography system is illuminated with a light source. An object-containing tube, a portion of which is located within the region illuminated by the light source, contains an object of interest that has a feature of interest. A detector is located to receive emerging radiation from the object of interest. A lens, including optical field extension elements, is located in the optical path between the object region and the detector, such that light rays from multiple object planes in the object-containing tube simultaneously focus on the detector. The object-containing tube moves relatively to the detector and the lens operate to provide multiple views of the object region for producing an image of the feature of interest at each view.

Claims

1. An optical projection tomography system for imaging an object of interest, the optical projection tomography system comprising:

(a) a light source;

(b) an object-containing tube, a portion of which is located within the region illuminated by the light source, wherein the object of interest has at least one feature of interest located within the object-containing tube;

(c) at least one detector, wherein the at least one detector is located to receive emerging radiation from the object of interest; and

(d) at least one lens located in the optical path between the object region and the at least one detector, wherein the at least one lens includes at least one optical field extension element such that light rays from multiple object planes in the object-containing tube simultaneously focus on the at least one detector.

2. The system of claim 1 wherein the object moves relatively to the at least one detector and the at least one lens operates to provide multiple views of the object region for producing at least one image of the at least one feature of interest at each view.

3. The system of claim 1, wherein the object of interest comprises a biological cell.

4. The system of claim 1, wherein each of the multiple object planes that come to a focus on at least one of the at least one detectors registers light of a different optical wavelength on the at least one detector.

5. The system of claim 1, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the object of interest.

6. The system of claim 1, wherein the object of interest is stained to impart an absorption coefficient of at least one wavelength that registers on the at least one detector.

7. The system of claim 1, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the diameter of the object-containing tube.

8. The system of claim 1, wherein the system further comprises a chromatic filter array located between the at least one lens and the at least one detector.

9. The system of claim 1, wherein the system further comprises a piezoelectric transducer coupled to the at least one lens for moving the at least one lens along an optical path between each of the multiple views of the object region, such that the object of interest remains within an interval spanned by the multiple object planes.

10. The system of claim 1, further comprising a wavefront-coded optical element located between the at least one lens and the at least one detector.

11. The system of claim 1, wherein the at least one detector comprises a detector selected from the group consisting of charge coupled devices, complementary metal-oxide-semiconductor devices, solid-state image sensors, and solid-state image sensor detector arrays.

12. The system of claim 6, wherein the object of interest is stained with hematoxylin.

13. The system of claim 1, wherein the light from multiple object planes includes light having a wavelength range of 550 nm to 620 nm spanning a focus interval of up to 50 microns.

14. The system of claim 1, wherein the object-containing tube has a diameter of at least 50 microns.

15. The system of claim 2, wherein the system further comprises a piezoelectric transducer attached to the at least one lens for moving the at least one lens along an optical path between each of the multiple views of the object region, such that the object of interest remains within the focus interval.

16. The system of claim 1, wherein the system further comprises a chromatic filter array located between the at least one lens and the at least one detector.

17. The system of claim 1, wherein an interval spanned by the multiple object planes comprises a focus interval of at least 12 microns.

18. The system of claim 1, wherein the light from multiple object planes includes a wavelength range of 550 nm to 620 nm.

19. The system of claim 1, further comprising a band pass filter for passing light in the range of 550 nm to 620 nm located to band limit light reaching the at least one detector.

20. The system of claim 1, wherein the light source includes a chromatic filter located to filter light reaching the object-containing tube.

21. The system of claim 1, wherein the at least one lens comprises a hyperchromatic lens.

71. The system of claim 1, wherein the at least one optical field extension element comprises a polarizing filter array located between the at least one lens and the at least one detector.

72. The system of claim 1, wherein the at least one optical field extension element comprises a wavefront-coding element located between the hyperchromatic lens and each of the at least two detectors.

73. The system of claim 1, wherein each of the multiple object planes that come to a focus on the at least one detector is formed by light having a different polarization.

22. An optical tomography system for viewing an object of interest comprising:

a microcapillary tube viewing area for positioning the object of interest;

at least one detector;

a motor located to attach to and rotate a microcapillary tube;

means for transmitting broadband light having wavelengths between 550 nm and 620 nm into the microcapillary tube viewing area;

a hyperchromatic lens located to receive light transmitted through the microcapillary tube viewing area; and

a tube lens located to focus light rays transmitted through the hyperchromatic lens, such that light rays from multiple object planes in the microcapillary tube viewing area simultaneously focus on the at least one detector.

23. The system of claim 22, wherein the object of interest comprises a biological cell.

24. The system of claim 22, wherein the hyperchromatic lens and the tube lens operate to simultaneously focus multiple object planes from the microcapillary tube viewing area on the at least one detector.

25. The system of claim 24, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the biological cell.

26. The system of claim 23, wherein the biological cell is stained to impart an absorption coefficient of at least one wavelength that registers on the at least one detector.

27. The system of claim 23, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the microcapillary tube viewing area.

28. The system of claim 22, wherein the system further comprises a chromatic filter array located between the hyperchromatic lens and the at least one detector.

29. The system of claim 22, wherein the light from the multiple object planes includes light having a wavelength range of 550 nm to 620 nm spanning a focus interval of up to 50 microns.

30. The system of claim 22, further comprising a chromatic filter array located between the hyperchromatic lens and the at least one detector, so that light coming to a focus on the detector is separated into two or more wavelength bands, each wavelength band being transmitted through the chromatic filter array to a separate set of pixels on the at least one detector.

31. An optical tomography system for viewing all object of interest comprising:

a microcapillary tube containing the object of interest;

a motor, attached to the microcapillary tube, for rotating the microcapillary tube;

a light source located to illuminate the microcapillary tube;

a hyperchromatic lens located to receive light transmitted through the microcapillary tube;

a dichroic beamsplitter located to split a plurality of ray paths originating in multiple object planes in the microcapillary tube as transmitted through the hyperchromatic lens; and

at least two detectors, where a first detector of the at least two detectors is located to receive light transmitted along one of the plurality of ray paths, and a second detector of the at least two detectors is located to receive light transmitted along another of the plurality of ray paths, where light rays from the multiple object planes are simultaneously focused on at least one of the at least two detectors.

32. The system of claim 31, wherein the system further comprises a piezoelectric transducer coupled to the hyperchromatic lens.

33. The system of claim 31, wherein the object of interest comprises a biological cell.

34. The system of claim 31, wherein each of the multiple object planes that come to a focus registers light of a different optical wavelength on at least one of the at least two detectors.

35. The system of claim 31, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the object of interest.

36. The system of claim 31, wherein the biological cell is stained to impart an absorption coefficient of at least one wavelength.

37. The system of claim 31, wherein an interval spanned by the multiple object planes comprises an interval spanning the thickness of the microcapillary tube.

38. The system of claim 31, wherein the system further comprises a chromatic filter array located between the hyperchromatic lens and each of the at least two detectors.

39. The system of claim 36, wherein the object of interest is stained with hematoxylin.

40. The system of claim 31, wherein the light from multiple object planes includes light having a wavelength range of 550 nm to 620 nm covering a focus interval of up to 50 microns.

41. The system of claim 40, wherein the object-containing tube has a diameter of at least 50 microns.

42. The system of claim 40, wherein an interval spanned by the multiple object planes comprises a focus interval of at least 12 microns.

43. The system of claim 31, wherein the light from multiple object planes includes a wavelength range of 550 nm to 620 nm.

44. The system of claim 31, further comprising a band pass filter for passing light in the range of 550 nm to 620 nm located to band limit light reaching the at least two detectors.

45. The system of claim 31, wherein the light source includes a chromatic filter located to filter light reaching the object-containing tube.

46. A method for finding a midpoint of a focus-invariant region in a focus-invariant optical tomography system, where the optical tomography system includes an objective lens with an optical axis and a rotating tube containing an object of interest, comprising:

panning the objective lens along the optical axis to acquire multiple images of an object of interest;

evaluating focus quality for each image;

determining upper and lower boundaries of a focus invariance region;

acquiring a pseudo-projection image, where the pseudo-projection'"'"'s scanning range is centered between the upper and lower boundaries; and

rotating the tube and repeating panning, evaluating and acquiring steps above until a plurality of pseudo-projections have been acquired at a plurality of projection angles.

47. A method for computing a 3D reconstruction by finding a midpoint of a focus-invariant region in a focus-invariant optical tomography system, where the optical tomography system includes an objective lens with an optical axis and a microcapillary tube containing an object of interest, comprising:

incrementally stepping the objective lens to focus at a plurality of focal planes and acquiring a plurality of pseudo-projections of the object of interest at a plurality of focal planes in the microcapillary tube;

repeating until a limit condition on the range of focal planes is met;

rotating the tube through a plurality of viewing angles and repeating the above steps of stepping and repeating until a limit condition on the range of focal planes is met for each viewing angle;

for each viewing angle, summing the acquired pseudo-projections acquired at that viewing angle to generate a set of summed pseudo-projections; and

computing a 3D reconstruction using the summed pseudo-projections.

48. The method of claim 47 wherein the limit condition comprises meeting or exceeding the limit produced by the formula ε

nD=tube diam, where 0<

ε

<

1 and n is an incremental step value.

49. A method for applying a focus score to identify a pseudo-projection having the best focus, comprising:

incrementally stepping the objective lens to focus at a plurality of focal planes and acquiring a plurality of pseudo-projections of the object of interest at a plurality of focal planes in the microcapillary tube;

repeating until a limit condition on the range of focal planes is met;

performing a 2.5-D focus evaluation to determine a best focus mapped to the current viewing angle;

rotating the tube through a plurality of view angles and repeating the above steps of stepping and repeating for each viewing angle;

for each viewing angle, summing the acquired pseudo-projections acquired at best focus for that viewing angle to generate a set of summed pseudo-projections; and

computing a 3D reconstruction using the summed pseudo-projections.

50. The method of claim 49 wherein the limit condition comprises meeting or exceeding the limit produced by the formula ε

nD=tube diam, where 0<

ε

<

1 and n is an incremental step value.

51. An autofocusing system comprising:

a first autofocusing camera;

a second autofocusing camera;

an image comparator coupled to receive digitized input from the first and second autofocusing cameras, where the image comparator provides a feedback signal.

a transducer drive coupled to receive the feedback signal;

a microcapillary tube within a field of view of the first and second autofocusing cameras containing an object of interest, where the microcapillary tube is coupled to a rotation motor and separate images from an upper spectrum object volume and a lower spectrum object volume are captured by the first and second autofocusing cameras;

where the separate images are compared and analyzed by the comparator that provides the feedback signal to drives the transducer drive which, in turn, controls an objective lens focus drive so that the focus range of an objective lens moves closer to a region of poorer focus quality, and when the difference in focus quality between the two images becomes sufficiently small, the transducer drive stops shifting the focus range; and

an image camera located to receive images from the microcapillary tube.

52. The autofocusing system of claim 51 further comprising

a primary beam splitter;

a secondary beam splitter;

a mirror located to directing light rays to the second autofocusing camera;

a first filter located to pass light rays from the primary beam splitter to the image camera through a first image forming lens;

a second filter located to pass light rays from the secondary beam splitter directed to be first autofocusing camera through a second imaging lens; and

a third filter located to direct light rays from the mirror through a third imaging lens to the second autofocusing camera.

53. The autofocusing system of claim 52 wherein the first filter passes light having wavelengths between 550 nm and 620 nm.

54. The autofocusing system of claim 52 wherein the second filter passes light having wavelengths between 585 nm and 620 nm.

55. The autofocusing system of claim 52 wherein the third filter passes light having wavelengths between 550 nm and 585 nm.

56. An autofocusing system comprising:

a source of spectrally uniform light illumination located to illuminate an object of interest within a field of view of an image camera in line with an objective lens positioned to view an object of interest;

a primary beam splitter;

a secondary beam splitter;

a mirror;

a first filter located to pass light rays from the primary beam splitter to the image camera through a first image forming lens;

a second filter optically aligned with the secondary beam splitter;

a third filter optically aligned with the mirror;

a first Fourier plane forming lens located to transmit light from the second filter through a first Fourier plane spatial filter to a first photo sensor;

a second Fourier plane forming lens located to transmit light from the third filter through a second Fourier plane spatial filter to a second photo sensor;

where the first and second Fourier spatial filters provide analog feedback to a focus control controller through the first and second photo-diodes, further where the high frequency content of upper and lower portions of the focal range respectively, is compared in a signal conditioning and difference amplification processor and the signal conditioning and difference amplification processor provides an output which is used to a drive connected to control the objective lens position.

57. The autofocusing system of claim 56 wherein the first filter passes light having wavelengths between 550 nm and 620 nm.

58. The autofocusing system of claim 56, wherein the second filter passes light having wavelengths between 585 nm and 620 nm.

59. The autofocusing system of claim 56 wherein the third filter passes light having wavelengths between 550 and 585 nm

60. A 2.5-dimensional image analysis method for optical tomography using focus-invariant optics comprising:

identifying features, G_i=G₁, . . . G_Ω

in image stack S_k;

for each feature G₁, identify pixels of best focus X_i, Y_i, Z_i in S_k;

generating a blank composite image PP_k;

adding intensity values of X_i, Y_i, Z_i to intensity values of pixels X_i, Y_i in PP_k repeating for all features (G₁, G₂ . . . G_Ω

) until all features have been incorporated into PP_k; and

repeating for multiple perspectives.

61. The method of claim 60, wherein the focus-invariant optics include a 4-color CFA, the method further comprising processing each 2×

2 block of a 4-color CFA by selecting pixels containing the best focus.

62. The method of claim 60, wherein the focus-invariant optics include a 4-color CPA including a plurality of 2×

2 blocks, the method further comprising processing each 2×

2 block of a 4-color CFA using a weighted sum of two or more pixels.

63. The method of claim 60, wherein the focus-invariant optics include a PFA including a plurality of 2×

2 blocks, the method further comprising processing each 2×

2 block of a PFA by selecting pixels containing the best focus.

64. The method of claim 60, wherein the focus-invariant optics include a PFA including a plurality of 2×

2 blocks, the method further comprising processing each 2×

2 block of a PFA using a weighted sum of two or more pixels.

65. A folded system for optical tomography comprising:

an objective lens for transmitting collimated light;

a first dichroic beam-splitting cube located downstream from the objective lens to split the collimated light transmitted by the objective lens into a first arm and a second arm;

the first arm has a wavelength λ

1 and originates in a first focal plane;

a first tube lens is positioned to transmit the first arm through second beam-splitter cube, and onto a first region of the camera sensor'"'"'s active area;

the second arm has a wavelength λ

2 and originates in a second focal plane;

first and second mirrors positioned to pass the second arm through a tube lens after reflecting within a second dichroic beam-splitter cube and onto a second region of a camera sensor'"'"'s active area, whereby the first region and the second region of the camera acquire focused images originating from different focal planes in object space.

66. The system of claim 65 where a relative lateral shift in images is achieved by laterally shifting the second dichroic beam-splitter cube, so that the reflected light of the second arm is laterally shifted relative to the first arm and to the first tube lens.

67. A multiple camera system for optical tomography comprising:

a primary objective lens;

first and second mirrors located to receive light transmitted through the primary objective lens;

at least three beam-splitters located to direct light along different ray paths;

at least four primary tube lenses, each located along one of the ray paths;

at least four secondary objective lenses, each located along one of the ray paths;

at least four secondary tube lenses, each located along one of the ray paths; and

at least four cameras each located along one of the ray paths, where space between one of the at least four primary tube lenses and one of the at least four secondary objective lenses differs for each ray path, so that a different object plane is focused on each of the at least four cameras.

68. The system of claim 67 wherein images acquired by the at least four cameras retain the same orientation at the image planes of the at least four cameras.

69. The system of claim 67 where the primary objective lens and the at least four primary tube lenses provide at least 10×

magnification, and the secondary objectives and tube lenses provide at least and additional 10×

magnification.

70. A multiple camera system for optical tomography comprising:

a primary objective lens;

first and second mirrors located to receive light transmitted through the primary objective lens;

at least three beam-splitters located to direct light along different ray paths;

at least four primary tube lenses, each located along one of the ray paths;

at least four secondary objective lenses, each located along one of the ray paths;

at least four secondary tube lenses, each located along one of the ray paths;

at least four cameras, each located along one of the ray paths; and

wavefront coded optics positioned between the primary objective lens and the at least four cameras so that a different object plane is focused on each of the at least four cameras.

74. An optical projection tomography method for imaging an object of interest, the optical projection tomography method comprising:

(a) illuminating an object of interest; and

(b) simultaneously focusing light rays from multiple object planes in the object of interest onto at least one detector for plurality of different views.

75. The method of claim 74 further comprising filtering the light rays to pass only light rays having wavelengths within the range of 550 nm to 620 nm.

76. The method of claim 75 wherein the filtering further comprises passing the light through a band pass filter located to limit light reaching at least one detector.

77. The method of claim 74 further comprising delivering the object of interest for imaging from an object containing tube;

wherein the object containing tube comprises a tube selected from the group consisting of a capillary tube, a microcapillary tub, a microcapillary tube bundle, and a microcapillary tube cassette.

78. The method of claim 77 wherein the object containing tube has a diameter of at least 50 microns.

79. The method of claim 74, wherein simultaneously focusing comprises moving at least one lens of a hyperchromatic lens system relative to at least one detector.

80. The method of claim 74 further comprising splitting a first plurality of light ray paths via a beamsplitter.

81. The method of claim 74 wherein the simultaneous focusing comprises focusing a hyperchromatic lens.

82. The method of claim 79 wherein the simultaneous focusing further comprises filtering the light by transmitting the light through at least one chromatic filter array.

83. The method of claim 74 wherein the at least one detector comprises a detector selected from the group consisting of charge coupled devices, complementary metal-oxide-semiconductor devices, solid-state image sensors, and solid-state image sensor detector arrays.

84. The method of claim 82, further comprising locating the at least one chromatic filter array between the hyperchromatic lens and the at least one detector, so that light coming to a focus on the detector is separated into two or more wavelength bands, each wavelength band being transmitted through the chromatic filter array to a separate set of pixels on the least one detector.

85. The method of claim 82, wherein the at least one chromatic filter array is substituted with at least one polarization filter array.

86. The method of claim 74, wherein simultaneously focusing of the multiple object planes comprises extending focusing through a focus interval spanning the thickness of the object of interest.

87. The method of claim 74, wherein the simultaneously focusing of the multiple object planes comprises extending focusing through a focus interval of at least 12 microns.

88. The method of claim 74, wherein the simultaneously focusing of the multiple object planes comprises extending focusing through a focus interval of up to 50 microns.

89. The method of claim 74 further comprising impinging the light rays onto a wave front-coded optical element.

90. The method of claim 74, further comprising staining the object of interest to impart an absorption coefficient of at least one wavelength.

91. The method of claim 90, wherein the staining of the object of interest is performed with hematoxylin.

92. The method of claim 74, wherein simultaneously focusing of the multiple object planes comprises simultaneously focusing with an autofocusing system.

93. The method of claim 89, wherein the simultaneously focusing with an autofocusing system comprises focusing using chromatic balance.

94. The method of claim 74, wherein simultaneously focusing light rays comprises transmitting the light rays through a polarizing filter array.

95. The method of claim 74, wherein simultaneously focusing light rays comprises transmitting the light rays through a wavefront-coding element.

96. The method of claim 74, wherein simultaneously focusing light rays comprises focusing each of the multiple object planes on the at least one detector by light having a different polarization.

97. The method of claim 74, wherein simultaneously focusing light rays comprises performing a 2.5-D focus evaluation to determine a best focus.

98. The method of claim 74, wherein the object of interest comprises a biological cell.