Kolenbrander, Jeremy P. (Brighton, CO, US)
Ladtkow, James R. (Broomfield, CO, US)
Scibona, Joseph A. (Littleton, CO, US)
Steward, Jeffrey A. (Lakewood, CO, US)
Fletcher, Chris (Superior, CO, US)
Plaque It!
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International Search Report for PCT/US2004/021344 mailed Nov. 17, 2004.
CROSS-REFERENCE TO RELATED APPLICATIONS
This application claims priority under 35 U.S.C. 119(e) to provisional patent application 60/485,015, filed Jul. 2, 2003, which is hereby incorporated by reference in its entirety to the extent not inconsistent with the disclosure herein.
1. A system for processing blood by separating fluid components comprising: a density centrifuge blood processing system having a separation chamber rotating about a central rotation axis—and operable to extract and collect a separated, selected component from the chamber; a light source in optical communication with said density centrifuge blood processing system for providing an incident light beam for illuminating an observation region on said separation chamber of said density centrifuge blood processing system, thereby generating light transmitted, scattered or both from said observation region; a light collection element in optical communication with said density centrifuge blood processing system for collecting at least a portion of said light transmitted, scattered or both from said observation region and for directing at least a portion of said light transmitted, scattered or both from said observation region onto a two-dimensional detector; the two-dimensional detector position to receive and detect said light transmitted, scattered or both from said observation region provided by said light collection element, thereby measuring a two-dimensional distribution of intensities of said light transmitted, scattered or both from said observation region; said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprising an image of at least a portion of said density centrifuge blood processing system, said two-dimensional distribution of the intensities of said light transmitted, scattered or both providing a measurement of the positions of one or more phase boundary between optically differentiable blood components in said seperation chamber, and wherein said two-dimensional detector generates an output signal corresponding to said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation; and a centrifugation device controller in communication with said two-dimensional detector for receiving said output signal, wherein said controller controls the positions of one or more phase boundary by adjusting a flow-related operating parameter of the density centrifuge blood processing system.
2. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of said at least a portion of said separation chamber.
3. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprise an image of said at least a portion of an optical cell of said separation chamber.
4. The system of claim 3 wherein the image of a phase boundary between optically differentiable blood components provides said measurement of the position of the phase boundary.
5. The system of claim 1 wherein at least a portion of said light transmitted, scattered or both from said observation region is light transmitted through the separation chamber.
6. The system of claim 1 wherein at least a portion of said light transmitted, scattered or both from said observation region is light scattered by the separation chamber or materials therein.
7. The system of claim 1 wherein said centrifugation device controller and said two-dimensional detector provide feedback control of the positions of one or more phase boundary between optically differentiable blood components in said separation chamber.
8. The system of claim 1 wherein said centrifugation device controller is capable of adjusting a plurality of operating parameters of the density centrifuge blood processing system.
9. The system of claim 1 wherein said operating parameter is selected from the group consisting of: a flow rate of a fluid component out of the separation chamber; a flow rate of a fluid into the separation chamber; and the rate of rotation of the separation chamber about the central rotation axis.
10. The system of claim 1 wherein the light collection element and two-dimensional detector are stationary and provided in fixed positions relative to the density centrifuge blood processing system.
11. The system of claim 1 wherein the light collection element and two-dimensional detector are stationary and provided in fixed orientations relative to said density centrifuge blood processing system.
12. The system of claim 1 wherein at least a portion of said observation region is positioned such that a phase boundary between optically differentiable blood components in said separation chamber is viewable by the two-dimensional detector.
13. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of a phase boundary between optically differentiable blood components.
14. The system of claim 1 wherein at least a portion of said observation region is positioned such that a separated blood component in said separation chamber is viewable by two-dimensional detector.
15. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of a separated blood component in the separation chamber.
16. The system of claim 15 wherein the image of the separated blood component in the separation chamber provides a measurement of the composition of the separated blood component.
17. The system of claim 15 wherein said separated blood component is selected from the group consisting of: a white blood cell containing component; a platelet containing component; a plasma containing component; and a red blood cell containing component.
18. The system of claim 1 wherein at least a portion of the observation region is positioned such that an extraction port of the separation chamber is viewable by the two dimensional detector.
19. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of an extraction port of the separation chamber.
20. The system of claim 19 wherein the image of the extraction port of the separation chamber provides a measurement of the composition of cellular components in said extraction port.
21. The system of claim 20 wherein said cellular components are selected from the group consisting of: red blood cells; white blood cells; and platelets.
22. The system of claim 19 wherein the image of the extraction port of the separation chamber provides a measurement of the flux of cellular components through the extraction port.
23. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of at least one phase boundary between optically differentiable blood components in the separation chamber and at least one extraction port of the separation chamber.
24. The system of claim 23 wherein the image provides simultaneous measurements of said position of at least one phase boundary between optically differentiable blood components and the composition of blood components in the extraction port of the separation chamber.
25. The system of claim 1 wherein the observation region is positioned such that leakage of fluid in said separation chamber is viewable by the two-dimensional detector.
26. The system of claim 1 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of a sample identification marker or kit identification marker corresponding to a blood sample undergoing processing.
27. The system of claim 26 wherein said sample identification marker or kit identification marker comprises a bar code.
28. The system of claim 1 wherein the observation region is positioned such that improper alignment of said separation chamber is detectable by the two dimensional detector.
29. The system of claim 1 wherein the two-dimensional detector is selected from the group consisting of: a charge coupled device; a two-dimensional photodiode array; a two-dimensional photoconductive array; a two-dimensional pyroelectric array; a CCD camera; and a complementary metal oxide semiconductor detector.
30. The system of claim 1 wherein the two-dimensional detector is a monochrome CCD camera.
31. The system of claim 1 wherein the two-dimensional detector is a color CCD camera.
32. The system of claim 1 wherein the light collection element comprises a lens.
33. The system of claim 1 wherein the light collection element comprises a lens system.
34. The system of claim 33 wherein the lens system has a selectively adjustable focal length.
35. The system of claim 1 wherein the light collection element comprises a fixed focus lens system.
36. The system of claim 1 wherein the light collection element has a fixed field of view.
37. The system of claim 1, wherein the light collection element has a selectively adjustable field of view.
38. The system of claim 1 wherein said two-dimensional detector and light collection element are selectively positionable along a detector axis oriented orthogonal to the central rotation axis.
39. The system of claim 1 wherein the light source is selected from the group comprising: a xenon lamp; a light emitting diode; and an array of light emitting diodes;.
40. The system of claim 1 wherein the light source is selected from the group comprising: a white light emitting diode; blue light-emitting diode; a red light-emitting diode; and a green light-emitting diode.
41. The system of claim 1 wherein the light source comprises a parabolic reflector element.
42. The system of claim 1 further comprising an additional light source, wherein said light source illuminates a first side of said separation chamber and said additional light source illuminates a second side of said separation chamber, and wherein said first and second sides are different sides.
43. The system of claim 42 wherein the additional light source is selected from the group comprising: a xenon lamp; a light emitting diode; and an array of light emitting diodes.
44. The system of claim 1 wherein the light source provides an incident light beam comprising visible light.
45. The system of claim 1 wherein the light source provides an incident light beam comprising infrared light.
46. The system of claim 1 wherein the light source provides an incident light beam comprising ultraviolet light.
47. The system of claim 1 wherein the light source provides continuous illumination of the observation region on the separation chamber.
48. The system of claim 1 wherein the light source is capable of being switched on and off in a manner providing pulsed illumination of the observation region on the separation chamber.
49. The system of claim 1 wherein said separation chamber comprises an optical cell.
50. The system of claim 49 wherein said optical cell comprises: an extraction chamber having a first external optical surface and a second external optical surface, wherein said first and second external optical surfaces of said extraction chamber are opposing planar surfaces; and an extraction port having a first external optical surface and a second external optical surface, wherein said first and second external optical surfaces of said extraction chamber are opposing planar surfaces; wherein said first optical surface of said extraction chamber and said first optical surface of said extraction port are both in the depth of field of the light collection element.
51. The system of claim 50 wherein first optical surface of said extraction chamber and said first optical surface of said extraction port are both substantially in the same plane.
52. The system of claim 1 further comprising a calibration marker, wherein said calibration marker is in said observation region.
53. The system of claim 52 wherein the calibration marker is in the depth of field of the light collection element.
54. The system of claim 52 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of said calibration marker.
55. The system of claim 1 further comprising an additional light collection element and an additional two-dimensional detector, wherein light transmitted, scattered or both from an additional observation region is collected by the additional light collection element and detected by the additional two-dimensional detector.
56. The system of claim 55 wherein said additional two-dimensional detector measures a two-dimensional distribution of the intensities of said light transmitted, scattered or both from said additional observation region.
57. The system of claim 1 wherein said fluid components are blood components.
58. The system of claim 1, wherein said centrifugation device controller determines the position of phase boundaries between said optically differentiable blood components.
59. The system of claim 1, wherein said centrifugation device controller determines one or more of the following: a flux of cellular material out of the separation chamber; a flux of noncellular material out of the separation chamber; a composition of an extracted blood component; a hematocrit of an extracted blood component; and an extent of hemolysis in said blood.
60. A method of monitoring a density centrifuge blood processing system comprising the steps of: providing said density centrifuge blood processing system having a separation chamber rotating about a central rotation axis and operable to extract and collect a separated, selected component from the chamber; illuminating an observation region on a density centrifuge blood processing system with an incident light beam provided by a light source, thereby generating light transmitted, scattered or both from said observation region; collecting at least a portion of light transmitted, scattered or both from said observation region and directing said light transmitted, scattered or both from said observation region onto a two-dimensional detector; measuring two-dimensional distributions of intensities of said light transmitted, scattered or both from said observation region; said two-dimensional distributions of the intensities of said light transmitted, scattered or both from said observation region comprising images of at least a portion of said density centrifuge blood processing system, said two-dimensional distributions of the intensities of said light transmitted, scattered or both providing measurements of the positions of one or more phase boundary between optically differentiable blood components in said separation chamber; generating output signals corresponding to said two-dimensional distributions of the intensities of said light transmitted, scattered or both from said observation region; and providing a centrifugation device controller for receiving said output signals, wherein said controller provides feedback control of said positions of said one or more phase boundary based on said output signals, wherein said controller controls the positions of said one or more phase boundary by adjusting a flow-related operating parameter of the density centrifuge blood processing system.
61. The method of claim 60 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of said at least a portion of said separation chamber.
62. The method of claim 60 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of said at least a portion of an optical cell of said separation chamber.
63. The method of claim 62 wherein said phase boundary between optically differentiable components in the separation chamber is viewable in said image.
64. The method of claim 63 further comprising measuring the position of said phase boundary.
65. The method of claim 62 wherein an extraction port of the separation chamber is viewable in said image.
66. The method of claim 65 further comprising determining the average intensity of light transmitted through a selected portion of said extraction port.
67. The method of claim 66 further comprising measuring a composition of a component passing through said extraction port.
68. The method of claim 60 further comprising the step of providing a calibration marker in the depth of field of the light collection element.
69. The method of claim 60 wherein said step of illuminating the observation region comprises providing a pulsed incident light beam.
70. The method of claim 69 further comprising the step of selecting the rotational position of the separation chamber corresponding to the two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region by selecting the timing and duration of the pulsed incident light beam.
71. The method of claim 70 further comprising the step of selecting different rotational positions of the separation chamber corresponding to the two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region for different rotations of said separation chamber.
72. The method of claim 60 wherein said step of illuminating the observation region comprises providing a plurality of pulsed incident light beams.
73. A method of controlling a blood centrifuge blood processing system for separating fluid components comprising the steps of: providing said density centrifuge blood processing system having a separation chamber rotating about a central rotation axis and operable to extract and collect a separated, selected component from the chamber; illuminating an observation region on a density centrifuge blood processing system with an incident light beam provided by a light source, thereby generating light transmitted, scattered or both from said observation region; collecting at least a portion of light transmitted, scattered or both from said observation region and directing said light transmitted, scattered or both from said observation region onto a two-dimensional detector; measunng a two-dimensional distribution of intensities of said light transmitted, scattered or both from said observation region; said two-dimensional distributions of the intensities of said light transmitted, scattered or both from said observation region comprising an image of at least a portion of said density centrifuge blood processing system, wherein one or more phase boundary between optically differentiable blood components in said separation chamber is viewable in said image; measuring the positions of said one or more phase boundary between optically differentiable blood components in said separation chamber; and controlling the positions of said one or more phase boundary by selectively adjusting a flow-related operating parameter of said density centrifuge blood processing system based on said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region, wherein said flow-related operating parameter is selected from the group consisting of: a flow rate of a fluid component out of the separation chamber; a flow rate of a fluid into the separation chamber; and the rate of rotation of the separation chamber about the central rotation axis.
74. The method of claim 73 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of a phase boundary between optically differentiable components in said separation chamber.
75. The method of claim 74 further comprising the step of measuring the position of the phase boundary between optically differentiable components in said separation chamber.
76. The method of claim 75 wherein the operating parameter of said density centrifuge blood processing system is selectively adjusted on the basis of the position of the phase boundary between optically differentiable components in said separation chamber.
77. The method of claim 73 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of an extraction port of said separation chamber.
78. The method of claim 77 further comprising the step of measuring the composition of fluid components passing through said extraction port of said separation chamber.
79. The method of claim 78 wherein said operating parameter of said density centrifuge blood processing system is selectively adjusted on the basis of the composition of fluid components passing through said extraction port of said separation chamber.
80. The method of claim 78 wherein said operating parameter of said density centrifuge blood processing system is adjusted such that the composition of fluid components passing through said extraction port of said separation chamber is in a predefined range of compositions.
81. The method of claim 73 wherein said two-dimensional distribution of the intensities of said light transmitted, scattered or both from said observation region comprises an image of an extraction port of said separation chamber and said phase boundary between optically differentiable components in said separation chamber.
82. The method of claim 81 further comprising the step of measuring the composition of fluid components passing through said extraction port of said separation chamber and measuring the position of the phase boundary between optically differentiable components in said separation chamber.
83. The method of claim 81 wherein said operating parameter of said density centrifuge blood processing system is selectively adjusted on the basis of the composition of fluid components passing through said extraction port of said separation chamber and the position of the phase boundary between optically differentiable components in said separation chamber.
84. The method of claim 73 further comprising the step of providing a calibration marker in the depth of field of the light collection element.
85. The method of claim 73 further comprising the step of selectively adjusting a plurality of operating parameters of said density centrifuge blood processing system.
86. The method of claim 73 wherein said fluid components are blood components.
BACKGROUND OF INVENTION
Large scale blood collection and processing play important roles in the worldwide health care system. In conventional large scale blood collection, blood is removed from a donor or patient, separated into its various blood components via centrifugation, filtration and/or elutriation and stored in sterile containers for future infusion into a patient for therapeutic use. The separated blood components typically include fractions corresponding to red blood cells, white blood cells, platelets and plasma. Separation of blood into its components can be performed continuously during collection or can be performed subsequent to collection in batches, particularly with respect to the processing of whole blood samples. Separation of blood into its various components under highly sterile conditions is critical to most therapeutic applications.
Recently, apheresis blood collection techniques have been adopted in many large scale blood collection centers wherein a selected component of blood is collected and the balance of the blood is returned to the donor during collection. In apheresis, blood is removed from a donor and immediately separated into its components by on-line blood processing methods. Typically, on-line blood processing is provided by density centrifugation, filtration and/or diffusion-based separation techniques. One or more of the separated blood components are collected and stored in sterile containers, while the remaining blood components are directly re-circulated to the donor. An advantage of this method is that it allows more frequent donation from an individual donor because only a selected blood component is collected and purified. For example, a donor undergoing plateletpheresis, whereby platelets are collected and the non-platelet blood components are returned to the donor, may donate blood as often as once every fourteen days.
Apheresis blood processing also plays an important role in a large number of therapeutic procedures. In these methods, blood is withdrawn from a patient undergoing therapy, separated, and a selected fraction is collected while the remainder is returned to the patient. For example, a patient may undergo leukapheresis prior to radiation therapy, whereby the white blood cell component of his blood is separated, collected and stored to avoid exposure to radiation. Alternatively, apheresis techniques may be used to perform red blood cell exchange for patients with hematological disorders such as sickle cell anemia and thalassemia, whereby a patient's red blood cell component is removed and donated packed red blood cells are provided to the patient along with his remaining blood components. Further, apheresis may be used to perform therapeutic platelet depletion for patients having thrombocytosis and therapeutic plasma exchange for patients with autoimmune diseases.
Both conventional blood collection and apheresis systems typically employ differential centrifugation methods for separating blood into its various blood components. In differential centrifugation, blood is circulated through a sterile separation chamber which is rotated at high rotational speeds about a central rotation axis. Rotation of the separation chamber creates a centrifugal force directed along rotating axes of separation oriented perpendicular to the central rotation axis of the centrifuge. The centrifugal force generated upon rotation separates particles suspended in the blood sample into discrete fractions having different densities. Specifically, a blood sample separates into discrete phases corresponding to a higher density fraction comprising red blood cells and a lower density fraction comprising plasma. In addition, an intermediate density fraction comprising platelets and leukocytes forms an interface layer between the red blood cells and the plasma. Descriptions of blood centrifugation devices are provided in U.S. Pat. No. 5,653,887 and U.S. patent application Ser. No. 10/413,890.
To achieve continuous, high throughput blood separation, extraction or collect ports are provided in most separation chambers. Extraction ports are capable of withdrawing material from the separation chamber at adjustable flow rates and, typically, are disposed at selected positions along the separation axis corresponding to discrete blood components. To ensure the extracted fluid exiting a selected extraction port is substantially limited to a single phase, however, the phase boundaries between the separated blood components must be positioned along the separation axis such that an extraction port contacts a single phase. For example, if the fraction containing white blood cells resides too close to the extraction port corresponding to platelet enriched plasma, white blood cells may enter the platelet enriched plasma stream exiting the separation chamber, thereby degrading the extent of separation achieved during blood processing. Although conventional blood processing via density centrifugation is capable of efficient separation of individual blood components, the purities of individual components obtained using this method is often not optimal for use in many therapeutic applications. For example, centrifugation separation of blood samples is unable to consistently (99% of the time) produce separated platelet components which have less than 1×10 6 white blood cells per every 3×10 11 platelets collected. The presence of white blood cells in platelet products increases the risks of viral exposure and immunological complications upon infusion into a patient.
As a result of the inability to achieve optimal purity levels using centrifugation separation alone, a number of complementary separation techniques based on filtration, elutriation and affinity-based techniques have been developed to achieve the optimal purities needed for use of blood components as therapeutic agents. These techniques, however, often reduce the overall yield realized and may reduce the therapeutic efficacy of the blood components collected. Exemplary methods and devices of blood processing via filtration, elutriation and affinity based methods are described in U.S. Pat. No. 6,334,842 and International Patent Application Serial No. PCT/US03/117764.
The purity of extracted blood components using density centrifugation is currently limited by the control of the position of phase boundary layers between separated components provided by conventional centrifugation devices and methods. The position of phase boundaries along the separation axis depends on a number of variables. First, phase boundary positions depend on the relative flow rates of individual blood components out of the separation chamber. Second, phase boundary positions depend on the rotational velocity of the separation chamber about the central rotation axis and the temperature of the blood undergoing separation. Third, phase boundary positions vary with the composition of the blood undergoing processing. Blood sample composition may vary considerably from donor to donor and/or from patient to patient. In addition, blood composition may vary significantly as function of time for a given donor or patient, especially as blood is recycled through the separation chamber multiple times. Given the sensitivity of the phase boundary position to many variables which change from person to person and during processing, it is important to monitor the position of the phase boundaries during blood processing to ensure optimal separation conditions are maintained and the desired purity of selected blood components is achieved. In addition, accurate characterization of the positions of phase boundaries allows for separation conditions to be adjusted and optimized for changes in blood composition during processing.
Although capable of measuring the position of one or more phase boundaries, conventional optical monitoring and control methods for blood processing have substantial limitations. First, conventional optical monitoring systems and methods, such as those discussed in U.S. Pat. Nos. 5,316,667 and 5,260,598, utilize one-dimensional optical detection or one-dimensional optical scanning. Accordingly, these methods are unable to characterize the intensities of transmitted and/or scattered light from a two-dimensional or three-dimensional region of a blood processing device. Moreover, these methods are unable to measure the flux or purities of cellular material exiting the separation chamber through a selected extraction port. Second, conventional optical monitoring methods lack the signal-to-noise ratios needed for many blood processing applications because light intensities characterized are limited to a single optical axis. For example, conventional optical monitoring methods lack the sensitivity needed to accurately resolve the position of the phase boundaries between white blood cells and other blood components because white blood cells comprise less than 1% of total blood volume. Therefore, these methods are not capable of providing blood components, such as platelets and red blood cells, with white blood cell levels reduced to the extent needed to avoid immunological complications and viral transmission. Third, conventional optical monitoring methods are limited to fixed optical geometries and are incapable of monitoring regions of the density centrifuge device located on a plurality of different optical axes. As a result, the functional capabilities of conventional optical methods for monitoring and controlling separation by density centrifugation are substantially limited to monitoring the position of phase boundaries in the separation chamber.
It will be appreciated from the foregoing that a need exists for methods and devices for monitoring and controlling the processing of whole blood samples and blood component samples. Particularly, optical monitoring methods and devices are needed which are capable of accurately characterizing the separation, extraction and collection of blood components processed by density centrifugation. In addition, multifunctional optical monitoring and control systems for blood processing are needed which are capable of simultaneously monitoring a plurality of regions corresponding to a separation region, sample identification region and a blood component extraction region. Accordingly, it is an object of the present invention to provide methods, devices and device components for blood processing which are capable of high throughput separation, characterization and collection of individual blood components, particularly red blood cells, white blood cells, platelet enriched plasma and plasma.
SUMMARY OF THE INVENTION
This invention provides methods, devices and device components for improving the processing of fluids comprising fluid components, such as blood, components of blood and fluids derived from blood. Methods, devices and device components of the present invention are capable of monitoring and controlling separation of blood into discrete components and subsequent collection of selected components. The present invention includes methods, devices and device components for optically monitoring blood processing via a wide range of separation techniques, including density centrifugation, centrifugal elutriation, size and shape filtration, affinity chromatography or any combination of these techniques. The methods, devices and device components of the present invention are capable of characterizing the composition and purity of a collected blood component and capable of measuring the rate in which a blood component is extracted and collected. In addition, the methods, devices and device components of the present invention are capable of controlling blood processing by optimizing separation and extraction conditions to reproducibly achieve a desired selected purity and/or composition of a blood component. The present invention improves processing of static blood samples or flowing blood samples.
In one aspect, this invention provides methods, devices and device components for improving the separation of whole blood via density centrifugation and subsequent collection of selected, separated blood components. Particularly, the invention relates to optical methods, devices and device components for measuring two-dimensional distributions of light intensities corresponding to light transmitted and/or scattered by separated blood components in a rotating separation chamber, particularly a separation chamber having an optical cell with one or more extraction ports. In one embodiment, two-dimensional distributions of light intensities measured by the present invention comprise two- or three-dimensional images of device components of a density centrifuge systems, such as a separation chamber, an optical cell and/or one or more extraction ports, and materials disposed therein. The measured two-dimensional distributions of light intensities comprising images of device components of a density centrifuge provide quantitative information relating to important optimizing operating conditions of the centrifugation device. First, two-dimensional distributions of light intensities measured by the present invention provide an in situ and real time measurement of the position of one or more phase boundaries between optically differentiable blood components undergoing separation. Second, measured two-dimensional distributions of transmitted and/or scattered light intensities provide an in situ and real time measurement of the composition of one or more separated blood components such as separated blood components exiting an extraction port of an optical cell. Third, measured two-dimensional distributions of transmitted and/or scattered light intensities provide an in situ and real time measurement of the flux of cellular blood components exiting the separation chamber through one or more extraction ports of an optical cell. Fourth, measured two-dimensional distributions of transmitted and/or scattered light intensities provide a means of sensing identity information, such as identification number and/or lot identification number corresponding to a blood sample undergoing processing and the kit or container holding the blood sample. Automated sample and lot identification is beneficial because this information can be used to confirm that the appropriate blood processing procedure is selected and carried out for a given sample. Finally, measured two-dimensional distributions of transmitted and/or scattered light intensities provide a means of monitoring the alignment of the separation chamber in a blood processing device and identifying leakage of fluid out of the separation chamber.
In one aspect, the present invention relates to multifunctional optical monitoring systems for a blood processing device, particularly a density centrifuge. An optical monitoring system is provided which is capable of measuring a two dimensional distribution of transmitted and/or scattered light intensities corresponding to patterns of light transmitted and/or scattered from an observation region positioned on a density centrifuge, such as an observation region corresponding to an optical cell of a separation chamber. In an embodiment, a dynamic optical monitoring system of the present invention is capable of measuring a two-dimensional distribution of scattered any/or transmitted light comprising an image of an observation region having a position which is selectively adjustable before, during and/or after processing. Alternatively, the optical monitoring system of the present invention is capable of measuring a two-dimensional distribution of scattered any/or transmitted light corresponding to an observation region having a selectively adjustable size. Alternatively, the present invention includes optical monitoring systems having a selected, fixed position observation region. Use of a fixed position observation region provides highly stable monitoring systems capable of generating very reproducible images. Monitoring systems of the present invention are capable of monitoring the position of boundary layers between optically differentiable components, identifying and tracking a blood sample undergoing processing, detecting leaks and misalignment of the separation chamber, monitoring the composition of extracted blood components, monitoring the composition of a blood sample prior to processing, regulating the administration of anti-coagulation agents or other blood treatment agents added to the blood sample and characterizing the flux of cellular blood components extracted from the centrifuge.
In another aspect, the present invention relates to multifunctional control systems for a blood processing device, particularly a density centrifuge. Feedback control systems are provided wherein two-dimensional distributions of transmitted and/or scattered light intensities corresponding to patterns of light originating from an observation region on a separation chamber are generated and processed, preferably in real time. The two-dimensional distributions of transmitted and/or scattered light intensities acquired serve as the basis for control signals transmitted to various components of a density centrifuge. These control signals can be used to selectively adjust the separation conditions of the blood sample undergoing processing, such as the position of phase boundaries between optically differentiable components, and the composition, purities and flow rates of separated components out of the density centrifuge. In a preferred embodiment, images of the separation chamber identifying the positions of phase boundaries between separated blood components are used to select flow rates of these components out of the separation chamber. In this embodiment, flow rates can be selected to provide and maintain a desired extent of separation during processing and extraction. In another exemplary embodiment, two-dimensional distributions of transmitted and/or scattered light intensities comprising images of one or more extraction ports are acquired and processed in real time to determine the composition and/or fluxes of cellular material exiting the separation chamber via extraction ports. In this embodiment, fluxes of separated components can be utilized to select the processing times and flow rates needed to collect a selected amount of a particular blood component or can be utilized to determine the return rate of a selected blood component to a donor or patient in apheresis blood processing. In another embodiment, flow rates of blood components are selectively adjusted to select a desired composition and/or purity of an extracted blood component
An exemplary optical monitoring system for a density centrifuge having a separation chamber rotating about a central rotation axis comprises at least one light source, a light collection element and a two-dimensional detector. Rotation of the separation chamber about a central rotation axis results in separation of the blood components in the separation chamber according to density along rotating separation axes oriented perpendicular to the central rotation axis of the centrifuge. Both the light source and light collection element are arranged such that they are periodically in optical communication with an observation region positioned on the density centrifuge. In one embodiment, the light source and two dimensional detector are arranged such that an optical cell of the separation chamber is periodically rotated into and out of the observation region. The light source is capable of providing an incident light beam which illuminates at least a portion of the density centrifuge, preferably an optical cell of the rotating separation chamber, thereby generating light which is transmitted, scattered, or both, by blood components undergoing separation. Preferred light sources are capable of generating an incident light beam comprising light having a selected wavelength range including, but not limited to, visible light, infrared light and/or ultraviolet light. In one embodiment, a plurality of light sources are provided capable of illuminating a plurality of sides of an optical cell of a separation chamber.
The light collection element is capable of collecting light from an observation region. In one embodiment, collected light from the observation region corresponds to light which is transmitted and/or scattered by blood components undergoing separation, light which is transmitted and/or scattered by components of the centrifugation device, such as the separation chamber, or both. The light collection element directs the collected light onto the two-dimensional detector. The two-dimensional detector detects the light received from the light collection element and measures a two-dimensional distribution of transmitted and/or scattered light intensities corresponding to patterns of transmitted and/or scattered light. In one embodiment, the light collection element and two-dimensional detector are arranged such that the relative spatial distribution of scattered and/or transmitted light from the observation region is preserved during collection and detection. In a preferred embodiment, the two-dimensional detector is also capable of generating one or more output signals corresponding to the two-dimensional distribution of transmitted and/or scattered light intensities from the observation region. In one embodiment, the output signal is transmitted to a device, such as a computer, capable of displaying the two-dimensional distribution of intensities, storing the two-dimensional distribution of intensities and/or processing the two-dimensional distribution of intensities. Alternatively, the output signal is transmitted to a device, such as a computer, capable of controlling operating settings of the density centrifuge. In a preferred embodiment, the output signal is sent to a device controller which ascertains a number of important operating parameters from the two-dimensional distribution of intensities acquired. Device controllers of the present invention are capable of determining the position of phase boundaries between optically differentiable blood components, the fluxes of cellular materials and noncellular materials out of the separation chamber, the composition of extracted blood components, hematocrit, and the extent of hemolysis in a blood sample. In one embodiment, the device controller is also capable of quantifying in real time the uncertainty in operating parameters ascertained from two-dimensional distribution of scattered and/or transmitted light intensities.
In an embodiment having a dynamic observation region, the position of the observation region on the blood processing device is selectively adjustable. In an exemplary embodiment, the position of the observation region is adjusted by varying the position and/or field of view of the light collection element. For example, in one embodiment the light collection element and two-dimensional detector are arranged such that they are selectively positionable along a detection axis positioned orthogonal to the central rotation axis. In this embodiment, translation of the light collection element and two-dimensional detector along the detection axis allows selective adjustment of the position of the observation region along a separation axis of the centrifugation device. In an alternative embodiment, the size of the observation region is selectively adjustable, for example by adjusting the length, width, or radius of the observation region or any combination of these. For example, the size of the observation region can be adjusted by varying the field of view of one or more lenses or lens systems comprising the light collection element. In an embodiment, the ability to selectively adjust the position, size, or both, of the observation region before, during and after processing provides multifunctional optical monitoring systems capable of observing and controlling a plurality of important device operating conditions.
In another aspect, the present invention comprises an optical monitoring and control system capable of measuring the position of phase boundaries between optically differentiable blood components. In this embodiment, the observation region is positioned such that phase boundaries between optically differentiable components are viewable, for example once per rotation of the centrifuge. For example, in an embodiment, an interface area is periodically rotated into the observation region upon rotation of the separation chamber. Reference to an interface region in the present invention refers to an area of the separation chamber wherein two or more separated phases are viewable. Exemplary interface regions refer to a region of the separation chamber having one or more windows for transmitting light through the separated blood components, such as an optical cell. For example, in a preferred embodiment, the interface area is defined by an optical cell wherein the phase boundaries between optically differentiable blood components are viewable, such as the phase boundary between red blood cells and the buffy coat layer and the phase boundary between the buffy coat layer and the plasma. In an exemplary, phase boundaries within a mixed-phase layer, such as the buffy coat layer, are viewable. For example, the present invention provides a means of monitoring the phase boundary between a white blood cell-containing layer and a platelet enriched plasma layer.
In a preferred embodiment, illumination of the separation chamber generates patterns of light transmitted and/or scattered from separated blood fractions in the interface region. Optically differentiable blood components generate different intensities of transmitted or scattered light. Therefore, detection of patterns of transmitted light, scattered light, or both, corresponding to an observation region provides a direct measurement of the positions of phase boundaries along the separation axis of a density centrifuge. In a preferred embodiment optically differentiable components have transmitted and/or scattered light intensities that differ by about 30 relative intensity units, wherein a relative intensity unit reflects a range of 0-255 intensity units and a value of 0 corresponds to no detected light and a value of 255 corresponds to an intensity which saturates the detector. In an exemplary embodiment, at least one calibration marker is provided in the observation region. Calibration markers of the present invention have well known optical properties, such as absorption coefficients, scattering cross sections, lengths and widths, and provide spatial reference points for resolving the positions of optically differentiable blood components along the separation axis. Calibration markers also provide a reference for optimizing focusing of the light collection element and providing a brightness and/or color index to calibrate measured light intensities.
Measurement of a two-dimensional distribution of scattered and/or transmitted light intensities in the present invention is beneficial because it provides a sensitive measurement of the position of one or more phase boundaries along the separation axis. For example, acquisition of a two-dimensional distribution of scattered and/or transmitted light intensities from a 0.2-0.4 inches 2 observation region provides a measurement of the position of a phase boundary accurate to within about 0.0005±0.0002 inch 2 .
In another preferred embodiment, the present invention comprises an optical monitoring system capable of providing in situ measurements of the composition of one or more blood component undergoing processing in a density centrifuge, such as an extracted blood component. Reference to composition in this context relates to the amount, identity and purity of cellular materials, such as erythrocytes, leukocytes and thrombocytes, and non-cellular materials, such as blood plasma proteins, in a given blood component, such as an extracted component. Measurement of the composition of a selected blood component includes, but is not limited to, measurement of cell types and concentration, and purity of a given separated fraction or mixed fraction. Composition measurements can be used to predict yield and quality. Exemplary composition measurements are also be the basis of control signals for optimizing separation and extraction conditions to achieve desired compositions of one or more extracted components. In an embodiment of the present invention, the observation region is positioned such that at least one separated blood component is viewable. For example, in one embodiment a composition-monitoring region is periodically rotated into the observation region as the separation chamber is rotated about the central rotation axis. Reference to a composition-monitoring region in the present invention relates to a portion of the separation chamber occupied by at least one separated component, such as an extraction port of an optical cell of a separation chamber. In one embodiment, the separation chamber is arranged such that upon illumination, light is transmitted through at least one separated component to provide a measurement of composition. Transmitted light is collected by the light collection element and detected by the two-dimensional detector. In one embodiment, the observation region is positioned to provide a continuous measurement of composition along the separation axis. Alternatively, light collection element and detector are positioned such that one or more extraction port is periodically rotated into the observation region as the centrifuge rotates. Use of two-dimensional optical imaging allows for the accurate characterization of sample composition along a plurality of separation axes which allows for desirable signal-to-noise ratio averaging that enhances sensitivity.
The intensity of light transmitted by blood or a blood component depends on the concentrations and optical properties of cellular and noncellular components and the optical path length of light through the separation chamber. Accordingly, measurement of a pattern of light intensities transmitted through the separation chamber provides a plurality of measurements of the composition of a selected blood component. Measurement of a two-dimensional distribution of scattered and/or transmitted light intensities in the present invention is beneficial because it provides a method of measuring the purity and/or flux of an extracted, separated fraction, in contrast to conventional one-dimensional optical detection or scanning methods.
In another aspect, the present invention comprises an optical monitoring system capable of measuring the flux and/or composition of one or more cellular blood components exiting an extraction port of the separation chamber, such as an extraction port of an optical cell. In this embodiment, the observation region is positioned on the density centrifuge such that at least one extraction port of the separation chamber is viewable. For example, in one embodiment, at least one extraction port is periodically rotated into the observation region as the separation chamber is rotated about the central rotation axis. In a preferred embodiment, the separation chamber is illuminated in a manner such that light is transmitted through at least one extraction port. As cellular components pass through an extraction port, light is absorbed and/or scattered by a given component. By monitoring the two-dimensional distribution and temporal profile of transmitted and/or scattered light intensities, cellular matter exiting the separation chamber are able to be quantified and type-characterized as a function of time. In an embodiment, the observation region of the present invention is positioned such that a two-dimensional distribution of scattered and/or transmitted light intensities is acquired showing the passage of cellular and non-cellular materials out of the separation chamber, preferably for some applications showing the passage of cellular and non-cellular materials out of an optical cell of a separation chamber. As cellular material absorbs and/or scatters incident light, the flux of cellular material passing through a selected extraction port is determined by measuring the transmitted light area intensity as a function of time. In some instances, for example, larger transmitted and/or scattered light intensities correspond to larger concentrations of cellular material than smaller transmitted and/or scattered light intensities. The present invention includes embodiments wherein at least a portion of the observation region is positioned such that extraction ports in contact with separated fractions corresponding to red blood cells, white cells, platelet enriched plasma and/or plasma are periodically rotated into the observation region.
In another aspect, the present invention comprises an optical monitoring system capable of monitoring the composition of a blood sample prior to blood processing. For example, optical monitoring systems of the present invention generate a two-dimensional distribution of scattered and/or transmitted intensities of light from one or more inlets of a blood processing devise, such as the inlets of a density centrifuge. The levels of light transmitted and/or scattered by a blood sample flowing through the inlet provides real time measurements of important qualities of the incoming blood sample, such as the extent of hemolysis in the blood sample, hematocrit, abundance of lipids in the blood sample and other measurements of blood sample composition. A benefit of this aspect of the invention is that measurements of the composition of a blood sample prior to processing correlates to blood sample and blood component composition measurements taken during and after blood processing to provide a better understanding of a selected blood processing procedure or therapy.
The present invention includes embodiments wherein a plurality of centrifuge operating parameters is measured and analyzed upon acquisition of every two-dimensional distribution of scattered and/or transmitted light intensities. In an embodiment, for example, the present invention comprises an optical monitoring system capable of simultaneously determining the position of at least one phase boundary between at least two optically differentiable blood components, the composition of at least one separated blood component and the flux and/or composition of one or more cellular blood components exiting an extraction port of the separation chamber. In this embodiment, the observation region is positioned on the density centrifuge such that phase boundaries between optically differentiable components, one or more separated components, one or more inlets and at least one extraction port are each viewable upon rotation of the separation chamber about the central rotation axis. An exemplary separation chamber, for example, is designed such that phase boundaries, extraction ports, inlet ports and separated components are readily observable in an image provided by a single two-dimensional distribution of scattered and/or transmitted intensities of light from the separation chamber. This functional aspect of the present invention provides simultaneous monitoring of a plurality of operating conditions of a blood system, which allow correlations between two or more operating parameter to be analyzed and used for accurate device control. Further, methods of the present invention include device control methods wherein a blood processing system is controlled using output signals corresponding to real time measurements of a plurality of operating conditions of a density centrifuge. This functional capability provides improved device control with respect to the control provided by conventional one dimensional scanning or imaging techniques.
Observation regions of the present invention also includes regions other than those selected for viewing separated blood components in the separation chamber. In one embodiment, the observation region includes an identifying region of the blood sample, such as a bar code or other sample designation. This embodiment allows efficient identification and tracking of processed blood products. Alternatively, the observation region includes a region for detecting leaks of blood in the density centrifuge device or an alignment region for detecting improper or proper alignment of the separation chamber before, during or after blood processing. In addition, the present invention can detect spillover of one blood component into the collection port of another blood component. In this context, spillover refers to processes whereby the position of a separated layer in separation chamber changes such that the separated layer contact the orifice of an extraction port corresponding to different separated component.
In another aspect, the present invention comprises a control system for a density centrifuge device. In this embodiment, the optical monitoring system of the present invention is operationally coupled to one or more centrifugation device controllers. In an embodiment, centrifuge device controllers of the present invention receive an output signal from the two-dimensional detector, process the output signal in real time and adjust operating conditions of said centrifugation device to achieve a desired extent of separation and a desired composition of an extracted blood component. In another embodiment comprising a feedback device controller, the device controller and optical monitoring system are operationally coupled in a manner whereby an output signal corresponding to a two-dimensional distribution of scattered and/or transmitted intensities of light from an interface region including one or more phase boundaries and/or one or more extraction ports is sent to a controller capable of adjusting the flow rate of one or more separated blood components out of the separation chamber. In this embodiment, the controller adjusts the flow rates of individual blood components in a manner to selectively adjust the positions of one or more phase boundaries along the separation axis such that a selected extraction port is in fluid communication with a single blood component. Similarly, the present invention includes feedback device controllers, wherein output signals corresponding to a two-dimensional distribution of scattered and/or transmitted light intensities from light from one or more extraction port is sent to a controller capable of adjusting the flow rate of one or more separated blood components from the separation chamber. In this embodiment, the controller adjusts the flow rates of individual blood components in a manner to achieve desired compositions of extracted blood fractions.
In another aspect, the present invention is capable of measuring a two-dimensional distribution of scattered and/or transmitted light intensities comprising a three dimensional image of a region of the separation chamber occupied by one or more blood components, such as a region of an extraction port. In this embodiment, light produced upon illumination of an observation region is collected and detected. In one embodiment, a three dimensional image is generated statistically by modeling the scattering of light by cellular components located in different layers in the region of the separation chamber monitored. Generating a three dimensional image is beneficial because it provides a measurement of the composition of separated blood components along a third axis corresponding to the depth in the separation chamber. This measurement is useful for characterizing the flows of different blood components into the separation chamber and/or through exit ports disposed at different separation chamber depths. In an alternative embodiment, the present invention is capable of measuring a two dimensional distribution of light intensities from fluorescent materials present in the separation chamber. This aspect of the present invention is capable of generating two or three dimensional images from the acquired two-dimensional distributions of fluorescent light intensities. In this embodiment, fluorescence is excited by illumination with an excitation beam. The fluorescence generated is then collected and detected in a manner generating two-dimensional or three-dimensional images. This embodiment is especially useful for monitoring and controlling the separation of fluorescently labeled materials, such as fluorescently labeled cells or blood proteins.
In another embodiment, the present invention provides control systems for centrifuge blood processing of batch samples of blood, preferably whole blood samples or blood samples comprising one or more blood components contained in containers or bags. Exemplary methods and devices for processing batch samples are described in U.S. application Ser. No. 10/413,890. In one embodiment, one or more blood samples residing in an initial fluid containment container are connected to the rotors of a density centrifuge in a manner allowing rotation of the blood samples about a central rotation axis. Rotation of the centrifuge generates a centrifugal force which separates components of the sample according to density along rotating separation axes oriented orthogonal to the central rotation axis. Once the blood sample undergoes separation, discrete components are sequentially extracted out of the initial fluid containment container via one or more outlet ports operationally connected to a plurality of physically separated fluid-receiving containers. Discrete components are extracted via pumping or by the introduction of an inert fluid which is capable of forcing the fractionated sample to exit the fluid containment container. In a preferred embodiment, the present invention provides a means of monitoring and controlling the flow rates and the fluid paths of blood components to selected fluid-receiving containers corresponding to extracted components.
In one embodiment, the optical monitoring and control systems of the present invention is operationally coupled to a batch sample centrifuge in a manner such that phase boundaries between optically differentiable materials, purity and composition of extracted components and the flux of extracted components is monitored during processing in real time. Further, the present invention provides a means of controlling the withdrawal of separated blood components such that the discrete fractions can be separately collected in separate fluid-receiving containers. For example, two-dimensional distributions of scattered and/or transmitted light intensities comprising images of the rotating initial fluid containment container is used to select pumping rates out of the initial fluid containment container or inert fluid flow rates into the fluid containment container in a manner ensuring that only a selected component is directed to a selected fluid-receiving container. In a preferred embodiment, the monitoring system of the present invention is capable of monitoring the change in container of a given component as it is extracted by measuring two-dimensional distributions of scattered and/or transmitted light intensities of light from the separation chamber corresponding to phase boundaries between optically differentiable components or corresponding to one or more extraction ports. A optical monitoring and control system of the present invention is also capable of switching the fluid-receiving container in fluid communication with the initial fluid containment container upon substantially complete extraction of a selected component. Alternatively, an optical monitoring and control system of the present invention is capable of adjusting the pumping rate of a component being extracted to ensure that an adjacent component is not collected in the same fluid-receiving container. In a preferred embodiment, the optical monitoring and control system of the present invention is capable of generating an output signal triggering a multi-channel valve or clamp to divert the flow of sample corresponding to an adjacent component into separate fluid-receiving container.
Collection and processing two-dimensional distributions of scattered and/or transmitted light intensities corresponding to an image of an observation region have a number of advantages over conventional one-dimensional optical monitoring or scanning methods applied to centrifugation of blood samples. First, two-dimensional distributions of scattered and/or transmitted light intensities comprising images of an observation region provide a substantially improved means for discriminating between optically differentiable blood components and measuring the position of phase boundaries between these components as compared to one-dimensional measurements. One-dimensional optical scanning or monitoring provides a single profile of light intensities corresponding to a single optical axis. In contrast, two-dimensional distributions of scattered and/or transmitted light intensities provided by the present invention comprise a pattern of light intensities corresponding to a plurality of optical axes. Therefore, each two-dimensional distribution of scattered and/or transmitted light intensities provides a plurality of multiple measurements of the positions of phase boundaries along the separation axes. Averaging light intensities from each optical axis monitored improves signal-to-noise ratios over measurements derived from one-dimensional measurements by a factor of approximately 10. The improvement in signal-to-noise ratio observed in the present invention provides more reproducible measurements of the relative positions of phase boundaries and provides more accurate calibration of absolute phase boundary positions. In addition, the improved signal-to-noise ratio provides the present systems the capability of providing direct measurements of the composition and purity of any portion of a blood sample, particularly the composition and purity of a given separate blood component, in contrast to conventional one-dimensional scanning and imaging methods.
Second, measurement of light intensities over a two-dimensional area reduces problems arising from heterogeneity in the separated blood components. The various cellular components of blood exhibit distributions of cell types, sizes, shapes and optical properties, such as absorption constants and scattering coefficients. As a result, profiles of scattered and/or transmitted light intensities at different points along the separation axes show a substantial degree of variability for different regions of the separation chamber. Collecting light associated with a plurality of optical axes allows the effects of heterogeneity in the various cellular components to be treated statistically. In one aspect of the present invention, each two-dimensional distribution of scattered and/or transmitted light intensities is statistically analyzed to provide a measure of the average optical properties of a given blood component. Further, the devices and methods of the present invention provide a quantitative measurement of the uncertainties associated with compositions of blood components disposed along the separation chamber, which allows accurate characterization of the reproducibility in the purity levels of extracted components achieved. The ability to characterize uncertainty in the purity levels achieved allows for the quantitative assessment of quality assurance useful for establishing regulatory approval. Third, collection and detection of scattered light corresponding to a two-dimensional area allows for direct measurements of the composition and flux of cellular materials out of an extraction port of a separation chamber. Cellular components of blood undergoing separation are extracted from a separation chamber via extraction ports, which comprise tubes extending selected distances along the separation axis. The flux of cellular components through the extraction port is not spatially uniform.
Rather, the flow of cellular components routinely exhibits substantial spatial inhomogeniety. Therefore, to accurately measure the flux of cellular material exiting the separation chamber at a given time, a profile of transmitted light intensities across an area perpendicular to the flow of exiting cellular components is required. Two-dimensional distributions of scattered and/or transmitted light intensities provide measurements corresponding to a plurality of axes perpendicular to the flow of material out of the separation chamber. This provides a sensitive means of measuring fluxes and compositions of cellular material out of the separation chamber. Two dimensional detection is critical for characterizing fluxes and compositions of cellular material exiting the separation chamber because such material are typically inhomogeneously dispersed through an extraction port.
Fourth, detection of light corresponding to a two-dimensional area also provides optical systems capable of simultaneously monitoring a plurality of operating conditions important to controlling blood processing. In contrast to conventional optical monitoring techniques, the methods and devices of the present invention are capable of multifunctional operation because the measured two-dimensional distribution of scattered and/or transmitted light intensities correspond to a plurality of different optical axes. In the present invention reference to multifunctional operation relates to the ability of an optical monitoring system to monitor and/or control a plurality of operating or experimental conditions important to optimal operation of a density centrifuge. The ability to simultaneously generate and analyze a plurality of measurements from a single two-dimensional distribution of scattered and/or transmitted light intensities is beneficial in the present invention because it allows diverse measurements to be correlated and analyzed in combination to provide a greater understanding of the operating conditions of the centrifuge during blood processing. For example, optical methods of the present invention are capable of simultaneously monitoring the position of phase boundaries, the composition of extracted components, the fluxes of components out extraction ports, the identity of blood samples, the presence of leaks of blood components out of the separation chamber or any combination of these. In addition, the ability to selectively adjust the position and size of the observation region expands the functional capabilities of the optical monitoring system of the present invention. Optical monitoring and control systems capable of multifunctional operation are beneficial because they substantially reduce the time, effort and expense associated with personnel overseeing a blood processing device. In addition, the devices and methods of the present information provide highly reproducible separation conditions capable of generating separated blood components having well-characterized and highly reproducible compositions namely purities. Furthermore, multifunctional monitoring and control systems are capable of dealing with rapid changes in blood separation conditions and are well designed for overseeing processing of blood samples having atypical compositions, such as the samples encountered during therapeutic procedures.
In another aspect, the present invention provides optical monitoring and control systems for blood processing utilizing separation methods other than pure density centrifugation, such as separation on the basis of shape, size, sedimentation velocity, diffusion rate, surface chemistry characteristics or any combination of these techniques. For example, the present invention is capable of monitoring and controlling blood processing via multiple stage processing. In a preferred embodiment of multiple stage processing, a blood sample is first fractionated into discrete blood components by density centrifugation. Next, one or more selected blood components are extracted from the density centrifuge and further separated by shape and size filtration, centrifugal elutriation, affinity chromatography or any combination of these methods. In this embodiment, optical monitoring and control systems of the present invention control the extent of separation achieved in both stages.
In a preferred embodiment, two stage blood processing is achieved by a combination of density centrifugation and centrifugal elutriation methods. Exemplary methods and devices for blood processing by centrifugal elutriation are described in U.S. Pat. No. 6,334,842. In a preferred embodiment, a blood sample is separated into components via density centrifugation in a first stage and a selected blood component or plurality of blood components is extracted and subjected to further processing via centrifugal elutriation. In a preferred embodiment, the selected component is introduced into a flow of liquid elutriation buffer and passed into a funnel-shaped separation chamber located in a spinning centrifuge. As the liquid buffer flows through the separation chamber, the liquid sweeps smaller sized, slower sedimenting cells toward an elutriation boundary within the chamber. Larger, faster-sedimenting cells, however, migrate toward an area of the chamber having the greatest centrifugal force. By selecting the proper fluid flow rates through the funnel-shaped separation chamber, faster sedimenting cells and slower-sedimenting cells are separately extracted from the separation chamber and subsequently collected. Therefore, the combination of density centrifugation and centrifugal elutriation provides a method of separating blood components based on both density and sedimentation velocity.
The methods, devices and device components of the present invention are capable of monitoring and controlling multiple stage blood processing. Particularly, the optical monitoring and control systems of the present invention are capable of generating two-dimensional distribution of scattered and/or transmitted light intensities comprising images of blood separation in first and second stages of a blood processing device. First, the monitoring system of the present invention is capable of measuring two-dimensional distributions of scattered and/or transmitted intensities of light from a separation chamber of the density centrifuge, which characterize the composition, purity and extraction rate of the blood component selected for additional processing via centrifugal elutriation. Further, in one aspect of the present invention two-dimensional distributions of scattered and/or transmitted light intensities are used to optimize separation and extraction conditions in the first stage to achieve a desired composition for additional processing in the second stage. In one embodiment, for example, phase boundary positions in the first stage are selected and maintained in a manner minimizing the presence of red blood cells and/or white blood cells in a platelet-containing blood component selected for additional processing in the second stage. Second, the optical monitoring and control systems of the present invention are capable of measuring two-dimensional distributions of scattered and/or transmitted light intensities comprising images of the elutriation chamber itself as it is rotated about the central axis of a centrifuge. Two-dimensional distributions of scattered and/or transmitted light intensities of light from the elutriation chamber provide direct measurements of the composition of the blood component undergoing additional processing, which can be compared to measurements acquired by monitoring separation achieved in the first stage to evaluate the degree of separation achieved during extraction. For example, the brightness or color of a two-dimensional distribution of scattered and/or transmitted light intensities of light from an elutriation chamber provide measurements of the composition of a blood component selected for further processing, for example the abundance of red blood cells in the elutriation chamber.
In addition, two-dimensional distributions of scattered and/or transmitted light intensities generated by the present invention provide direct measurements of the composition, and flux of sub-components separated in the second stage. Characterization of the composition of a selected subcomponent is beneficial because it ensures that the collected subcomponent is adequate for use in transfusion or infusion therapies. For example, the methods of the present invention are useful for leukoreduction methods by optically characterizing platelet-containing sub-components to ensure levels of white blood cells are low enough as to avoid complication upon infusion related to undesirable immune responses and viral transmission. Alternatively, the methods of the present invention are useful in immunotherapy for characterizing extracted white blood cell-containing sub-components and to optimizing separation conditions in a second stage to minimize the levels of red blood cells and platelets in the purified sub-component or to collect a particular white blood cell-type.
The methods, devices and device components of the present invention are useful for monitoring and controlling blood processing other than separation of blood into components. Exemplary processing applications capable of being monitored and controlled by the present invention include, but are not limited to, blood component washing, pathogen reduction and pathogen removal, red blood cell deglycerolization and the addition of blood components and/or blood processing agents to blood samples.
In another aspect, the present invention provides a method of detecting the occurrence and extent of hemolysis of red blood cells during blood processing, particularly centrifugation. Hemolysis can occur during blood processing when motion of the blood sample results in a degradation of red blood cells leading to the release of hemoglobin. Upon its release, hemoglobin migrates to less dense blood components, such as the plasma containing component. The release and migration of free hemoglobin to lower density blood components is able to be optically monitored in the present invention because hemoglobin absorbs light strongly in the visible region of the spectrum, particularly over the wavelength range of about 500 nm to about 600 nm, and thus, decreases detected light intensities. Accordingly, measured two-dimensional distributions of scattered and/or transmitted light intensities can be used to determine light absorption over this wavelength range to characterize the extent of hemolysis during blood processing. In these measurements, large absorption over the wavelength range of 500 nm to 600 nm corresponds to separation conditions resulting in substantial hemolysis. Further, in one embodiment such measurements are used as the basis of control signals to optimize the flow conditions in a blood processing device to minimize the occurrence of hemolysis. In a one embodiment, the lower density blood component is illuminated with both green light and red light, and transmitted light, scattered light, or both, is collected and detected corresponding to each illumination color. A comparison of the intensities of scattered and/or transmitted light corresponding to each illumination color provides an accurate measurement of the extent of hemolysis in the sample.
In another aspect, the present invention provides methods of monitoring and controlling a density centrifuge capable of separating at least two optically differentiable components of a fluid and having a separation chamber rotating about a central rotation axis wherein said components in the centrifuge separation chamber separate along a separation axes which rotate about the central rotation axis, comprising the steps of: (1) illuminating the density centrifuge with an incident light beam provided by a light source; (2) collecting light from a observation region on the density centrifuge and directing said light onto a two-dimensional detector; (3) positioning at least a portion of said observation region such that phase boundaries are viewable; and (4) detecting said light with said two-dimensional detector, which generates a two-dimensional distribution of scattered and/or transmitted intensities of light from of said observation region; (5) measuring the position of at least one phase boundary between said components along said separation axis. Optionally, the methods of the present invention further comprise the step of measuring the composition of a component exiting the separation chamber via an extraction port. Optionally, the methods of the present invention also include the step of adjusting the operating conditions of said centrifugation device to achieve substantial separation of said optically differentiable components.
The invention is further illustrated by the following description, examples, drawings and claims.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a schematic drawing showing an optical monitoring and control system of the present invention.
FIG. 2 is a schematic drawing showing a side view of a light collection element and two-dimensional detector useable in the present invention.
FIG. 3 is a schematic drawing showing a front cut-away view of a light collection element and two-dimensional detector useable in the present invention.
FIG. 4 is a schematic drawing showing a top plan view of a mounting configuration providing for selective adjustment of the position of the light collection element and two-dimensional detector.
FIG. 5 is a schematic drawing showing observation regions of monitoring systems of the present invention.
FIG. 6 is a top plan view of an optical cell of a separation chamber showing an expanded region illustrated in FIGS. 6A and 6B. FIGS. 6A and 6B show schematics of images generated by the methods of the present invention of the expanded region shown in FIG. 6 having a human blood sample therein separated into blood components. The images in FIGS. 6A and 6B illustrate the ability of the methods and devices of the present invention to monitor and control the position of phase boundaries between separated blood components. In FIGS. 6A and 6B, triangles schematically represent white blood cells and platelets, circles schematically represent red blood cells and areas having lines schematically represent plasma.
FIG. 7 shows images of the rotating separation chamber of a density centrifuge generated by the methods of the present invention. The image in FIG. 7 includes a phase boundary monitoring region and a white blood cell extraction port monitoring region. Analysis of the image in FIG. 7 provides a measurement of the composition and flux of cellular material out of the separation chamber. In FIG. 7, triangles schematically represent white blood cells and platelets, circles represent red blood cells and areas having lines represent plasma.
FIG. 8 shows the temporal behavior of the measured phase boundary positions (bottom two curves) and transmitted light intensities through the extraction port monitoring region (top two curves) during white blood cell collection. FIG. 8A show corresponding 50 point moving averages. Solid diamonds (designated as RBC Pixels) correspond to the position of the phase boundary between the red blood cell containing component and the buffy coat layer, open squares (designated as Platelet Pixels) correspond to the position of the phase boundary between the platelet containing component and the buffy coat layer, solid triangles (designated as Extraction Port Tool #1) correspond to median transmitted intensities through a first flux monitoring region and X markers (designated as Extraction Port Tool #2) correspond to median transmitted intensities through a second flux monitoring region.
FIG. 9 shows a series of plots of the observed white blood cell concentrations as a function of the median intensity of light transmitted through the second flux monitoring region (X markers, + markers and − markers) corresponding to the rotational velocities (RPM) indicated in the legend. Also shown in FIG. 9, are plots of the hematocrit of the extracted material as a function of the median intensity of light transmitted through the second flux monitoring region (diamond markers, square markers and triangle markers) corresponding to the rotational velocities (RPM) indicated in the legend.
FIG. 10 shows a plot of the concentration of white blood cells in the extracted material as a function of the position of the phase boundary (in terms of pixel height of the collected image) between the red blood cell containing component and the buffy coat layer corresponding to the rotational velocities (RPM) indicated in the legend.
FIG. 11 shows a schematic of an exemplary master-smart slave control system of the present invention capable of controlling blood processing.
FIG. 12 provides a schematic flow diagram illustrating an automated, computer controlled process control system for a density centrifuge blood processing device.
FIG. 13 is a schematic diagram showing exemplary Control Driver and APC Sub-System architectural relationships useful in the methods of the present invention.
FIG. 14 is a schematic diagram showing exemplary Procedure Control and APC Sub-System architectural relationships useful in methods of the present invention.
FIG. 15 shows exemplary architectural relationships of the APC Executive with the APC Driver, Image Data List Container, and the APC components within the Control Sub-System useful in the methods of the present invention.
FIG. 16 is a schematic diagram providing a state chart for the image data analyzer task.
FIG. 17 shows an exemplary architecture of the APC Driver component of the present invention.
FIG. 18 shows an exemplary high level state diagram for a APC Driver task useful for the methods of the present invention.
FIG. 19 shows an exemplary architecture of a APC Image Processing Engine component of the present invention.
FIG. 20 provides an exemplary state chart for an image analyzer task useful in the methods of the present invention.
FIG. 21A provides a schematic diagram of a rotated side view of an optical cell of the present invention useful for monitoring blood processing via density centrifugation. FIG. 21B provides a cross sectional view of an exemplary extraction port design of the present invention. FIG. 21C provides a cross sectional view of an alternative extraction port design of the present invention, wherein first extraction port and second extraction port each have axial bores having a rectangular cross sectional profile.
FIG. 22 is a top view of an optical monitoring and control system of the present invention well suited for blood processing via density centrifugation.
FIG. 23 is a cut away view corresponding to cut away axis 1200 indicated in FIG. 22.
FIG. 24 is a side view of the optical monitoring and control system illustrated in FIGS. 22 and 23.
FIG. 25 provides a schematic diagram of an exploded, side view of a bottom pulsed LED source useful in the methods and devices of the present invention.
FIG. 26 shows a functional flow diagram representing a method of synchronizing light pulses generated by top and bottom pulsed LED light sources trigger and trigger delay settings.
FIG. 27 provides plots of measurements of the white blood cell concentration (square markers) and hematocrit (diamond markers) of a separated blood component passing through an extraction port as function of the measured average intensity of light transmitted through the extraction port.
DETAILED DESCRIPTION OF THE INVENTION
Referring to the drawings, like numerals indicate like elements and the same number appearing in more than one drawing refers to the same element. In addition, hereinafter, the following definitions apply:
The terms “light” and “electromagnetic radiation” are used synonymously and refer to waves of electric and magnetic fields that also exhibit particle-like behavior. Light useful for the methods of the present invention includes gamma rays, X-rays, ultraviolet light, visible light, infrared light, microwaves, radio waves or any combination of these. “Depth of field” refers to the zone of acceptable sharpness in a picture and/or image extending in front of and behind the plane of the subject. Depth of field may by quantitatively characterized as the range of distances reproduced in a picture and/or image over which the image is not unacceptably less sharp than the sharpest part of the image. The term “depth of field” is intended to be interpreted consistently with the mean of this term as understood by those having skill in the art. A light collection element may be characterized in terms of its depth of field.
“Optically differentiable” refers to differences in the optical characteristics of two or more illuminated materials. Optically differentiable materials can have different absorption coefficients, extinction coefficients, scattering cross sections, fluorescence excitation wavelengths, phosphorescence excitation wavelengths, emission wavelengths or any combinations of these characteristics. As the optical characteristics of most materials depend on wavelength, materials can be optically differentiable when illuminated by light having a selected wavelength range. Exemplary optically differentiable materials useable in the present invention include, but are not limited to, erythrocytes, eosinophils, basophils, monocytes, lymphocytes, granulocytes, platelets (thrombocytes), plasma proteins, and plasma. Exemplary optically differentiable materials further include the materials comprising a blood processing device or blood sample container, such as polymeric materials such as plastics, metals, and glass.
“Flux of cellular material exiting the separation chamber” refers to the amount of cells, such as erythrocytes, leukocytes, thrombocytes or any combination of these, which cross a defining area, such as the cross-sectional area of an extraction port of blood processing device, such as a density centrifuge, elutriation separation chamber or filtration separation device, per unit time. Flux of cellular material can be expressed in units of: (number of cells) cm −2 s −1 .
“Optical communication” refers to the orientation of two or more elements such that light is capable of propagating from one element to another element. Elements can be in optical communication via one or more additional elements such as reflectors, lenses, fiber optic couplers, wave guides or any combinations of these. In one embodiment of the present invention, one or more light sources and a light collection element can be positioned in optical communication with an observation region on a blood processing device, such as a density centrifuge. In this embodiment, at least a portion of light from one or both of the light sources is directed onto an observation region and the light collection element is positioned such that it is capable of collecting at least a portion of light scattered transmitted, or both from the observation region.
“Light collection element” refers to a device or device component which collects light and distributes the collected light in a desired way. Light collection elements useable in the present invention are capable of collecting at least a portion of transmitted light, scattered light or both generated upon illumination of an observation region on a blood processing device. Exemplary light collection elements of the present invention are capable of collecting light in a manner generating an image of an observation region on a two dimensional detector. Light collection elements of the present invention include, but are not limited to, fixed focus lenses, spherical lenses, cylindrical lenses, aspheric lenses, wide angle lenses, zoom lenses, concave lenses, convex lenses, biconcave lenses, biconvex lenses, lens systems comprising a plurality of lenses, wave guides, fiber optic couplers, reflectors, spherical mirrors, aspherical mirrors, prisms, apertures, lenses, or any combination or equivalents of these. Light collection elements of the present invention are capable of directing collected light onto another optical device or device component, such as a two-dimensional detector. Light collection elements include at least one lens system having a selectively adjustable field of view and/or focal length. Light collection elements can be translatable along a detection axis, which is perpendicular to a central rotation axis.
“Field of view” refers to the angular distribution of light rays which are collected and detected by an optical detection system, such as a light collection element in optical communication with a two dimensional detector. The field of view of a two dimensional imaging system of the present invention is the portion of an illuminated object or plurality of objects which is represented in a two dimensional image. Optical detection systems of the present invention can have a fixed field of view or a field of view which is selectively adjustable.
“Blood processing” refers to the manipulation of a blood sample or component thereof, to realize a change in composition. Blood processing includes methods of separating blood or a component thereof into components or subcomponents, leukoreduction, pathogen inactivation, blood filtering, oxygenating blood and blood components, dialysis, blood purification or clearing, pathogen removal, blood and blood component warming, blood component washing, and red blood cell deglycerolization. The present invention provides improved methods of blood processing wherein a blood sample or component thereof is separated into components or subcomponents on the basis of density, size, diffusion rate, sedimentation velocity, surface chemistry properties or combinations of these characteristics.
“Observation region” refers to an illuminated portion of an object or plurality of objects which generates transmitted light, scattered light or both at least a portion of which that is collected by a light collection element and detected by a two-dimensional detector. In preferred embodiments of the present invention, the observation region is positioned on a blood processing device, component of a blood processing device, such as an optical cell, or a blood sample container. The size and position of the observation region is determined by the field of view of the light collection element, the position of the light collection element from the blood processing device, the area of the two-dimensional detector and the position of the two-dimensional detector with respect to the light collection element. In an embodiment, the size, shape and position of the observation region is selectively adjustable by controlling the position of the light collection element with respect to the blood processing device and the field of view of the light collection element. In an embodiment of the present invention, one or more phase boundaries between optically differentiable components are viewable in the observation region. In another preferred embodiment, at least one separated component is viewable in the observation region. In another preferred embodiment, at least one extraction port is viewable in the observation region.
“Interface region” refers to a region of the a blood separation device wherein two or more optically differentiable phases are viewable. For example, in one embodiment the interface area is defined by a region of the separation chamber wherein the phase boundary between a red blood cell containing component and a plasma containing component is viewable. In another embodiment, the interface area is defined by a region of the separation chamber wherein the phase boundary between a red blood cell containing component and a mixed-phase white blood cell and platelet containing component and the phase boundary between the mixed-phase white blood cells and platelet containing component and plasma containing component are viewable. In another embodiment, the phase boundary between a white blood cell containing component and a platelet containing component are viewable. In the present invention, a two-dimensional distribution of scattered and/or transmitted light intensities of light from an interface region provides a measurement of the position of one or more boundary layers along a plurality of separation axes. In an exemplary embodiment, the interface region is an optical cell of a separation chamber.
“Composition-monitoring region” refers to portion of a blood processing device occupied by at least one separated phase. For example, the composition-monitoring region can be defined by a region of a separation chamber in a density centrifuge wherein light is transmitted through one or more discrete phase in the separation chamber upon illumination by an incident light beam. As the transmission of light through a separated compound depends on identity and concentration of cellular and non-cellular material, monitoring scattered light, transmitted light, or both, from a composition-monitoring region provides a measurement of the identity, concentration, cell type, purity of at least one component or any combination of these. In an exemplary embodiment, the composition monitoring region is an extraction port in an optical cell of a separation chamber.
“Blood sample” and “blood” are used synonymously to refer to whole blood,