Patent Publication Number: US-2020278341-A1

Title: Method and apparatus for performing hematologic analysis using an array-imaging system for imaging and analysis of a centrifuged analysis tube

Description:
This application is a divisional of U.S. patent application Ser. No. 15/207,074 filed Jul. 11, 2016, which is a continuation of U.S. patent application Ser. No. 13/016,347 filed Jan. 28, 2011, which claims the benefit of U.S. Provisional Patent Application No. 61/305,449 filed Feb. 17, 2010 and U.S. Provisional Patent Application No. 61/351,138 filed Jun. 3, 2010, each of which applications is hereby incorporated by reference in its entirety. 
    
    
     BACKGROUND INFORMATION 
     U.S. Pat. Nos. 4,027,660; 4,091,659; 4,137,755; 4,209,226; 4,558,947; 4,683,579; 5,132,087; 5,888,184; and 6,441,890 describe methods and apparatus for hematological analysis using a capillary tube and a space occupying insert that floats on the centrifuged red blood cells thereby expanding the surrounding buffy coat and permitting the measurement and quantization of the blood&#39;s layers. This method permits the determination of a compete blood count (CBC) consisting of hematocrit, a hemoglobin determination, a total white blood cell count with the latter presented as a total and percent granulocytes and total and percent lymphocytes plus monocytes, as well as a platelet count and a mean red cell hemoglobin concentration. It is widely used through the world for performing point of care CBC in human and veterinary medicine. The device, formerly manufactured and sold by Becton Dickinson, Inc. of New Jersey U.S.A. is now manufactured and sold by QBC Diagnostics, Inc., of Pennsylvania, U.S.A. The apparatus is sold under the trademark of QBC® hematology. The capillary tubes are referred to in the industry as “QBC® tubes”. 
     The QBC® hematology system includes a number of different complex instruments for reading the QBC® tubes, each of which has an illumination system, a power source, an imaging and optical system, a microprocessor, and a display. These devices can cost anywhere from several hundred to many thousands of U.S. dollars. The current versions of both the stand-alone reader and the integral reader-centrifuge (QBC® STAR reader) provide for a linear scan of the tube, either while it is stationary in the case of the stand-alone reader or while the centrifuge is in motion, as is the case with the QBC® STAR reader. In both cases, the linear scan is limited to scanning a single axially extending line scan of the tube, which evaluates only a thin stripe of the area of interest within the tube. Because this method of scanning can only scan a thin stripe of the area of interest at a given time, it is necessary to take multiple axially extending scans taken at different circumferential positions of the tube to determine which of the scans can be used for analytical purposes. By looking at several different scans, each taken at a different circumferential position, it is possible to ascertain whether any particular scan is representative of the sample or if it contains an unrepresentative anomaly. Also, because of the narrow scan, the mechanical and optical alignment of the instrument must be held to a very high tolerance, which also increases the cost of the device. 
     This is particularly true in the case of the QBC® STAR reader, because the QBC® tube is read while the centrifuge is in motion, necessitating an elaborate timing system to ensure that illumination occurs exactly when the tube is in position under the linear scanning device (e.g., CCD scanner). Another, related problem is the need to provide elaborate vibration damping so that the relative tube and reader position be maintained during this process. 
     These considerations force the analysis tube readers to have a relatively high price, which limits the market size for the QBC® hematology system because health care providers are reluctant and/or unable to make the requisite equipment investment when the equipment is only used for a few tests per day. In those instances when the point of care giver does not have the analysis equipment, the patient is subjected to the significant inconvenience, harm and expense of having to go to a private laboratory and having to wait often several days to get the result. The lack of an analysis device also makes the physician&#39;s job more difficult by precluding immediate results at the point of care. Additionally, regulatory requirements of the United States require that the providers of the test be subject to regulatory supervision under the CLIA (Clinical Laboratory Improvement Act) laws. 
     What is needed, therefore, is a simple, inexpensive, robust method for reading the centrifuged blood sample at the point of care with immediate availability of results while the health care providers are still with the patient. In addition, a method and device are needed that can provide accuracy results and methodological adherence to proper analytic techniques, as well as quality control measures, particularly those that will permit CLIA waiving, which is subject to less burdensome regulations. 
     SUMMARY OF THE INVENTION 
     According to one aspect of the present invention, a device for analyzing a hematologic sample centrifuged within a capillary tube is provided. The tube has an internal compartment with a radial width and an axial length and a float disposed within the tube. The device includes a tube holder, a sample imaging device, a processor, and a sample data display. The sample imaging device is operable to create a digital image of the sample within a region of the tube. The region is defined by substantially all of the radial width and axial length of the sample residing within the internal cavity of the tube in the region where the float resides after centrifugation. The sample imaging device is operable to produce signals representative of the image. The processor is adapted to produce information relating to bands of interest within the image based on the signals from the sample imaging device. The sample data display is adapted to display the results therefrom and/or a digital image of the sample within the region. 
     According to another aspect of the present invention, a method of analyzing a hematologic sample deposited within a capillary tube is provided. The tube has an internal cavity with a radial width and an axial length, and a float disposed within the tube. The method includes the steps of: a) centrifuging the sample to create constituent bands within the sample disposed in the tube; b) creating an image of a region of the centrifuged sample, which region is defined by substantially all of the radial width and axial length of the sample residing within the internal cavity of the tube in a region where the float resides after centrifugation; c) determining a position for one or more band boundaries using the image; and d) producing analysis results based on the determined band boundaries. 
     Advantages associated with the present analysis device include the provision of a less expensive, and easier to manufacture, analysis device. The imaging of substantially all of the radial width and a significant portion of the axial length of a centrifuged sample within a capillary tube eliminates many problems associated with narrow linear array sensing. For example, prior art linear array sensing is susceptible to circumferentially located bandwidth anomalies; e.g., if the bandwidth at a particular circumferential position is irregularly too small or too big, data based on that band width will be inaccurate. For this reason, the prior art devices take multiple linear array sensings at non-contiguous circumferential positions and average those sensings, or otherwise compare them to one another for accuracy. The prior art devices, therefore, require hardware that can rotate one or both of the linear sensing array and the sample. The hardware must also be able to provide very accurate mechanical and optical alignment of the instrument relative to the sample, and in the case of a dynamic sensing device like the QBC® STAR reader, also provide elaborate imaging controls and vibration damping. The present device also provides significant quality control mechanisms. 
     On the other hand, the prior art linear imaging had the advantage of minimal geometric distortion. Since all prior art imaging data was in the form of a narrow linear segment taken at a right angle to the tube as it was scanned, each band position was exactly related to its digital representation. In the case of the image array as used in the present device, in which the tube is positioned some distance from the imaging lens and camera, the bands in the tube are foreshortened in proportion to their distance from the center of the optical axis, and the sides of the tube are particularly affected by this effect, sometimes making them appear crescent shaped. This geometric distortion, in addition to any other distortions from the lens, is preferably accounted for in order to enhance the accuracy of the results. For example, the geometric distortion can be accounted for by using a correction table which accounts for each pixel, or regions in the image. The correction table can be used to re-map the image so that the image positions correctly correspond to the actual locations on the tube surface. This type of correction table can be automatically generated by imaging and analyzing a known ‘calibration’ standard or if only geometric distortion is involved, the corrections can be simply calculated based on the known distances involved. Alternatively, the geometric distortion can be accounted for by correcting the band lengths following their preliminary measurement. 
     The foregoing and other objects, features and advantages of the present invention will become more apparent in light of the following drawings and detailed description of the present invention. 
    
    
     
       BRIEF DESCRIPTION OF THE DRAWINGS 
         FIG. 1  is a schematic diagram of the present invention hematology analysis device. 
         FIG. 2  is a schematic diagram of a capillary tube. 
         FIG. 3  is an enlarged partial view of a tube such as that shown in  FIG. 2 . 
     
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
     Referring to  FIGS. 1-3 , a blood sample for analysis within the QBC® hematology system is typically obtained either from a venous or capillary sample, centrifuged in a simple, small dedicated centrifuge which may be either battery powered or AC powered. U.S. Pat. Nos. 4,027,660; 4,683,579 and 6,441,890, each of which is hereby incorporated by reference in its entirety, describe methods and apparatus for hematological analysis using a capillary tube and a space occupying insert that floats on the centrifuged red blood cells thereby expanding the surrounding buffy coat and permitting the measurement and quantization of the blood&#39;s layers. The capillary tube  10  includes a body that extends between a closed bottom  12  and an open top  14 . In some embodiments, the “closed bottom” may be vented to allow the escape of gas. The open top  14  provides access to an internal cavity  16  that has a radial width  18  and an axially extending length  20 . In those embodiments where the tube  10  is cylindrical, the radial width  18  is the inner diameter of the tube  10 . The present invention is not limited to use with any particular type of capillary tube. U.S. Pat. No. 4,027,660, for example, describes a QBC® style capillary tube operable to contain a fluid sample and a volume occupying mass  22  (hereinafter referred to as a “float”), and the information available by virtue of the relative positioning of the float  22  within the sample after centrifugation. U.S. Pat. No. 6,444,436 describes a different style of capillary tube that can be used with the present invention; e.g., one having a polynomial (e.g., rectilinear) cross-sectional geometry.  FIGS. 2 and 3  of the present application diagrammatically illustrate a capillary tube  10  with a sample and a float  22  disposed in the internal cavity  16  of the tube  10 . The centrifuged sample disposed in the tube  10  illustrates the constituent bands  24  ( 24   a ,  24   b ,  24   c ,  24   d ,  24   e ) and the band boundaries  25  ( 25   a ,  25   b ,  25   c ,  25   d ) therebetween. U.S. Pat. Nos. 4,683,579 and 6,441,890 describe automated devices for reading the centrifuged sample by way of an axially extending linear scan of a limited portion of the sample disposed within the QBC® tube, which limited linear portion is disposed at a particular circumferential position of the tube  10 . 
     The present invention analysis device operates with a capillary tube  10  such as those provided within a QBC® hematology system; i.e., a tube  10  filled with a sample that has been centrifuged to produce the separated constituent layers (i.e., “bands”)  24  within the sample. One embodiment of the present analysis device  28  includes a housing  30  containing a tube holder  32 , a sample imaging device  34 , a processor  36  adapted to produce information relating to bands  24  of interest within the image based on the signals from the sample imaging device  34 , a sample data display  38 , and may include an operator input device  40  that enables the operator to enter relevant patient information. 
     In some embodiments, the analysis device  28  further includes a centrifuge  42  with a platen  44  configured to hold one or more capillary tubes  10  in a position where the tubes  10  extend radially outward from a central axis. In these embodiments, the analysis device  28  can perform both the centrifugation and the image analysis. The centrifuge  42  is operable to centrifugally spin the tube  10  containing the sample about the central axis at speeds sufficient to create constituent layer separation within the sample disposed in the tube  10 . In these embodiments, the platen  44  is an example of a tube holder  32 . In other embodiments, the tube holder  32  may be independent of the centrifuge  42 . 
     The sample imaging device  34  includes a digital camera operable to image substantially all of the radial width  18  and axial length  20  of the sample residing within the internal cavity  16  of the tube  10  in the region  46  (see  FIG. 3 ) where the float  22  resides after centrifugation in a single image, and to produce signals representative of the image. In the preferred embodiments, the sample imaging device  34  is operable to image a region  48  comprising substantially all of the radial width  18  and axial length  20  of the sample within the tube  10  in a single image, and to produce signals representative of the image. Alternately, two or more cameras can be used to image separate portions of the tube  10 , which portions are contiguous with one another. The images of the contiguous regions can be subsequently combined and analyzed or are separately analyzed. Either the digital camera itself, or an independent light source within the sample imaging device  34 , provides sufficient lighting so that bands  24  of interest within the centrifuged sample may be differentiated within the sample image. The optical resolution of the camera must be sufficient to provide adequate clarity within the image for the analysis at hand; e.g., to differentiate bands  24  of interest. As indicated above, the sample imaging device  34  may be incorporated into a QBC® tube type reader, or may be an independent device (e.g., a portable digital camera, a cell phone camera, etc.) configured for use with such a reader. An example of an acceptable digital camera is a Bayer-type matrix color camera. If, for example, a standard Aptina® five megapixel color camera chip with a frame width of 2592 pixels is used, it can produce a theoretical image resolution of 0.02 mm, which is acceptable for most analyses. If a color camera is used, color filters and different illumination types are likely not required. A grey scaled camera may also be used because the separated buffy coat layers have different light scattering properties and may therefore be detected using a black and white camera, although this measurement is less robust and requires more controlled illumination. The sample imaging device  34  may be described as an “area-array imaging device” because it images substantially all of the radial width  18  and axial length  20  of the sample within the interior cavity  16 . If a plurality of cameras is used within the present sample imaging device  34 , the images they produce are contiguous with one another thereby permitting the plurality of images to be combined into a single representative image. The linear scan devices of the prior art, in contrast, are limited to producing narrow linear segments that do not extend across the full radial width  18 , and the circumferential linear segments are not contiguous with each other. As a result, the circumferentially positioned linear segments cannot be combined into a single representative image. Examples of acceptable independent light sources include white and/or blue LEDs, operable either in a steady state mode or in the case of the QBC® STAR type reader, in a pulsed mode. The relative blue spectrum of a white LED or the inclusion of a separate blue LED can excite the fluorescence of a dye such as Acridine Orange in the tube  10 . 
     The processor  36  is adapted (e.g., programmed) to perform several tasks, including: a) controlling the sample imaging device  34  based on the analysis at hand; b) controlling the centrifuge  42  for those embodiments that include one; c) receiving and acting on operator input entered through the operator input device  40 ; and d) producing information relating to bands  24  of interest within the image based on the signals from the sample imaging device  34 . The extent of the information relating to the bands  24  can vary depending upon the embodiment of the device  28 . For example, the processor  36  may be adapted to provide information relating to the adequacy of the sample image, and/or with algorithmic capability that is operable to analyze the signals representative of the sample image and produce data (e.g., CBC, hematocrit, WBC count, etc.) relating thereto based on characteristics of the different bands  24  within the centrifuged sample. In some applications, the processor  36  can be adapted to produce graphic markings based on the analysis of the sample that can be superimposed over the sample image when displayed to illustrate the calculated band boundaries  25  relative to the sample image. Using the analysis of a blood sample as an example, graphic markings can be used to identify features such as the: a) bottom of the tube  10 ; b) bottom of the float  22 ; c) red blood cell/granulocyte interphase; d) granulocyte/lymphocyte and monocyte interphase; e) lymphocyte and monocyte/platelet interphase; f) platelet/plasma interphase; g) top of the float  22 ; h) plasma/air interphase; etc. 
     The sample data display  38  is in communication with the processor  36  and includes a display screen. The display screen is an electronic screen (e.g., flat screen LED, LCD, etc.) operable to display the calculated results and/or a digital image of the sample residing within the centrifuged sample with sufficient optical resolution so that the image can be evaluated by a technician operator to provide the information pertaining to the bands  24  of interest within the centrifuged sample. In those embodiments that include an operator input device  40  (e.g., key pad, touch screen, etc.), the operator input device  40  allows the operator to enter relevant patient or other information, if desired. The sample display  38  may be integral with the housing  30 , or it may be an independent device in communication with the processor  36 . For example, universal monitors are often used in medical facilities, which monitors have the capability of displaying data from more than one analysis device. In such an application, the data to be displayed may be viewed on an integral display screen and/or a remotely located display device in communication with the processor  36 . 
     In some embodiments, the analysis device  28  includes a communication port  50  for sending the signals representative of the sample image to a remote location. The communication port  50  can be a hardwire port for communicating by hardwire connection to a remote site, or it can be a wireless communication connection (e.g., similar to that used in a wireless phone). 
     In some embodiments, fiduciary marks  52  (i.e., calibration, measurement marks, etc.) may be placed on or in the capillary tube  10 , or the tube holder  32 , or on a measuring device positioned adjacent the tube  10  (e.g., a ruler) to facilitate geometric and/or optical calibration and thereby account for any image distortion introduced by the camera. In those instances where the fiduciary marks are placed on or in the tube, a particularly useful embodiment is one wherein the marks are positioned relative to the internal cavity to permit geometric evaluation of sample within the internal cavity. In those instances where fiduciary marks  52  are disposed on a measuring device positioned adjacent the tube  10 , the measurement device can measure along an axis that is maintained parallel to the lengthwise axis (e.g., axial direction) of the tube  10 . In such embodiments, the measurement device is preferably in close proximity (e.g., in the same focal plane) as the sample tube  10 . Alternately, a look-up-table can be provided by factory calibration to serve this function. During the image processing and analysis steps, the calibration information can be used to ensure correct length measurements of the tube  10  features, regardless of their position in the image frame or distance from the camera and can compensate for instrument-to-instrument differences. 
     Operation: 
     A fluid sample (e.g., whole blood) is collected from a patient and deposited into a capillary tube  10  such as those used in the QBC® hematology system for subsequent centrifugation. As indicated above, the centrifuge may be independent of, or incorporated into, the analysis device  28 . The sample is centrifuged for a period of time adequate to create constituent layer separation within the sample disposed in the tube  10 , and the representative bands  24  associated therewith. The centrifuged sample is then imaged using the sample imaging device  34 . The image includes substantially all of the radial width  18  and axial length  20  of the sample residing within the internal cavity  16  of the tube  10  in the region where the float  22  resides after centrifugation. Because capillary tubes  10  are not always filled with the exact same volume of fluid sample, the sample imaging device  34  preferably images the region  48  of the tube  10  from the top meniscus to the bottom of the red blood cell layer. It is desirable, but not required, that the bottom of the tube  10  be imaged as well. If the sample being imaged is disposed within a STAR type QBC® tube, for example, the total length between the tube bottom to the tube top fill position is approximately 53 mm. The distance from the tube top fill position to the bottom of the float  22  in most instances is about 37 mm. In those device  28  embodiments that include a centrifuge, the sample may be centrifuged and the centrifuge subsequently stopped or slowed to a very low RPM prior to the imaging. The sample imaging device  34  produces signals representative of each image and communicates those signals to the processor  36 . 
     The images signals are subsequently analyzed within the processor  36  using image processing algorithms to isolate and analyze the bands  24  of interest within the sample, and in some instances relevant sections of the bands  24 . Before or after the image signals are analyzed, the image signals may be sent to the sample data display  38  for evaluation by the operator. The ability to have an operator visually evaluate an image that includes substantially all of the radial width  18  of the sample within the tube  10 , and a relevant portion of the axial length  20  of the sample is a substantial advancement of the technology. A person of skill in the art will recognize that no automated system can account for all potential variables within the sample image. For example, during the centrifugation process, there is a chance that sample will exit the capillary tube  10  and pass into the retaining tube of the centrifuge. In such instances, the released sample can contaminate the exterior of the capillary tube  10  and inhibit accurate analysis. Similarly, a misplaced tube label or debris deposited on the exterior of the capillary tube  10  during handling can also inhibit or prevent accurate analysis. In these instances, the ability of the present device  28  to produce a single substantially complete image of the centrifuged sample will enable the operator to identify such potential problems and take appropriate action. As another example, the image available with the present device  28  will also enable the operator to evaluate other aspects of the sample image for potential problems; e.g., overall image quality, accuracy of sample coloration, the degree to which a blood sample may be lipemic or icteric, etc. In those applications where the operator evaluates the image after processing and boundary markings are assigned by the processor  36 , the operator can evaluate whether the assigned boundary markings are accurately positioned relative to the sample image. Hence, the ability to have an operator visually evaluate an image using the present device  28  provides considerable quality controls to the analysis process. It should be emphasized that the present instrument, as described herein, may be used in locations where trained operators are present, and also locations where no trained operators are present (e.g., a CLIA-waived environment). In the latter type location, the sample images captured by the present device  28  may be sent to a remotely located trained operator for analysis. If it is not possible to have a trained operator review the image and/or results within a predetermined period of time, the present device  28  may be programmed to prevent the release of any data if the sample image has any detectable anomalies. A visual image analysis is preferable in that the criteria for analysis rejection can be loosened, but a purely automated analysis (e.g., that checks for anomalies) is preferable to no analysis at all. 
     The extent to which the present device  28  images the centrifuged sample within the tube  10  makes possible another quality control mechanism. As indicated above, the present device  28  images substantially all of the radial width  18  and a significant portion of the axial length of the centrifuged sample. In some instances, the radial portion of the image can be expanded to a point outside of the capillary tube  10  to include other imageable features such as calibration markers or areas. The image characteristics associated with the regions outside of the capillary tube  10  can be compared against the characteristics of the region inside the tube  10 . Inconsistencies identified by the comparison of the characteristics (e.g., brightness) can be used to evaluate the accuracy of the image. This type of quality control is not possible using the prior art reading devices that utilize a linear scanning device, which has essentially only a one pixel width. 
     Although the invention has been described and illustrated with respect to exemplary embodiments thereof, the foregoing and various other additions and omissions may be made therein and thereto without departing from the spirit and scope of the present invention.