Patent Abstract:
An apparatus and method are provided for determining boundaries between fractions in a fractionated sample. The method includes the steps of providing the fractionated sample in a receptacle, the receptacle having a transparent window, positioning the receptacle in a known position relative to datum means, capturing an image of the receptacle and datun means, and processing the image to determine the position of boundaries between the fractions in the sample relative to the datum means. An apparatus and method for extracting a selected fraction of a fractionated sample are also disclosed.

Full Description:
BACKGROUND OF THE INVENTION  
       [0001]     The present invention relates to a method and apparatus for automatically determining the boundaries between fractions in a fractionated sample and to a method and apparatus for automatically extracting fractions from a fractionated sample and in particular to such a method and apparatus wherein the fractionated sample is a fractionated blood sample.  
         [0002]     Blood samples are often analysed or processed by fractionation to separate particular components. This allows particular components of interest to be extracted from a sample. Fractionation is achieved by centrifugation of the sample, which results in a sample having a number of layers or fractions, each fraction consisting of particular components of the blood. Typically, the centrifuged sample comprises three fractions, the uppermost fraction containing (amongst other components) plasma, the lowermost fraction containing (amongst other components) red blood cells and the middle fraction, known as the ‘buffy coat’, containing (amongst other components) white blood cells.  
         [0003]     The fractions are typically extracted in turn by a pipette means under manual control. The fraction containing the component of interest is retained and the other fractions may be retained or disposed of as desired. Manually controlled extraction in this manner is time consuming and expensive. It is also requires considerable skill as, to the naked eye, the boundaries between fractions can be difficult to distinguish. These problems are exacerbated if the ‘buffy coat’ is the fraction of interest, for instance if DNA analysis of the sample is required, as the buffy coat is typically relatively thin in relation to the other fractions.  
         [0004]     It is therefore an object of the present invention to provide a method and apparatus for automating some or all of this process.  
       SUMMARY OF THE INVENTION  
       [0005]     According to a first aspect of the present invention there is provided a method of determining the boundaries between fractions in a fractionated sample, the method comprising the steps of: providing the fractionated sample in a receptacle, the receptacle having at least a transparent window; providing a datum means; positioning the receptacle in a known position relative to the datum means; capturing an image of the receptacle and datum means; and processing the image to determine the position of boundaries between the fractions in the sample relative to the datum means.  
         [0006]     This thus provides an automatic method for determining the position of the boundaries between fractions within a sample. Once the boundaries are determined automatically, extraction of fractions of interest can also be carried out automatically.  
         [0007]     The image capture may be carried out by a digital imaging means, such as a digital camera.  
         [0008]     The digital image is preferably processed using an edge detection algorithm to determine the position of the boundaries between fractions.  
         [0009]     If a receptacle of known dimensions is used, the processing may further include the step of calculating the volume of each fraction.  
         [0010]     The receptacle may be illuminated by white, coloured or filtered light in order to improve the quality of the captured image or the contrast between fractions. In particular, red light may be used. Additionally or alternatively, the image may be captured through a filter, if desired, to improve the contrast between fractions.  
         [0011]     The datum means is preferably a datum plate upon which the receptacle may be positioned. The datum plate may be movable between an operational position and a stowed position. The digital imaging means is preferably mounted in a fixed relation to the operational position of the datum plate.  
         [0012]     In use, a receptacle is held at a known orientation and positioned on the datum plate when the datum plate is in the operational position. This allows the position of the lower end of the receptacle to be determined and hence the relative positions of the fraction boundaries in the sample to be determined.  
         [0013]     A calibration means may be provided on the datum plate if desired. The calibration means may be provided with suitable markings or formations to allow any or all of the focus, colour, contrast or brightness levels or alignment of the imaging means to be determined and adjusted.  
         [0014]     Preferably, the method further comprises the additional step of extracting a selected fraction or fractions of interest from the sample. Extraction is preferably achieved by a pipette means. The pipette means is preferably inserted into a selected fraction and operated so as to aspirate a volume substantially equal to the volume of the selected fraction.  
         [0015]     Preferably, the fractions are aspirated in order starting with the uppermost fraction. If the fraction of interest is not the uppermost fraction, then once the uppermost fraction is aspirated it may be discarded and the next fraction then aspirated. Once the fraction of interest is aspirated, it may be dispensed into another receptacle for storage or analysis.  
         [0016]     As the fraction boundaries may be intermingled, it may be desirable to extract a volume of material equal to a volume slightly larger or slightly smaller than the fraction providing the maximum amount of the component of interest or the maximum concentration of the component of interest respectively.  
         [0017]     The receptacle is preferably a test tube of known dimensions, such as a falcon™ tube. A processing means is preferably provided for determining the positions of the fraction boundaries and calculating the volume of each fraction. The processing means may additionally control the pipette means.  
         [0018]     According to a second aspect of the present invention there is provided a method of extracting a selected fraction of a fractionated sample, the method comprising the following steps: providing the fractionated sample in a receptacle, the receptacle having at least a transparent window; providing a datum means; positioning the receptacle in a known position relative to the datum means; capturing an image of the receptacle and datum means; processing the image to determine the position of boundaries between the fractions in the sample relative to the datum means; calculating the volume of each fraction in the receptacle from the determined positions of the boundaries between the fractions and the dimensions of the receptacle; inserting a pipette means into the selected fraction and operating the pipette means so as to aspirate a volume substantially equal to the calculated volume of the selected fraction.  
         [0019]     The second aspect of the invention may incorporate any or all of the features described in relation to the first aspect of the present invention as appropriate or as desired.  
         [0020]     According to a third aspect of the present invention there is provided an apparatus suitable for determining the boundaries between fractions in a fractionated sample, the apparatus comprising: a receptacle for holding a fractionated sample, the receptacle having at least a transparent window; a datum means; positioning means for positioning the receptacle in a known position relative to the datum means; imaging means for capturing a image of the receptacle and datum means; and processing means for processing the image to determine the position of the fraction boundaries relative to the datum means.  
         [0021]     According to a fourth aspect of the present invention there is provided an apparatus suitable for extracting a selected fraction of a fractionated sample, the apparatus comprising: a receptacle for holding a fractionated sample, the receptacle having at least a transparent window; a datum means; positioning means for positioning the receptacle in a known position relative to the datum means; imaging means for capturing a image of the receptacle and datum means; and processing means for processing the image to determine the position of the fraction boundaries relative to the datum means and for calculating the volume of each fraction of a fractionated sample from the determined positions of the boundaries between the fractions and the dimensions of the receptacle.  
         [0022]     The apparatus according to the third or fourth aspects of the invention may incorporate means relating to any or all of the features described in relation to the methods of the first aspect or second aspects of the present invention as appropriate or as desired.  
         [0023]     The apparatus may include automated robotic apparatus to achieve full automation of the process.  
         [0024]     Preferably the above method and apparatus is adapted for determining the boundaries between fractions in a fractionated sample and extracting desired fractions from the fractionated sample in the case wherein the sample is a fractionated blood sample. In particular this method and apparatus may be applied to extracting the buffy coat fraction from a fractionated blood sample. It is of course possible to apply this technique to the determination of boundaries between fractions and the extraction of fractions in other types of fractionated sample.  
         [0025]     The skilled man will of course appreciate that the above method and apparatus may be adapted to allow a plurality of samples to be processed at any one time. 
     
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0026]     In order that the invention is more clearly understood, one embodiment will now be described in greater detail below, with reference to the accompanying drawings, in which:  
         [0027]      FIG. 1  is a schematic diagram showing a fractionated blood sample in a test tube and features of an apparatus for identifying and extracting desired fractions of the sample;  
         [0028]      FIG. 2  is a schematic diagram of features of an apparatus for identifying and extracting desired fractions from a plurality of samples; and  
         [0029]      FIG. 3  is a schematic block diagram of the control circuitry for such an apparatus. 
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS  
       [0030]     Referring now to  FIG. 1 , a fractionated blood sample is provided in a test tube  10 . The sample has three fractions, an uppermost or ‘plasma’ fraction  12 , a lowermost or ‘red blood cell’ fraction  16  and an intermediate or ‘buffy coat’ fraction  14 , which contains white blood cells.  
         [0031]     The test tube  10  of this embodiment is entirely transparent such that the entire test tube acts as a viewing window. As an alternative, the test tube  10  could have a portion of the tube acting as a viewing window. The window need not be 100% transparent but may be partially transparent.  
         [0032]     In order to determine the position of the fraction boundaries, the test tube  10  is held vertically and positioned with its lower end in contact with a datum plate  18 . An image of the test tube  10  is then captured by digital camera  20 . The image is then processed by a suitable processing means  100  to determine the positions of the fraction boundaries relative to the datum plate  18 . The processing means  100  typically achieves this by use of edge detection algorithms.  
         [0033]     In order that a better quality image can be captured the test tube  10  may be illuminated by white, coloured or filtered light as required. A calibration means  24  may be provided adjacent to the test tube  10 . The calibration means  24  may be provided with suitable markings to allow the focus, colour, brightness or contrast levels or alignment of camera  20  to be adjusted. A filter  36  may be provided to improve the contrast between fractions in the sample.  
         [0034]     The test tube  10  is of known dimensions. Thus from the determined position of each fraction boundary, the volume of each fraction can be calculated by the processing means  100 . The processing means may then control an automatic pipette means  22 . The pipette means can be inserted into the sample to a desired position and used to aspirate a volume equal to the calculated volume of a particular fraction. Typically, the fractions are aspirated in turn starting with the uppermost. Those fractions which are not of particular interest may be discarded after aspiration and those fractions which are of further interest may be dispensed into other receptacles for storage or analysis.  
         [0035]     As an example, if it is desired to extract the buffy coat  14  for further analysis, the pipette means is used to aspirate the plasma fraction  12  to a level just above the upper boundary of the buffy coat  14 . The aspirated plasma fraction  12  is then discarded. The pipette means is then used to aspirate the buffy coat  14  to a level just below the lower boundary of the buffy coat  14 . The buffy coat  14  material in the pipette means  22  may then be transferred to a further receptacle for storage or analysis.  
         [0036]     It is of course possible that if a more concentrated sample of buffy coat fraction  14  is required, the plasma fraction  12  can be aspirated to a level just below the upper boundary of the buffy coat  14  and subsequently, the buffy coat fraction  14  can be aspirated to a level just above the lower boundary of the buffy coat  14 . In a further alternative, it is possible that the buffy coat fraction  14  may be extracted in a single operation by positioning the tip of the pipette means  22  either just above or just below the lower boundary of the buffy coat fraction  14  as desired and aspirating a volume of material equal to the volume of the buffy coat fraction  14 .  
         [0037]      FIG. 2  illustrates schematically how this method and apparatus may be applied to process a plurality of fractionated samples. In  FIG. 2 , a plurality of samples are each retained in test tubes  10 . The test tubes  10  are held in a tube rack  30 . The tube rack  30  may optionally be provided with releasable rack clamps  32  to further secure the test tubes  30 .  
         [0038]     Back light  44  illuminates an optical booth  42  with white, coloured or filtered light as required. Back light  44  is used generally to illuminate the top level of the samples and the orientation of the test tubes  10 . Light  34  is provided above the samples and is used to identify the buffy coat fraction  14 .  
         [0039]     A tube pop-up actuator  28  may be used to raise one or more tubes  10  into a position whereby the tube can be gripped by tube gripping means  26 . This is facilitated by utilization of an X-Y table  40  enabling the plurality of tubes to be positioned suitably corresponding to the actuator  28 . The tube gripping means has gripper jaws  38  ensuring vertical compliance of the tubes  10 . The tube gripping means  26  is then operable to position the tubes  10  on the datum plate  18 . The gripper jaws  38  enable the tubes  10  to be held securely until reaching the datum plate  18  at which point they are able to rest on the datum plate  18 . The datum plate  18  may itself be movable from a stowed position to an operational position to facilitate the movement of the tubes  10  by the tube gripping means  26  into position on the datum means  18 . After the fraction boundaries are determined and any desired fractions are extracted, the tube gripping means  26  is operable to either return the tubes  10  to their original rack  30  or to place the tubes  10  in a different tube rack  30 . The tube gripping means  26  is moved between the required positions by an actuator (not shown).  
         [0040]     In the case wherein more than one tube  10  is selected from the rack, the tubes  10  may be imaged and have their fractions extracted simultaneously or in turn. If the imaging is to be carried out simultaneously either individual cameras  20  may be provided for each tube or alternatively one camera may be provided for imaging a plurality of tubes. Additionally, if fractions in a plurality of tubes  10  are to be extracted simultaneously the pipette means  22  is provided with a plurality of pipette heads, each pipette head being individually controllable. This allows the pipette heads to extract the correct amount of material from each test tube  10 .  
         [0041]      FIG. 3  shows how the various components of the apparatus may be connected. The processing means  100  is connected directly to the camera  20 , the pipette means  22 , the tube gripping means  26 , the pop-up actuators  28  and to datum plate moving means  118 . The processing means may also be provided with a user interface  102 . The processing means may also be connected to means  110  for controlling the illumination of the tubes  10  during image capture. In one preferred embodiment, the processing means and user interface may be provided by a computer or computer system.  
         [0042]     It is of course to be understood that the invention is not to be limited to the details of the above embodiment, which is described by way of example only. Many variations are possible within the scope of the following claims.

Technology Classification (CPC): 6