Abstract:
A liquid aspiration method which includes aspirating partial aliquots of liquid at a number of different depths of penetration of a pipette tip into a liquid. The different depths are selected so that partial aliquot segments are obtained throughout the entire depth of sample, optionally excluding the extreme top and bottom levels.

Description:
FIELD OF THE INVENTION  
       [0001]     The present invention relates to the transfer of an amount of liquid from one container to another, and more particularly, to a method for aspiration of whole blood from a sample tube using a vacuum actuated pipette.  
       BACKGROUND OF THE INVENTION  
       [0002]     Clinical analyzers that automatically perform chemical assays and immunoassays of biological fluids such as urine, blood serum, plasma, cerebrospinal liquids and the like are in widespread commercial use. Generally, chemical reactions between an analyte in a patient sample and reagents used during analysis generate some sort of signal that can be measured by the analyzer. The concentration of analyte in the patient sample may be determined from this signal.  
         [0003]     Clinical analyzers generally use aspirating means such as a hollow sampling probe or needle to transfer predetermined volumes of liquid samples or liquid reagents between receptacles, such as between sample containers, reagent containers and reaction cuvettes disposed on the analyzer. The aspirating means typically includes an elongated, needle-like pipette member having a hollow passage whereby liquid may be aspirated into and/or dispensed from the sample probe. The pipette is carried by a transport mechanism conventionally adapted to provide horizontal and vertical movement enabling the pipette tip to be lowered into a liquid for aspiration of the liquid, and for transporting the liquid to a site whereat the pipette is lowered to an optimal position for dispensing the liquid into another container. Some type of aspirating means, such as a piston and/or vacuum assembly, is controlled to aspirate liquid into the pipette and to dispense liquid from the pipette. Hereinafter, variations of the term aspirate refer to all of such processes for extracting liquid from one container and depositing at least some of the liquid into the same or another container and further includes the supporting devices required to complete the liquid handling operations.  
         [0004]     U.S. Pat. No. 4,794,085 describes a method which permits the detection of penetration of liquid by an apertured container used for aspirating and dispensing the liquid. The apparatus has control means for advancing the container an increment of the maximum possible distance to the liquid, means to generate a pressure differential within the dispensing container that is sufficient to generate a signal that is indicative of whether the container aperture is closed by the liquid, and devices to detect and signal the pressure produced within the container by such a pressure differential.  
         [0005]     U.S. Pat. No. 4,926,701 describes a pipetting device comprising a probe for dipping into a reservoir, reaction vessel or the like, a metering pump connected to the probe and a shutoff valve disposed between the probe and the pump are provided. In the intake phase of the pump with the valve open, first air and then a predetermined quantity of liquid is taken in.  
         [0006]     U.S. Pat. No. 4,951,512 provides for providing access to a sealed container which temporarily provides an opening in the closures of the containers, and either removes contents, senses properties of the contents, or dispenses material into the container. A lift assembly moves each sample container upward against a puncture tube to produce an opening in the closure of the container. The system takes a sample through this opening or inserts a probe through the opening to measure the properties of the sample.  
         [0007]     U.S. Pat. No. 5,163,582 covers an apparatus and method for dispensing a predetermined volume of liquid from a closed, liquid-containing blood collection tube. The apparatus includes a dual conduit providing a passageway for liquid to be dispensed from a closed blood collection tube and a gas conduit providing a passageway for gas to be introduced into the blood collection tube. Included in the apparatus is insertion of the dual conduit into the blood collection tube, turning the tube away from a vertical, upright orientation, connecting and disconnecting the gas passageway from a gas supply, displacing a volume of gas through the gas passageway, and controlling the operation of the apparatus.  
         [0008]     U.S. Pat. No. 5,413,246 discloses a disposable apparatus to dispense an amount of liquid from a closed container using a stopper piercing means to access the interior of a closed blood collection tube, a gas passage means to allow a metered amount of gas to be forced into the blood collection tube, and a liquid passage means to allow fluid to be dispensed from the tube in proportion to the amount of gas forced into the tube.  
         [0009]     U.S. Pat. No. 5,499,545 is a method for improving measurement accuracy by eliminating the influence of changes in the atmospheric and internal pressures on the quantity of a liquid absorbed or discharged. A pipetting device inducts a specified quantity of liquid into a tip portion or discharges a specified quantity of liquid from the tip portion by controlling the pressure inside a cylinder portion including a cylinder and a piston.  
         [0010]     From the above art, it is evident that much attention has been given to extracting liquid samples from containers, without addressing another critical factor in obtaining accurate analytical measurements, that being aspiration of a uniform and representative aliquot portion of a patient sample from a sample container. This is an especially critical factor when the liquid to be analyzed is one of several body fluids as these frequently have a non-uniform composition. In particular, when the sample to be analyzed is a sample of whole blood that has not been preprocessed, a natural sedimentation occurs during a 30 minute period after the sample has been mixed, for instance by inversion of a primary whole blood tube. To compensate for such sedimentation, automated analyzers may be adapted with an automatic mixing mechanism, thereby adding complexity and cost as well as decreasing the reliability of an analyzer.  
       SUMMARY OF THE INVENTION  
       [0011]     It is therefore an object of this invention to provide a method for aspirating a uniform aliquot of liquid into a pipette tip when the liquid has been at rest for a sufficient period of time so that sedimentation has occurred. This invention is accomplished by aspirating partial aliquots of liquid at a number of different depths of penetration of the pipette tip into the liquid. The different depths are selected so that equal-volume, partial aliquot segments are obtained throughout the entire depth of sample, optionally excluding the extreme top and bottom levels. In an alternate embodiment, the aspiration may be a continuous aspiration process between the uppermost and lowermost levels of fluid within the container. This invention then integrates the number of partial aliquot segments into a single full aliquot representative of a uniform sample, thereby avoiding the expense and complexity of an automatic mixing mechanism. 
     
    
     BRIEF DESCRIPTION OF THE DRAWINGS  
       [0012]     The invention will be more fully understood from the following detailed description thereof taken in connection with the accompanying drawings which form a part of this application and in which:  
         [0013]      FIG. 1  is a schematic representation of an aspiration system in which the present invention may be practiced;  
         [0014]      FIG. 2  is a graphical representation of one method of defining aspiration levels using the system of  FIG. 1 ;  
         [0015]      FIG. 3  is a graphical representation of an alternate method of defining aspiration levels using the system of  FIG. 1 ;  
         [0016]      FIG. 4  illustrates a dual indicia bar code useful for identifying sample tubes like those seen in  FIG. 1 ; and,  
         [0017]      FIGS. 4A and 4B  illustrate the dual indicia bar code of  FIG. 4  applied to a sample tubes like those seen in  FIG. 1   
     
    
     DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS  
       [0018]     Referring to  FIG. 1 , there is illustrated a liquid aspiration system  10  according to the present invention which includes a pipette  12  for aspirating and dispensing liquid such as a sample liquid  14  contained in a sample tube  16 . Although one such sample tube  16  is shown for the purpose of describing the liquid aspiration system  10 , it will be apparent to those skilled in the art that any number of sample tubes  16  may be present. In an exemplary embodiment, the liquid aspiration system  10  would be used in an automated clinical analyzer (not shown). Such automated clinical analyzers are well known in the art and those skilled in the art know the functions of the elements of the analyzers to which reference is made.  
         [0019]     Pipette  12  is of conventional design and typically has an elongate central hollow lumen  18 , shown in dashed lines and extending lengthwise between a tip  20  and a vacuum line  22 . Tip  20  may have a conically narrowing nose shape terminating in a distal orifice through which liquid is aspirated into lumen  18 , and through which liquid is dispensed out from lumen  18 . Liquid aspiration system  10  is typical of many such systems and comprises a piston assembly  24  in vacuum communication with pipette  12  via vacuum line  22 . A piston  26  extends into a vacuum chamber  28  and is controlled by motor  30  to increase or decrease the vacuum pressure level within chamber  28 , vacuum line  22  and lumen  18 . A pressure transducer  32 , typically placed in vacuum communication with vacuum line  22 , monitors vacuum pressure within liquid aspiration system  10  and is conventionally employed as a feedback means operated in conjunction with motor  30  responsive to commands from a system computer  24  in order to precisely control vacuum pressure within lumen  18 . Such devices and operations are well known in the art. Commercially available pipettors  12  made from metals like stainless steel or plastics like polypropylene and similar materials, and tubing  26  made from vinyl, polypropylene, polyethylene, metal, etc, may used in the present invention. Pressure transducer  32  measures air pressure within the pipette  12  both continuously and periodically during the aspiration method of the present invention. An exemplary pressure transducer  32  is Model SCXL004DN from SenSym, Miltipas, Calif., and is interfaced to computer  15  to provide a measured air pressure within tubing  22  and lumen  18 .  
         [0020]     Aspiration system  10  further comprises a transport mechanism  34  of conventional type adapted for moving pipette  12  laterally (X-direction), vertically (Z-direction) and from front to back (Y-direction) to move pipette  12  to an aspirating location above container  16 , aspirate liquid  14  into lumen  18  from container  16 , and to dispense a desired aliquot of sample liquid into an assay cuvette (not shown). Generally, a stepper-motor and limit-switches are used within transport mechanism  34  for transporting pipette  12  and these are interfaced to system computer  15  adapted and programmed to practice the present invention. Conventional electronic drivers and interface circuits are used to interface transport mechanism  34  and liquid aspiration system  10  to computer  15 .  
         [0021]     As mentioned previously, the present invention is directed at a method for overcoming the previous necessity to mix liquid having liquid susceptible to sedimentation in a sample container prior to aspiration. Table 1 shows settling rates for red blood cells in whole blood samples from various types of patients.  
                           TABLE 1                                   Patient   Red Blood Cell Settling Rate                           Newborn Infant    0-2 mm/hr           Young Children   3-13 mm/hr           Post Adolescent Male   1-15 mm/hr           Post Adolescent Female   1-20 mm/hr                      
 
         [0022]     In accordance with the present invention, pipette  12  is lowered into container  16  to ascertain the uppermost level of fluid  14 . Various techniques are known in the art for such level determination, including capacitive, optical, and air pressure, and the choice is not critical in practicing this invention. Container  16  is supported so that the bottom of container  16  is at a known height, on an operating table for example, so that the overall total height or depth of liquid  14  may be determined by the difference between the uppermost level of fluid  14  and the known height of the bottom of container  16 . According to one embodiment of this invention, seen in  FIG. 2 , the number of different aspiration levels at which a constant volume aspiration is effected may be determined from the height of the uppermost level of fluid  14  by assigning a constant difference between next adjacent aspiration levels. Alternately, as seen in  FIG. 3 , the number of aspiration levels at which a constant volume aspiration is effected may be predetermined, in which case the height of the uppermost level of fluid  14  may be used to determine the difference between next adjacent aspiration levels.  
         [0023]     The present invention is practiced by operating transport mechanism  34  in conjunction with piston assembly  24  so as to aspirate a partial aliquot of sample liquid  14  at each of several next adjacent aspiration levels, the volume of each partial aliquot being controlled to provide a final aliquot of desired volume. In an illustrative embodiment, container  16  is a whole blood primary tube, for example of the EDTA type, and it has been predetermined to employ eight aspiration levels at which eight constant volume aspirations of partial aliquots of 25 uL blood each are effected at each of the eight next adjacent aspiration levels, providing a final aliquot of 200 uL. In an alternate embodiment, the aspiration may be a continuous aspiration process between the uppermost and lowermost levels of fluid within the container by continuously operating transport mechanism  34  in conjunction with piston assembly  24  so as to continuously lower pipettor  12  into sample liquid  14  and continuously operate piston assembly  24  so as to continuously aspirate liquid  14  between the uppermost and lowermost levels.  
         [0024]      FIG. 2  illustrates an exemplary embodiment of the present invention in an instance that three tubes contain different amounts of a liquid susceptible to sedimentation (the liquid is shaded more darkly at the bottom of the column of liquid than at the top) and it has been predetermined to employ eight aspiration levels at which eight constant volume aspirations of partial aliquots are effected. The numbered dashed lines indicate where aspiration of a partial aliquot of liquid is taken for eight different levels. The tube marked “L has a first amount of liquid therein and the eight different aspiration levels are a first distance apart; the tube marked “M” has a second volume which is less than the first amount of liquid therein, consequently the eight different aspiration levels are a smaller distance apart than the first distance; finally, the tube marked “R” has a third volume which is more than the first amount of liquid therein, consequently the eight different aspiration levels are a greater distance apart than the first distance. This illustration uses eight aspiration levels only as an example that is easy to illustrate in drawings. It must be appreciated by the reader that any number of aspiration levels, for example twenty, may be employed and remain within the bounds of the present invention. In practice, the number of aspiration levels may be empirically determined for different types of sample liquids, like those seen in Table 1, as a function of the total amount of liquid  14  in container  16  or alternately as a function of the distance between the uppermost and lowermost levels of liquid  14  in container  16 .  
         [0025]     Under control of computer  15 , transport mechanism  34  positions pipette  12  at an aspirating location above tube L and lowers pipette  12  into tube L to ascertain the uppermost level of fluid  14  therein. From the known location of the bottom of tube L, computer  15  is programmed to calculate the eight different depths at which eight constant volume partial aliquots of sample liquid  14  are to be aspirated into lumen  18  from tube L. Computer  15  then operates transport mechanism  34  to lower pipette  12  to the first of the eight different depths and controls piston assembly  24  so as to aspirate the desired partial aliquot of sample liquid  14  into lumen  18 . This process is repeated at each of the remaining seven different depths to produce the desired full aliquot of sample liquid  14 . If desired, computer  15  may operate transport mechanism  34  and piston assembly  24  to dispense a desired amount of sample liquid  14  from lumen  18  into an assay cuvette.  
         [0026]     This same process may be employed for aspiration of liquid from tubes M and R, except that, as explained above, the eight different depths at which eight constant volume partial aliquots of sample liquid  14  are aspirated will be smaller and larger, respectively, than the depths at which sample liquid  14  is from tube L.  
         [0027]      FIG. 3  illustrates an alternate embodiment of the present invention in an instance that three tubes L. M. and R contain different amounts of a liquid susceptible to sedimentation (the liquid is shaded more darkly at the bottom of the column of liquid than at the top) and it has been predetermined to employ equally spaced apart eight aspiration levels at which eight constant volume aspirations of partial aliquots are effected. In this instance, computer  15  operates transport mechanism  34  to lowers pipette  12  into tube L to ascertain the uppermost level of fluid  14  therein. From the known location of the bottom of tube L, computer  15  is programmed to determine the number of different depths at which constant volume partial aliquots of sample liquid  14  are to be aspirated into lumen  18  from tube L. In this illustration, for tube L it is possible to aspirate at eight different depths. Computer  15  then operates transport mechanism  34  to lower pipette  12  to the first of the eight equally spaced depths and controls piston assembly  24  so as to aspirate the desired partial aliquot of sample liquid  14  into lumen  18 . This process is repeated at each of the remaining seven different depths to produce the desired full aliquot of sample liquid  14 .  
         [0028]     This same process may be employed for aspiration of liquid from tubes M and R, except that, as explained above, the number of different depths at which constant volume partial aliquots of sample liquid  14  may be aspirated will be smaller and larger, respectively, than the number of depths at which sample liquid  14  is from tube L.  
         [0029]      FIG. 4  illustrates a dual indicia bar code  12  useful for identifying sample tubes L, bar code  12  having a conventional bar code indicia  14  printed thereon. Bar code  12  further comprises a tab section  16  affixed thereto by means of a narrow extension  18 , tab section  16  having a miniaturized bar code indicia  20  printed thereon. Both bar code indicia  14  and bar code indicia  20  are encoded so as to identify the contents of tube L.  FIG. 4A  illustrates the dual indicia bar code  12  of  FIG. 4  applied to a sample tube L with bar code indicia  14  around the external side of tube L while  FIG. 4B  illustrates miniaturized bar code indicia  20  on the bottom of tube L. Bar code indicia  14  may be read using a conventional bar code reader as is often mounted on the operating plane of a clinical analyzer; bar code indicia  20  however may be read by a bar code reader located below operating plane of a clinical analyzer when tube L is carried in a slot in a tube rack, the slot having a hole in its bottom so that bar code indicia  20  is visible from beneath the rack.  
         [0030]     Those skilled in the art will appreciate that the embodiments of the invention disclosed herein are illustrative of the principles of the invention and that other modifications may be employed which are still within the scope of the invention. For example, as may be seen by comparison of tubes L and M in  FIG. 3 , it may be desired to not aspirate a partial aliquot of sample liquid  14  at the uppermost level of fluid  14 , as is shown for tube M but not for tube L. Similarly, it may be desired to aspirate or not aspirate a partial aliquot of sample liquid  14  at the lowermost level of fluid  14 .  
         [0031]     The method also could also be used to aspirate a variable volume partial aliquot of sample liquid  14 , for example a larger amount at the middle range than at the uppermost and lowermost ranges. Accordingly, the present invention is not limited to those embodiments precisely shown and described in the specification but only by the following claims.