Abstract:
A device and method for separating heavier and lighter fractions of a fluid sample. The device includes a plurality of constituents comprising a container and a composite element in the container. The composite element is a separator comprising a deformable bellows, a ballast mounted to the lower end of the bellows, and a float is engageable with an upper end of the bellows. A fluid sample is delivered to the container and the device is subjected to centrifugation whereby the centrifugal load causes the ballast to move toward the bottom of the tube and causes an elongation and narrowing of the bellows. The separator then moves down the tube and stabilizes in a position between the separated phases of the fluid sample. Termination of the centrifugal load enables the bellows to return to its original condition in sealing engagement with the walls of the tube. The dense formed phase of the fluid sample will lie between the separator and the bottom of the tube, while less dense liquid phase of the fluid sample will be the separator.

Description:
BACKGROUND OF THE INVENTION  
         [0001]    1. Field of the Invention  
           [0002]    This invention relates to a device and method for separating heavier and lighter fractions of a fluid sample. More particularly, this invention relates to a device and method for collecting and transporting fluid samples whereby the device and fluid sample are subjected to centrifugation in order to cause separation of the heavier fraction from the lighter fraction of the fluid sample.  
           [0003]    2. Description of Related Art  
           [0004]    Diagnostic tests may require separation of a patient&#39;s whole blood sample into components, such as serum or plasma, the lighter phase component, and red blood cells, the heavier phase component. Samples of whole blood are typically collected by venipuncture through a cannula or needle attached to a syringe or an evacuated collection tube. Separation of the blood into serum or plasma and red blood cells is then accomplished by rotation of the syringe or tube in a centrifuge. Such arrangements use a barrier for moving into an area adjacent the two phases of the sample being separated to maintain the components separated for subsequent examination of the individual components.  
           [0005]    A variety of devices have been used in collection devices to divide the area between the heavier and lighter phases of a fluid sample.  
           [0006]    The most widely used device includes thixotropic gel materials such as polyester gels in a tube. The present polyester gel serum separation tubes require special manufacturing equipment to prepare the gel and to fill the tubes. Moreover, the shelf-life of the product is limited in that overtime globules may be released from the gel mass. These globules may be present in the serum and may clog the measuring instruments, such as the instrument probes used during the clinical examination of the sample collected in the tube. Such clogging can lead to considerable downtime for the instrument to remove the clog.  
           [0007]    No commercially available gel is completely chemically inert to all analytes. If certain drugs are present in the blood sample when it is taken, there can be an adverse chemical reaction with the gel interface.  
           [0008]    Therefore, a need exists for a separator device that (i) is easily used to separate a blood sample; (ii) is independent of temperature during storage and shipping; (iii) is stable to radiation sterilization; (iv) employs the benefits of a thixotropic gel barrier yet avoids the disadvantages of placing a gel in contact with the separated blood components; (v) minimizes cross contamination of the heavier and lighter phases of the sample during centrifugation; (vi) minimizes adhesion of the lower and higher density materials against the separator device; (vii) is able to move into position to form a barrier in less time than conventional methods and devices; (viii) is able to provide a clearer specimen with less cell contamination than conventional methods and devices; and (ix) can be used with standard sampling equipment.  
         SUMMARY OF THE INVENTION  
         [0009]    The present invention is a method and assembly for separating a fluid sample into a higher specific gravity phase and a lower specific gravity phase. Desirably, the assembly of the present invention comprises a plurality of constituents. Preferably, the assembly comprises a container and a composite element.  
           [0010]    Most preferably, the container is a tube and the composite element is a separator arranged to move in the tube under the action of centrifugal force in order to separate the portions of a fluid sample.  
           [0011]    Most preferably, the tube comprises an open end, a closed end and a sidewall extending between the open end and closed end. The sidewall comprises an outer surface and an inner surface. The tube further comprises a closure disposed to fit in the open end of the tube with a resealable septum. Alternatively, both ends of the tube may be open, and both ends of the tube may be sealed by elastomeric closures. At least one of the closures of the tube may include a needle pierceable resealable septum.  
           [0012]    Preferably, the separator element comprises an overall specific gravity at a target specific gravity of σ t . The target specific gravity is that required to separate a fluid sample into at least two phases.  
           [0013]    Preferably, the separator comprises at least two or more regions of differing specific gravities. Preferably, at least one of the regions is higher than the target specific gravity and at least one of the regions is lower than the target specific gravity.  
           [0014]    The separator is disposed in the tube at a location between the top closure and the bottom of the tube. The separator includes opposed top and bottom ends and comprises a bellows, a ballast and a float. The components of the separator are dimensioned and configured to achieve an overall density for the separator that lies between the densities of the phases of a fluid sample, such as a blood sample.  
           [0015]    The bellows of the separator is molded from a resiliently deformable material that exhibits good sealing characteristics when placed against an adjacent surface. The bellows has an upper end that is at or in proximity to the top end of the separator and an opposed lower end that is disposed between the opposed ends of the separator.  
           [0016]    The upper end of the bellows may be formed from a needle pierceable material that may be pierced by a needle cannula for depositing a fluid sample into the tube. Additionally, the upper end of the bellows initially may be engaged releasably with the closure mounted in the open top end of the tube.  
           [0017]    Preferably, the bellows includes a toroidal sealing section which, in an unbiased state of the bellows, defines an outer diameter that exceeds the inside diameter of the tube. However, the bellows can be deformed slightly so that the outer circumferential surface of the toroidal sealing section is biased against the inner circumferential surface of the tube to achieve a sealing engagement between the bellows and the tube. The bellows may be elongated by oppositely directed forces in proximity to the opposed upper and lower ends thereof. Elongation of the bellows in response to such oppositely directed forces will reduce the outside diameter of the toroidal sealing section of the bellows. Sufficient elongation of the bellows will cause the toroidal sealing section of the bellows to be spaced inwardly from the internal surface of the blood collection tube.  
           [0018]    Desirably, the toroidal sealing section may be comprised of any natural or synthetic elastomer or mixture thereof, that is inert to the fluid sample of interest and is flexible.  
           [0019]    Preferably, the toroidal sealing section comprises a qualitative stiffness, expressed as follows:  
         S   *     =     k     a                   ρ   w          D   2                               
 
           [0020]    whereby S* is the non-dimensional stiffness coefficient, k is a force required to deflect the bellows a given length, a is the applied acceleration, D is the diameter of the toroidal sealing section and ρ w  is the density of water.  
           [0021]    Desirably, the qualitative stiffness of the toroidal sealing section is from about 0.00006 to about 190.  
           [0022]    Preferably, the toroidal sealing section may be subjected to a characteristic or radial deflection under an applied load such as an axially applied load. The characteristic or radial deflection is defined as a change in length of the toroidal sealing section relative to the change in cross section diameter of the toroidal sealing section. Preferably, the toroidal sealing section has a characteristic or radial deflection ratio of about 1.5 to about 3.5.  
           [0023]    Preferably, the toroidal sealing section when subjected to an applied load, such as centrifugation, to cause axial deformation of the toroidal sealing section, the change in cross section diameter of the toroidal sealing section may be expressed as follows:  
               D   before     -     D   during         D   before       ×   100      %     =     Δ                   D   m                             
 
           [0024]    wherein ΔD m  is from about 5% to about 20%.  
           [0025]    Therefore, a change in cross section diameter of the toroidal sealing section is proportional to the undeflected cross section diameter of the toroidal sealing section. Preferably, the proportion is from about 0.03 to about 0.20.  
           [0026]    Preferably, the ballast is a substantially tubular structure formed from a material having a greater density than the heavy phase of blood. The generally tubular ballast has a maximum outside diameter that is less than the inside diameter of the tube. Hence, the ballast can be disposed concentrically within and spaced from a cylindrical sidewall of the tube. The ballast may be securely and permanently mounted to the lower end of the bellows.  
           [0027]    Preferably, the float is formed from a material having a density less than the density of the lighter phase of the blood and may be engaged near the upper end of the bellows. Additionally, the float is movable relative to the ballast. For example, the float may be substantially tubular and may be slidably telescoped concentrically within the tubular ballast. Hence, the float and the ballast can move in opposite respective directions within the tube.  
           [0028]    In use, a fluid sample enters the assembly by needle. The needle pierces a portion of the bellows adjacent the top end of the separator and partially through the hollow interior of the float. The needle is withdrawn from the assembly and the septum of the closure and the bellows reseals.  
           [0029]    The assembly is then subjected to centrifugation. Forces exerted by the centrifuge causes a gradual separation of the phases of the fluid sample such that the more dense phase moves toward the bottom end of the tube, and the less dense liquid is displaced to regions of the tube above the more dense phase. Simultaneously, the centrifugal load will cause the dense ballast to move outwardly relative to the axis of rotation and toward the bottom of the tube. This movement of the ballast will generate an elongation and narrowing of the bellows. Thus, the outside diameter of the toroidal sealing section of the bellows will become less than the inside diameter of the tube. Additionally, the centrifugal load and the deformation of the bellows will cause the separator to disengage from the top closure. Hence, the separator will begin to move toward the bottom of the tube. Air trapped between the fluid sample and the separator initially will move through the circumferential space between the separator and the tube. After sufficient movement, the bottom end of the separator will contact the surface of the fluid sample. At this point, air trapped within the hollow interior of the separator can impede further downward movement of the separator into the fluid sample. However, this air can pass through the defect in the bellows caused by the needle or through some other manufactured defect in the bellows.  
           [0030]    The ballast will cause the separator to sink into the fluid sample while the float will buoyantly remain near the surface of the fluid sample thereby causing an elongation and narrowing of the bellows. The separator is not able to move in the tube without friction between the separator and the inner wall surface of the tube. The less dense liquid phase of the fluid sample will move through the space between the separator and the walls of the tube. As noted above, the overall density of the separator is selected to be less than the density of the formed phase of the fluid sample, but greater than the density of the less dense liquid phase of the fluid sample. Thus, the separator will stabilize at a location between the formed and liquid phases of the fluid sample after a sufficient period of centrifugation. The centrifuge then is stopped. The termination of the centrifugal load enables the toroidal sealing section of the bellows to return toward its unbiased dimensions, and into sealing engagement with the interior of the tube. The less dense liquid phase of the fluid sample can be separated from the tube by either removing the closure or passing a needle through the closure. Alternatively, in certain embodiments, the more dense formed phase can be accessed through a sealed opening in the bottom end of the tube.  
           [0031]    The separator of the present invention comprises a useful range of parameters and there are two principle driving equations for defining the parameters:  
           σ t   V   t =σ f   V   f +σ s   V   s    
           (conservation of mass)  
           [0032]    [0032]           (         (       σ   f     -     σ   t       )          V   f       -       (       σ   s     -     σ   t       )          V   s         )          ρ   w       =         δ   ·   Δ                     D   ·   k       a                           
 (force balance)  
           [0033]    The following non-dimensional parameters may then be substituted into the force balance:  
           
         V 
         s 
         *=V 
         s 
         /D 
         3 
         ; V 
         f 
         *=V 
         f 
         /D 
         3 
         ; S*=k/a ρ 
         w 
         D 
         2  
       
           [0034]    to arrive at:  
         (         (       σ   f     -     σ   t       )          V   f   *       -       (       σ   s     -     σ   t       )          V   s   *         )     =         δ   ·   Δ                     D   ·     S   *         D                           
 
           [0035]    So as to scale prototypes to any size device, wherein the following are defined:  
           [0036]    σ t , σ f , σ s  are the specific gravities of the separator device, float and ballast, respectively;  
           [0037]    V t , V f , V s  are the volumes of the separator device, float and ballast, respectively;  
           [0038]    σ w  is the density of water;  
           [0039]    k is the separator spring constant;  
           [0040]    a is the applied acceleration; and  
           [0041]    δ is the deflection ration defined by: ΔL/ΔD, where ΔL is the change in length.  
           [0042]    The left side of the equation can be an infinite number of combinations of materials and geometries and if it is equal to the product of the right side it can be concluded that the device will function.  
           [0043]    Desirable values for the right side of the equation are as follows:  
           δ=1.5−3.5  
           Δ D/D= 0.05 to 0.2  
             S*= 0.043 to 0.220.  
           [0044]    The assembly of the present invention is advantageous over existing separation products that use gel. In particular the assembly of the present invention will not interfere with analytes as compared to gels that may interfere with analytes. Another attribute of the present invention is that the assembly of the present invention will not interfere with therapeutic drug monitoring analytes.  
           [0045]    Most notably, the time to separate a fluid sample into separate densities is achieved in substantially less time with the assembly of the present invention as compared to assemblies that use gel.  
           [0046]    Another notable advantage of the present invention is that fluid specimens are not subjected to low density gel residuals that are at times available in products that use gel.  
           [0047]    A further attribute of the present invention is that there is no interference with instrument probes.  
           [0048]    Another attribute of the present invention is that samples for blood banking tests are more acceptable than when a gel separator is used.  
           [0049]    Another attribute of the present invention is that only the substantially cell-free serum fraction of a blood sample is exposed to the top surface of the separator, thus providing practitioners with a clean sample.  
           [0050]    A further attribute of the present invention is that the separator moves in the tube without friction between the separator and the inner wall of the tube under the action of centrifugal force.  
           [0051]    Additionally, the assembly of the present invention does not require any additional steps or treatment by a medical practitioner, whereby a blood or fluid sample is drawn in the standard fashion, using standard sampling equipment. 
       
    
    
     DESCRIPTION OF THE DRAWINGS  
       [0052]    [0052]FIG. 1 is an exploded perspective view of the assembly of the present invention.  
         [0053]    [0053]FIG. 2 is a perspective view of the closure of the assembly of FIG. 1.  
         [0054]    [0054]FIG. 3 is a bottom plan view of the closure of FIG. 2.  
         [0055]    [0055]FIG. 4 is a cross-sectional view of the closure of FIG. 3 thereof.  
         [0056]    [0056]FIG. 5 is a perspective view of the bellows of the separator of the assembly of FIG. 1.  
         [0057]    [0057]FIG. 6 is a cross-sectional view of the bellows of FIG. 5 taken along line  6 - 6  thereof.  
         [0058]    [0058]FIG. 7 is a bottom plan view of the ballast of the separator of the assembly of FIG. 1.  
         [0059]    [0059]FIG. 8 is a cross-sectional view of the ballast of FIG. 7 taken along line  8 - 8  thereof.  
         [0060]    [0060]FIG. 9 is a perspective view of the float of the separator of the assembly of FIG. 1.  
         [0061]    [0061]FIG. 10 is a side elevational view of the float of the separator of the assembly of FIG. 1.  
         [0062]    [0062]FIG. 11 is a cross-sectional view of the float of FIG. 10 taken along line  11 - 11  thereof.  
         [0063]    [0063]FIG. 12 is a side elevational view of the assembly of the present invention.  
         [0064]    [0064]FIG. 13 is a cross-sectional view of the assembly of FIG. 12 taken along line  13 - 13  thereof.  
         [0065]    [0065]FIG. 14 is a cross-sectional view of the assembly of FIG. 12 taken along line  13 - 13  thereof, showing the separator under a centrifugal load.  
         [0066]    [0066]FIG. 15 is a cross-sectional view of the assembly of FIG. 12 taken along line  13 - 13  thereof, showing the separator sealingly engaged with the tube between the liquid and formed phases of the fluid sample.  
         [0067]    [0067]FIG. 16 is a cross-sectional view similar to FIG. 13, but showing an alternate embodiment of the present invention.  
     
    
     DETAILED DESCRIPTION  
       [0068]    The present invention may be embodied in other specific forms and is not limited to any specific embodiments described in detail, which are merely exemplary. Various other modifications will be apparent to and readily made by those skilled in the art without departing from the scope and spirit of the invention. The scope of the invention will be measured by the appended claims and their equivalents.  
         [0069]    The present invention is illustrated in FIGS. 1 and 13- 16 , wherein assembly  10  includes a tube  12 , a closure  14  and a separator assembly  16 . Tube  12  includes a closed bottom  18 , an open top  20  and a cylindrical sidewall  22  extending therebetween. Sidewall  22  includes an inner surface  23  with an inside diameter “a” extending from top end  20  to a location substantially adjacent bottom end  18 .  
         [0070]    Closure  14 , as shown in FIGS.  2 - 4 , is unitarily molded from an elastomeric material and includes a top end  24  and a bottom end  26 . Portions of closure  14  adjacent top end  24  define a maximum outside diameter which exceeds the inside diameter “a” of tube  12 . Additionally, portions of closure  14  at top end  24  include a central recess  28  which defines a needle pierceable resealable septum. Portions of closure  14  extending upwardly from bottom end  26  taper from a minor diameter which is approximately equal to or slightly less than the inside diameter “a” of tube  12  to a major diameter that is greater than inside diameter “a”. Thus, bottom end  26  of closure  14  can be urged into portions of tube  12  adjacent open top end  20  thereof, and the inherent resiliency of closure  14  will ensure a sealing engagement with the inner circumferential surface of cylindrical sidewall  22  of tube  12 .  
         [0071]    Closure  14  is formed to include a bottom recess  30  extending into bottom end  26 . Bottom recess  30  is characterized by a central convex cone  32 . Additionally, a plurality of spaced apart resiliently deflectable arcuate flanges  34  extend around the entrance to recess  30 . Flanges  34  function to releasably hold separator assembly  16 .  
         [0072]    Separator assembly  16  includes a bellows  36 , a ballast  38  and a float  40 . Bellows  36 , as shown in FIGS. 5 and 6, is unitarily molded from a resiliently deformable material, that exhibits good sealing characteristics. More particularly, bellows  36  is symmetrical about a center axis and includes an upper end  42  a lower end  44 , and a hollow interior  45  that is open at lower end  44 . Portions of bellows  36  adjacent upper end  42  define an enlarged mounting head  46  with a top section that is convexly conical in an initial unbiased condition of bellows  36 . The conical section of bellows  36  adjacent upper end  42  can be deflected into a conical concave configuration that abuts conical portion  32  in recess  30  of closure  14 . Bellows  36  further includes a generally toroidal sealing section  47  intermediate upper and lower ends  42  and  44 . Toroidal sealing section  47  defines an outside diameter “b” which, in an unbiased condition of bellows  36 , slightly exceeds inside diameter “a” of tube  12 . However, oppositely directed forces on upper and lower ends  42  and  44  of bellows  36  will lengthen bellows  36  simultaneously reducing the diameter of toroidal sealing section  47  to a dimension less than “a”. A narrow neck  48  is defined between mounting head  46  and toroidal sealing section  47 . Neck  48  is dimensioned to be engaged within the area defined by arcuate flanges  34  on closure  14 . Hollow interior  45  of bellows  36  includes an annular float mounting bead  49  at a location substantially aligned with neck  48 .  
         [0073]    Portions of bellows  36  between toroidal sealing section  47  and lower end  44  define a generally cylindrical ballast mounting section  50  of outside diameter “c”, inside diameter “d” and length “e”. Ballast mounting section  50  terminates at an outwardly projecting flange  51  substantially adjacent lower end  44  of bellows  36 .  
         [0074]    Ballast  38  of separator  16  is generally cylindrical tube unitarily formed from a material that will not react with blood or other liquid being separated and that has a density higher than the blood or other liquid being separated. Ballast  38  preferably is substantially tubular and includes opposed upper and lower ends  52  and  54 , as shown in FIGS. 7 and 8. Outer circumferential surface areas of ballast  38  define a maximum outside diameter “f” that is less than inside diameter “a” of tube  12 . Inner circumferential surface regions of ballast  38  are characterized by an inwardly directed flange  56  adjacent upper end  52 . Flange  56  defines an inside diameter “g” which is approximately equal to outside diameter “c” of ballast mounting section  50  of bellows  36 . Additionally, flange  56  of ballast  38  defines a length “h” which is approximately equal to length “e” of ballast mounting section  50  on bellows  36 . As a result, ballast  38  can be securely mounted to ballast mounting section  50  of bellows  36  at locations between flange  51  and toroidal sealing section  47 . Portions of ballast  38  between flange  56  and lower end  54  of ballast  38  will project downwardly below lower end  44  of bellows  36  in this interengaged position.  
         [0075]    Float  40  of separator  16  is a generally stepped tubular structure unitarily molded from a foam material having a density less than the density of the liquid phase of blood. Float  40  may be unitarily formed from a low density polyethylene. As shown in FIGS.  9 - 11 , float  40  has an upper end  58 , a lower end  60  and a passage  62  extending axially therebetween. Float  40  is formed with an annular groove  64  extending around the outer circumferential surface thereof at a location spaced slightly from upper end  58 . Annular groove  64  is dimensioned to be resiliently engaged by inwardly directed annular bead  49  of bellows  36  for securely retaining portions of float  40  near upper end  58  to portions of bellows  36  near lower end  44  thereof. Additionally, groove  64  is configured to define apertures  65  that enable an air flow that insures narrowing of bellows  36  in the assembled condition of separator  16 , as explained below.  
         [0076]    Float  40  further includes narrow neck  66  at locations approximately midway between top and bottom ends  58  and  60 . Neck  66  defines a diameter “i” which is less than inside diameter “d” of ballast mounting section  50  of bellows  36 . As a result, neck  66  is freely movable in an axial direction within ballast mounting section  50  of bellows  36 .  
         [0077]    Float  40  further includes a substantially cylindrical base  68  defining a diameter “j” which is less than the inside diameter of ballast  38  between flange  56  and lower end  54 . Thus, base  68  of float  40  can be slidably moved in an axial direction relative to portions of ballast  38  adjacent bottom end  54  thereof.  
         [0078]    Separator  16  is assembled by resiliently engaging ballast mounting section  50  of bellows  36  with flange  56  of ballast  38 . Float  40  then is urged upwardly through ballast  38  and into lower end  44  of bellows  36 . After sufficient insertion, annular groove  64  of float  40  will engage annular bead  49  of bellows  36 . Thus, bellows  36 , ballast  38  and float  40  will be securely engaged with one another.  
         [0079]    Portions of separator  16  adjacent upper end  42  of bellows  36  then are urged into recess  30  in bottom end  26  of closure  14 . This insertion will cause arcuate flanges  34  of closure  14  to deflect. After sufficient insertion, arcuate flanges  34  will resiliently return toward an undeflected condition in which flanges  34  engage neck  48  of bellows  36 . Additionally, the concave cone at upper end  42  of bellows  36  is deflected downwardly and into a convex shape by cone  32  of closure  14 .  
         [0080]    The subassembly comprised of closure  14  and separator  16  then is inserted into open top  20  of tube  12  such that separator  16  and lower end  26  of closure  14  lie within tube  12 , as shown in FIGS. 12 and 13. Closure  14  will sealingly engage against interior surface regions and top end  20  of tube  12 . Additionally, toroidal section  48  of bellows  36  will sealingly engage against inner surface  23  of tube  12 .  
         [0081]    As shown in FIG. 13, a liquid sample is delivered to the tube by a needle that penetrates septum  28  of closure  14  and upper end  42  of bellows  36 . For purposes of illustration only, the liquid sample is blood. Blood will flow through central opening  62  of float  40  and to bottom end  18  of tube  12 . The needle then will be withdrawn from assembly  10 . Upon removal of the needle septum  28  of closure  14  will reseal itself. Upper end  42  of bellows  36  also will reclose itself in a manner that will render it substantially impervious to fluid flow.  
         [0082]    As shown in FIG. 14, when assembly  10  is subjected to centrifugation or to an axial centrifugation force, the respective phases of the blood will begin to separate so that the more dense phase comprising red blood cells will be displaced toward the bottom end  18  of tube  12  and so that the less dense phase comprising serum will be displaced to a location immediately above the denser phase and simultaneously, the centrifugal loads will urge ballast  38  toward bottom end  18  of tube  12  relative to float  40 . This movement of ballast  38  will generate a longitudinal deformation of bellows  36 . As a result, toroidal sealing section  48  will become longer and narrower and will be spaced concentrically inwardly from the inner surface  23  of sidewall  20  of tube  12 . The smaller cross-section of toroidal section  48  will permit a movement of portions of bellows  36  adjacent lower end  44  to move toward bottom  18  of tube  12 . Upper end  42  of bellows  36  initially will be retained adjacent closure  14  by arcuate flanges  34 . However, all of closure  14  is resiliently deformable, and hence arcuate flanges  34  will resiliently deform downwardly in response to centrifugal loads created on separator  16 , and particularly on ballast  38 . Hence, separator  16  will separate from closure  14  and will begin moving in tube  12  toward bottom end  18 , as shown in FIG. 14. Air in portions of tube  12  between the blood and separator  16  will flow around separator  16  and into sections of tube  12  between separator  16  and closure  14 . After sufficient movement of separator  16 , bottom end  54  of ballast  38  and/or bottom end  60  of float  40  will contact the top surface of the blood. This will leave trapped air within aperture  62  of float  40  that could impede further downward movement of separator  16 . However, the defect in top  42  of bellows  36  caused by the needle cannula will enable trapped air to escape to regions of tube  12  between separator  16  and closure  14 . Thus, ballast  38  will continue to urge separator  16  down into the separating blood. As noted above, separator  16  has an overall density between the densities of the formed and liquid phases of the blood. Consequently, separator  16  will stabilize in a position within tube  12  such that the formed phase of the blood will lie between bottom end  18  of tube  12  and separator  16 , as shown in FIG. 15. The liquid phases of the blood will lie between separator  16  and closure  14 .  
         [0083]    After this stabilized state has been reached, the centrifuge will be stopped. The termination of the centrifugal load will cause toroidal sealing section  48  of bellows  36  to resiliently return toward its unbiased condition and into sealing engagement with interior surface  23  of tube  12 . Thus, the formed and liquid phases of blood will be separated efficiently and can be accessed separately for analysis.  
         [0084]    An alternate embodiment of the tube assembly in accordance with the subject invention is identified generally by the numeral  110  in FIG. 16. Assembly  110  includes a tube  112 , a closure  114  and a separator  116 .  
         [0085]    Tube  112  includes an open top  118 , a bottom  120  and a cylindrical wall  122  extending therebetween. Bottom  120  of tube  112  has an opening  124  extending therethrough. A bottom closure  126  is sealingly engaged in opening  124 . Bottom closure  126  is formed from a needle pierceable elastomer and enables the formed phase of a blood sample to be accessed directly from bottom  120  of tube  112 .  
         [0086]    An alternate embodiment of the tube assembly of the present invention includes tube  112 , closure  114  and separator  116  wherein separator  116  is not mated with closure  114 .  
         [0087]    Closure  114  includes an elastomeric stopper  128  sealingly engaged in open top  118  of tube  112 . Stopper  128  is provided with a centrally disposed needle pierceable septum  130 . Stopper  128  further includes a bottom recess  132  having a plurality of inwardly directed resiliently deflectable arcuate flanges  134  extending thereabout. Recess  132  is not provided with a concave cone.  
         [0088]    Closure  114  further includes an outer cap  136  having an annular top wall  138  and a generally cylindrical skirt  140  depending downwardly from top wall  138 . Cap  136  is securely mounted around stopper  128  and is removably mountable over open top  118  of tube  112 . Top wall  138  of stopper  136  is provided with a central opening  142  that substantially registers with septum  130 .  
         [0089]    Separator  116  includes a bellows  144 , a ballast  146  and a float  148 . Bellows  144  includes an upper end  150 , a lower end  152  and a toroidal sealing  154  therebetween. Unlike the prior embodiment, portions of bellows  144  adjacent upper end  150  are not conically generated. Rather, these upper portions of bellows  144  are substantially spherically generated and will nest with recess  132  in stopper  128  without the inward deformation that had been described with respect to the first embodiment. Portions of bellows  144  adjacent lower end  152  and adjacent toroidal sealing  154  are substantially the same as in the prior embodiment.  
         [0090]    Ballast  146  includes an upper end  156  and a lower end  158 . Portions of ballast  146  in proximity to lower end  158  defer from the prior embodiment in that inwardly directed flanges  160  are provided for trapping float  148 . Thus, any post-assembly downward movement of float  148  relative to ballast  146  is substantially prevented. However, upward movement of float  148  relative to ballast  146  is possible, and will occur during centrifugation.