Abstract:
A device and method is provided for separating components of a fluid sample. The device includes a plurality of constituents comprising a container, a liner in the container, a closure for the container and a composite element. The composite element is a seal plug with a density between the densities of the components of the fluid sample is releasably engaged with the container closure and with the liner. A needle cannula is used to deposit a fluid sample in the liner and the entire device is placed in a centrifuge. The centrifugal load causes the seal plug to separate from the closure and causes the liner to expand outwardly against the container. The seal plug migrates into the fluid sample and stabilizes between the densities of the components of the fluid sample. The liner will resiliently return to its initial configuration upon termination of centrifugal load such that the liner sealingly engages the seal plug and separates the components of the fluid sample.

Description:
This application claims the benefit of Provisional Application No. 60/110,927, filed Dec. 5, 1998. 
    
    
     BACKGROUND OF THE INVENTION 
     1. Field of the Invention 
     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 to cause separation of the heavier fraction from the lighter fraction of the fluid sample. 
     2. Description of Related Art 
     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. 
     A variety of devices have been used in collection devices to divide the area between the heavier and lighter phases of a fluid sample. 
     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 have a specific gravity that is less than the separated serum and may float in the serum and may clog the measuring instruments, such as the instrument probes used during clinical examination of the sample collected in the tube. Such clogging can lead to considerable downtime for the instrument to remove the clog. 
     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. 
     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 many 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) can be used with standard sampling equipment; (viii) is able to move into position to form a barrier in less time than conventional methods and devices; and (ix) is able to provide a clearer specimen with less cell contamination than conventional methods and devices. 
     SUMMARY OF THE INVENTION 
     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, a liner and a composite element. 
     The container may be a conventional tube having a closed bottom, an opened top and a rigid cylindrical wall extending therebetween. The tube may include an inwardly directed rim near the open top. 
     The assembly further comprises a liner having a closed bottom, an open top and a tubular side wall. The liner is positioned in the tube such that the closed bottom of the liner is near the closed bottom of the tube. The liner, in an unbiased condition, is cross-sectionally dimensioned along most of its length to lie in spaced relationship to the tube. However, the liner may include an outwardly directed flange substantially adjacent the top of the liner. The flange may be dimensioned for engagement against the rim of the tube to position the liner longitudinally within the tube. 
     Preferably, the liner comprises a qualitative stiffness that may be characterized by a non-dimensional stiffness coefficient, S* and expressed as follows:          S   *     =       E        (     OD   -   D     )         a                   ρ   w          D   2                                
     where E is the modulus of elasticity, OD is the thickness defined by the outside diameter, D is the seal diameter, a is the applied acceleration, and ρ W  is the density of water. The stiffness coefficient is about 0.003 to about 190. 
     Preferably, the liner has a thickness of about 1.0 mm to about 2.5 mm, a modulus of elasticity of about 13.8 MPa to about 69 MPa. 
     Preferably, the assembly of the present invention will function under load created by an applied acceleration of about 300 g to about 3000 g. 
     Preferably, the liner deforms due to hydrostatic pressure under applied acceleration and returns to its initial state upon removal of the acceleration, thereby forming a seal by constricting the seal plug which is positioned in a target density region between the higher density portion and the lower density portion of a fluid sample. 
     The assembly further includes a tube closure that is sealingly engageable in the open top of the tube. The tube closure may include a tube end seat having an outside diameter at least equal to the outside diameter of the tube for disposition substantially adjacent the open top of the tube. The tube closure may include a tube stopper dimensioned for sealed engagement in portions of the tube between the top of the tube and the top of the liner. The tube closure may further include a liner stopper dimensioned for sealing engagement in the open top of the liner. 
     A plug recess extends into the bottom end of the tube closure. The entry to the plug recess may have a plurality of inwardly extending circumferentially spaced flexible walls. 
     Preferably, the composite element comprises a seal plug. The seal plug may be a single constituent or a plurality of constituents and comprises a specific density at a target density range as defined by separable fluid components densities. The seal plug may migrate freely when under an applied acceleration to settle at a location in the fluid sample in the target density region and thereby become a barrier at a desired level between the components of the fluid sample after the acceleration is removed. 
     Preferably, the seal plug has an aggregate specific gravity of about 1.028 to about 1.09. Most preferably, the seal plug has an aggregate specific gravity so that it will rest after centrifugal force, between the heavier and lighter phases of a blood sample. 
     The seal plug preferably has an overall density between the densities of two phases of a blood sample. The seal plug comprises a hard plastic shell having opposed first and second ends and an aperture extending between the ends. Outer circumferential portions of the hard plastic shell in proximity to the first end are dimensioned and configured for releasable engagement within the plug recess of the tube closure. Outer circumferential portions of the hard plastic shell in proximity to the second end are dimensioned for sealing engagement by the unbiased tube liner. The seal plug further includes an elastomeric septum that is securely mounted around the first end of the hard plastic shell to provide a pierceable barrier extending across the central passage through the shell. 
     Preferably, the seal plug 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 two phases. 
     In use, a fluid sample enters the assembly by a needle. The needle penetrates the closure and through the elastomeric septum on the seal plug for delivering a fluid sample into the liner. The needle is withdrawn from the assembly and the assembly is subjected to centrifugation. Forces exerted by the centrifuge cause the seal plug to separate from the tube closure and cause the liner to expand outwardly against the tube. Centrifugal forces then cause the seal plug to move through the expanded liner and toward the closed bottom of the tube. Sufficient movement will cause the seal plug to contact the fluid. Air trapped in the passage through the hard plastic liner and between the fluid and the elastomeric septum could create a buoyancy that might prevent further sinking of the seal plug into the fluid. However, the trapped air will be vented through a defect in the septum, such as the defect caused by the needle cannula. This venting of air permits further movement of the seal plug into the fluid. Simultaneously, the phases of the fluid will be separating such that the heavier phase component of the fluid will concentrate closer to the closed bottom, and such that the lighter phase component of the fluid will be closer to the open top. The seal plug will move primarily through the lighter phase component and toward the heavier phase component of the fluid. 
     The centrifuge may be stopped after the seal plug stabilizes between the separate phases of the fluid. Upon termination of the centrifugal load, the liner will resiliently return toward its unexpanded condition and will sealingly engage outer circumferential regions of the seal plug. As a result, the phases of the fluid sample are isolated from one another by the seal plug and may be separated for subsequent analysis. 
     When the fluid sample is blood, the higher specific gravity portion that contains the cellular components is between the separator and the bottom of the container after centrifugation. The lower specific gravity portion that contains the cell-free serum fraction or plasma is between the top surface of the separator and the top of the container after centrifugation. 
     Therefore, at the final position of the separator after centrifugation, the separator is able to substantially eliminate the presence of red blood cells in the lower specific gravity portion and the lower specific gravity is substantially free of cellular contamination. 
     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. 
     Most notably, is that 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. 
     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. 
     A further attribute of the present invention is that there is no interference with instrument probes. 
     Another attribute of the present invention is that samples for blood banking tests are more acceptable than when a gel separator is used. 
     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. 
     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. 
    
    
     BRIEF DESCRIPTION OF THE DRAWINGS 
     FIG. 1 is a perspective view of a blood collection tube device in accordance with the subject invention. 
     FIG. 2 is a cross-sectional view taken along line 2—2 in FIG.  1 . 
     FIG. 3 is a cross-sectional view similar to FIG. 2, but showing the device after insertion of a blood sample therein by a needle cannula. 
     FIG. 4 is a cross-sectional view similar to FIGS. 2 and 3, but showing the device at an early stage during centrifugation. 
     FIG. 5 is a cross-sectional view similar to FIG. 4, but showing the device at a later stage during centrifugation. 
     FIG. 6 is a cross-sectional view similar to FIG. 5, but showing the device upon completion of centrifugation, and with the respective phases of blood separated. 
     FIG. 7 is an enlarged cross-sectional view of portions of the device adjacent the seal plug during the early stages of centrifugation, as shown in FIG.  4 . 
     FIG. 8 is an enlarged cross-sectional view of portions of the device adjacent the seal plug at a later stage during centrifugation, as shown in FIG.  5 . 
     FIG. 9 is an exploded perspective view of the device. 
    
    
     DETAILED DESCRIPTION 
     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. 
     The preferred apparatus of the present invention is illustrated in FIGS. 1 to  9 , wherein device  10  comprises a tube  12 , an elastomeric liner  22 , a closure  34 , and a seal plug assembly  64 . 
     Device  10  includes rigid plastic tube  12  having an open top  14 , a closed bottom  16  and a cylindrical sidewall  18 . Sidewall  18  defines a constant inside diameter “a” along a major portion of its length. 
     Device  10  further includes an elastomeric liner  22  having an open top  24 , a closed bottom  26  and a tubular sidewall  28 . Side wall  28 , in an unbiased condition, defines an inside diameter “c” and an outside diameter “d” along a major portion of the length of liner  22 . Outside diameter “d” is less than inside diameter “a” of sidewall  18  on tube  12 , and is approximately equal to or slightly less than inside diameter “b” of rim  20 . Liner  22  is characterized by an outwardly directed flange  30  adjacent open top  24 . Portions of flange  30  immediately adjacent open top  24  are cylindrically generated and define an outside diameter “e”. However, portions of flange  30  spaced from open top  24  taper to outside diameter “d” which exists elsewhere on sidewall  28 . Diameter “e” of flange  30  is greater than inside diameter “b” of rim  20  on tube  12 , and is approximately equal to or slightly less than inside diameter “a” existing at locations on tube  12  between rim  20  and open top  14  thereof. With these relative diametrical dimensions, portions of liner  22  below flange  30  can be slid through rim  20  on tube  12 . Additionally, flange  30  can be slid into portions of tube  12  between rim  20  and open top  14 . However, flange  30  will interfere with rim  20  and will prevent liner  22  from sliding entirely past rim  20 . The length of liner  22  is selected to ensure that closed bottom  26  of liner  22  is spaced slightly from closed bottom  16  of tube  12  when flange  30  of liner  22  engages rim  20  of tube  12 . 
     A liner stopper  48  extends from liner end seat  44  to bottom end  38  of tube closure  34 . Liner stopper  48  has a cylindrical outer surface along most of its length with an outside diameter slightly greater than inside diameter “c” of liner  22 . However, portions of liner stopper  48  adjacent bottom end  38  are chamfered to a diameter that is less than inside diameter “c” of liner  22 . The chamfered bottom end of liner stopper  48  facilitates the inward compression required to urge liner stopper  48  of tube closure  34  into open top end  24  of liner  22 . Liner stopper  48  may further include by at least one axially extending vent groove [ 50 ] to permit venting of gas from liner  22  during insertion of tube closure  34 . 
     Tube closure  34  is further characterized by a plug recess  52  extending axially into bottom end  38 . The entrance to plug recess  52  is defined by a plurality of circumferentially spaced flexible release walls  54  that have inner surfaces  56  generated as part of a single cylinder with a diameter “f”. Each flexible release wall  54  may further include a chamfered surface extending from the cylindrically generated surface  56  to the bottom end  38  of tube closure  34 . Flexible release walls  54  each also include a radially aligned plug-gripping surface  60  facing into plug recess  52 . The top central portion of plug recess  52  is defined by downwardly pointing conical surface  62 . 
     Device  10  further includes a tube closure that can be pierced by a needle cannula and that will reseal itself after removal of the needle cannula. Tube closure  34  includes a top end  36  and a bottom end  38 . Top end  36  of tube closure  34  is characterized by a central recess  40  which defines a target area for piercing by a needle cannula. A radially aligned tube end seat  42  is defined between top and bottom ends  36  and  38  and faces toward bottom end  38 . Tube end seat  42  defines an outside diameter that exceeds the outside diameter of tube  12 . Thus, tube end seat  42  can be sealingly engaged against open top end  14  of tube  12 . 
     A liner end seat  44  is defined on tube closure  34  at a distance below tube end seat  42  to ensure that liner end seat  44  is substantially adjacent open top end  24  of liner  22  when tube end seat  42  is adjacent to open top end  14  of tube  12 . Portions of tube stopper  46  adjacent liner end seat  44  define a diameter approximately equal to inside diameter “a” of tube  12 . Portions of tube stopper  46  closer to tube end seat  42  define a larger diameter. Consequently, tube stopper  46  is compressed during insertion into tube  14  for achieving a tight sealing engagement. 
     A liner stopper  48  extends from liner end seat  44  to bottom end  38  of tube closure  34 . Liner stopper  48  has a cylindrical outer surface along most of its length with an outside diameter slightly greater than inside diameter “c” of liner  22 . However, portions of liner stopper  48  adjacent bottom end  38  are chamfered to a diameter that is less than inside diameter “c” of liner  22 . The chamfered bottom end of liner stopper  48  facilitates the inward compression required to urge liner stopper  48  of tube closure  34  into open top end  24  of liner  22 . Liner stopper  48  is further characterized by at least one axially extending vent groove  50  to permit venting of gas from liner  22  during insertion of tube closure  34 . 
     Tube closure  34  is further characterized by a plug recess  52  extending axially into bottom end  38 . The entrance to plug recess  52  is defined by a plurality of circumferentially spaced flexible release walls  54  that have inner surfaces  56  generated as part of a single cylinder with a diameter “f”. Each flexible release wall  54  further includes a chamfered surface  58  extending from the cylindrically generated surface  56  to the bottom end  38  of tube closure  34 . Flexible release walls  54  each also include a radially aligned plug-gripping surface  60  facing into plug recess  52 . The top central portion of plug recess  52  is defined by downwardly pointing conical surface  62 . 
     Device  10  further includes a seal plug assembly  64  which comprises a generally tubular hard plastic shell  66  and an elastomeric septum  68 . The components of seal plug assembly  64  are formed from materials to exhibit a combined density less than the density of the red blood cells, but greater than the density of the serum. Shell  66  includes a top end  70 , a bottom end  72  and a central passage  74  extending continuously between the ends. Annular sealing ribs  76  and  78  extend outwardly from shell  66  at locations near bottom end  72 . Annular sealing ribs  76  and  78  define diameters approximately equal to inside diameter “c” of liner  22 . Shell  66  further includes an outwardly projecting annular septum flange  80  substantially adjacent top end  70  and an annular closure engagement wall  82  between sealing flange  78  and septum flange  80 . Closure engagement wall  82  defines an outside diameter that is substantially equal to the inside diameter of plug recess  52  of tube closure  34 . A cylindrical wall  84  extends between closure engagement wall  82  and sealing flange  78 . Cylindrical wall  84  defines an outside diameter approximately equal to the inside diameter “f” defined by flexible release walls  54  of tube closure  34 . Additionally, cylindrical wall  84  defines a length approximately equal to the axial length of cylindrically generated portions  56  of flexible release walls  54 . 
     Elastomeric septum  68  is molded unitarily from a rupturable elastomeric material such as Kraton copolymer, a urethane or PVC. Septum  68  includes a bottom  86 , a generally cylindrical side wall  88  extending upwardly from bottom wall  86  and an initially conically convex top wall  90  extending upwardly from cylindrical side wall  88 . A shell recess  92  extends centrally into bottom  86  of septum  68 . Shell recess  92  includes a small diameter entry having a length substantially equal to the axial distance between septum flange  80  and closure engaging flange  82  on shell  66 . Shell recess  92  further includes a large diameter portion that dimensionally conforms to axial and diametric dimensions of septum flange  80  on shell  66 . 
     Device  10  is assembled by slidably inserting liner  22  into shell  12  until flange  30  of liner  22  is seated against annular rim  20  of tube  12 . As noted above, outside diameter “d” of cylindrical side wall  28  of liner  22  is less than inside diameter “a” of cylindrical side wall  18  of tube  12 . Accordingly, an annular space will exist between liner  22  and tube  12  at locations between annular rim  20  of tube  12  and closed bottom  16  thereof. 
     Seal plug  64  then may be assembled by mounting elastomeric septum  68  over top  70  of shell  66 . More particularly, septum flange  80  is forcibly urged into shell recess  92  in open bottom  86  of septum  68 . Small diameter portions of recess  92  will resiliently engage around portions of shell  66  between septum flange  80  and closure engagement flange  82 . Seal plug assembly  64  then is urged into plug recess  52  in bottom end  38  of tube closure  34 . This will require an initial outward stretching of portions of tube closure  34  adjacent bottom end  38 . However, tube closure  34  will resiliently return toward an undeflected condition with flexible release walls  54  engaged around cylindrical wall  84  between closure engagement flange  82  and annular sealing flange  78 . Additionally, conical surface  62  in shell recess  52  of tube closure  34  will cause convexly conical top wall  90  of septum  68  to deflect into concave configuration in nested engagement with conical surface  62 . 
     The assembly of closure  34  and seal plug  64  then is inserted into open top end  14  of tube  12 . Sufficient insertion causes annular sealing flanges  76  and  78  of shell  66  to sealingly engage in liner  22 . Liner end seat  44  will seat substantially adjacent open top  24  of liner  22 . Tube stopper  46  will compress into tight sealing engagement with inner circumferential portions of tube  12  between liner  22  and open top  14  of tube  12 . Insertion of tube closure  43  into tube  12  will terminate when tube end seat  42  seats against open top  14  of tube  12 . 
     A needle cannula  94  is used to insert a sample of blood into device  10 . More particularly, as shown most clearly in FIG. 3, needle cannula  94  is urged centrally into recess  40  at top end  36  of tube closure  34 . Continued advancement of needle cannula  94  will cause a rupturing of conical top wall  90  of septum  68 . An appropriate volume of blood  96  then is delivered from needle cannula  94  into liner  22 . Portions of tube closure  34  adjacent recess  40  will self-seal upon removal of needle cannula  94 . However, conical top wall  90  of septum  68  will remain with a defect. 
     Device  10  with blood  96  therein then is placed in a centrifuge which places a centrifugal load on device  10 . The centrifugal load deflects flexible release walls  54  sufficiently downwardly to permit separation of seal plug assembly  64  from tube closure  34 . Simultaneously, the centrifugal load causes an outward deflection of tubular sidewall  28  of elastomeric tube liner  22 . This outward deflection of liner  22  permits seal plug  64  to move toward closed bottoms  26  and  16  of liner  22  and tube  12  respectively. Air will be trapped in passage  74  of shell  66  approximately when bottom end  72  of shell  66  contacts blood  96 . This trapped air could restrict further downward movement of seal plug  64 . However, the defect in septum  68  caused by needle cannula  94  defines a path through which trapped air may escape passage  74 . Thus, seal plug  64  is permitted to sink into blood  96 . 
     The centrifugal load created by the centrifuge also separates serum from red blood cells in blood  96 . Thus, red blood cells, under the action of the centrifugal load, migrate around seal plug assembly  64  and toward closed bottom  26  of liner  22 . Simultaneously, the less dense serum of blood  96  will flow between shell  66  and outwardly deformed portions of elastomeric liner  22  as shown schematically in FIG.  5 . 
     Seal plug  64  will stabilize at a position in liner  22  between the red blood cells and the serum of blood  96 . This stabilized position is attributable to the formation of seal plug  66  and septum  68  from materials that will give seal plug  64  a density less than the density of the red blood cells, but greater than the density of the serum. After a specified time, the centrifuge will be stopped. The absence of the centrifugal load will cause side wall  28  of elastomeric liner  22  to resiliently return toward an undeformed condition and into tight sealing engagement with annular sealing flanges  76  and  78  of shell  66 . Thus, red blood cells  98  will be sealed between seal plug  64  and closed bottom  26  of liner  22 , while serum  100  will lie between seal plug  64  and closure  34 . 
     Liner  22  is compatible with most of the numerous additives used in sample collection tubes such as citrates, silicates, EDTA and the like that are used to condition the fluid sample either to facilitate or retard clotting, or to preserve the fluid sample for a particular analysis. It is within the purview of this invention that one or more additives may be used in the present invention for particular applications. 
     EXAMPLE 1 
     The separator of the present invention was made as follows: The three components of the separator, a liner, a septum, a seal plug were made by injection molding. The liner was made from DuPont Dow Engage® 8400 polyolefin elastomer with a flexural modulus of 22 MPa. The septum was made from GLS Dynaflex® 2712 (Dynaflex is a trademark of and manufactured by GLS Corp., Cory, Ill.), with a specific gravity of 0.889. The seal plug was made from Bayer Lustran® 348 ABS with a specific gravity of 1.06. 
     EXAMPLE 2 
     The separator of the present invention was made as follows: The two components of the separator, the liner was injection molded from DuPont Dow Engage® 8411 polyolefin elastomer with a flexural modulus of 29 MPa and the seal plug was injection molded from Bayer Lustran ® 348 with a specific gravity of 1.06. 
     EXAMPLE 3 
     The separator of the present invention was made as follows: The three components of the separator, the liner was injection molded from DuPont Dow Engage® 8411 polyolefin elastomer with a flexural modulus of 29 MPa and the septum was made from a Kraton® elastomer and the seal plug injection molded from Bayer Lustram® 348 with a specific gravity of 1.06.