Blood collection tube with separation barrier

PATENT APPLICATION FOR: BLOOD COLLECTION TUBE WITH SEPARATION BARRIER

BACKGROUND OF THE INVENTION

Field of the Invention

The invention relates to body fluid collection containment devices and methods, in particular improved blood collection tubes and methods, capable of separating phases of different density using a porous separating medium.

Discussion of the Related Art

Fluid col lection tubes containing a thixotropic gel for separating phases of different densities, e.g., in blood, are well known. See, e.g., U.S. Patents Nos. 3,997,442, 4,257,886, 4,426,290, 4,770,779, and 6,238,578. The gel is selected to have a density between the phases of blood which are to be separated. Upon centrifugation of a collected blood sample, the force of centrifugation causes movement of the gel from a substantial ly non-flowing state to a flowable state. In the flowable state, the gel migrates to a position between the two phases, e.g., between plasma / serum and hematocrit /clot portions. Upon cessation of centrifugation, the gel becomes substantial ly non-flowable, thereby maintaining the separation between phases. Therefore gel separators can be described as dynamic separators as they move during centrifugation. Gel movement, i .e., getting adequate movement and positioning of the gel upon centrifugation, and resolution or degree of separation within the liquid portions of biological samples can sometimes be an issue for the in-vitro diagnostic field. These devices also require special manufacturing equipment to prepare the gel and to fil l the tubes.

US 3,972,812 discloses another type of dynamic separator in which a porous disc is inserted into the top of a collection tube containing a previously collected and clotted blood sample. The porous disc has a specific density such that it descends down the tube during centrifugation until it reaches the serum-clot interface. The disc filters the clotted portions of blood (fi brin and cel l ular material) from the serum portion of blood. The insertion of the porous disc after collection of the sample exposes the operator to the blood sample. In addition getting adequate movement of the porous disc upon centrifugation relies on the porous disc having a specific density ( 1 .03 to 1.09 g/cc) intermediate to the materials to be separated. SUMMARY OF THE INVENTION

The various embodiments of the present invention are directed to an improved fluid coi Section container such as a blood collection tube, containing a porous material separation medium which provides a barrier to maintain separation of a sample such as blood after centri fugation, with minimal or no interference with common diagnostic anaiytes.

According to one embodiment of the present invention, a blood coliection tube comprises a first end and a second end and a sidewall between the first and second end having inner and outer walls. A separator made from a porous material and a pierceable closure to seal the first end. The porous material having a pore size such that all components of whole blood can flow through the separator.

In an additional embodiment of the present invention, a blood collection tube comprises a first end and a second end and a sidewall between the first and second end having inner and outer walls and a separator comprising a porous material and a pierceable closure to seal the first end. The porous material having a pore size greater than the particle size of all components of whole blood.

In another embodiment of the present invention, a blood collection tube comprises a first end and a second end and a sidewal l between the first and second end haying inner and outer walls and a separator affixed to the inner wall of the sidewall such that the separator does not substantially move in relation to said blood collection tube during centrifugation . The separator is composed of a porous material greater than the particle size of all components of whole blood. A pierceable closure seals the first end, wherein the blood collection tube is partial ly evacuated.

In a further embodiment of the present invention, a blood collection tube comprises a first end and a second end and a sidewal l between the first and second end having inner and outer wal ls and a separator comprising a porous materia! having a pore size that is greater than the particle size of all components of whole blood. A pierceable closure seals the first end, wherein the separator moves from a first position to a second position along the longitudinal axis of the blood col lection tube during centrifugation and wherein the blood collection tube is partially evacuated.

In a further embodiment of the present invention, a blood col lection tube comprises a first end and a second end and a sidewall between the first and second end having inner and outer wal l s, a separator and a pierceabie closure to seal the first end. The separator having a static component having a top end and bottom end and a lumen extending therethrough, which is fixed to the inner wall of the side wal l such that the static component remains substantially immobile during centrifugation, and a dynamic component which moves from a first position above the static component to a second position overlaying the top end of the static component during centrifugation. The separator further comprising a porous material,

BRIEF DESCRIPTION OF THE DRAWINGS

Fig. 1 is a cross sectional view of an evacuated blood collection tube containing a separator of porous material according to an embodiment of the invention.

Fig. 2 is a cross sectional view of an evacuated blood collection tube containing a separator of porous material from the perspective of Z-Z as shown in Fig. 1 according to an embodiment of the invention.

Fig. 3 is a cross sectional view of a blood col lection tube containing a separator of porous materia! after sample collection but before centrifugation according to an embodiment of the invention.

Fig. 4 is a cross sectional view of a blood collection tube containing a separator of porous material after centrifugation according to an embodiment of the invention.

Fig. 5 is a cross sectional view of a blood collection tube containing a dynamic separator in the first collection position according to an embodiment of the invention.

Fig. 6 is a cross sectional view of the blood collection tube in Fig. 5 after

centrifugation, with the dynamic separator in the second separation position.

Fig. 7 is a cross sectional view of an alternate embodiment of a blood collection tube having a dynamic separator in the first collection position, in accordance with an embodiment of the invention.

Fig. 8 is a cross sectional view of the blood col lection tube of Fig. 7 after

centrifugation, with the dynamic separator in the second separation position.

Fig. 9 is a partial cross sectional view of a blood collection tube having a dynamic separator in the first collection position, in accordance with an embodiment of the invention. Fig. 10 is a cross sectional view of a blood collection tube having a dynamic separator in the first col lection posiiion, in accordance with an embodiment of the invention.

Fig. 1 1 is a cross sectional and plan view of the dynamic separator of Fig. 10 in the undeformed state.

Fig. 12 is a cross sectional view of an alternate embodiment of a dynamic separator in the first collection position, in accordance with an embodiment of the invention.

Fig. 13 is a cross sectional view of the dynamic separator of Fig. 12 during collection,

Fig. 14a is a cross sectional view of an alternate embodiment of a dynamic separator in the first col lection position, in accordance with an embodiment of the invention.

Fig. 34b is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 14a in the second separation position.

Fig. 15a is a cross sectional view of an alternate embodiment of a dynamic separator, in accordance with an embodiment of the invention, just after launch during centrifugation.

Fig. 15b is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 15a in the second separation position.

Fig. 16 is a perspective view of an alternate embodiment of a dynamic separator.

Fig. 17 is a cross sectional view of the dynamic separator of Fig. 16.

Fig. 1 8 is a cross sectional view of a dynamic separator, in accordance with another embodiment of the invention.

Fig. 19 i s a cross sectional view of a blood collection tube having a dynamic separator of Fig. 16 in the first collection position.

Fig. 20 is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 16 in the second separation position.

Fig. 21 is a plan view of an alternate embodiment of a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 22 is a cross sectional view of a portion of a blood collection tube having a dynamic separator of Fig. 21 in the first collection position. Fig. 23 is a cross sectional view of a blood col lection tube having a dynamic separator of Fig. 21 in the second separation position.

Fig. 24 is a perspective view of an alternate embodiment of a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 25 is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 24 in the first collection position.

Fig. 26 is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 21 in the second separation position prior to closure of the diverter valve.

Fig. 27 is a cross sectional view of a blood collection tube having a dynamic separator of Fig. 21 in the second separation position after closure of the diverter valve.

Fig. 28 is a perspective view of a blood collection tube having a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 29 is a perspective view of an alternate embodiment of a dynamic separator of the present invention.

Fig. 30 is a perspective view of an alternate embodiment of a dynamic separator of the present invention.

Fig. 3 1 is a cross sectional view of a blood collection tube having a dynamic separator in the first col lection position, in accordance with an embodiment of the invention.

Fig, 32 is a cross sectional view of a blood collection tube, having a dynamic separator of Fig, 31 after sample collection but before centrifugation.

Fig. 33 is a cross sectional view of blood collection tube having a dynamic separator of Fig. 3 1 in the second separation position.

Fig. 34 is a cross sectional view of an alternate embodiment of a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 35 is a cross sectional view of an alternate embodiment of a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 36 is a cross sectional view of a blood collection tube having a dynamic separator in the first col lection position, in accordance with an embodiment of the invention. Fig. 37 is a cross sectional view of blood collection tube having a dynamic separator of Fig. 36 after centrifugation, in the second separation position.

Fig. 38 is a cross sectional view of a blood collection tube having a dynamic separator in the first col lection position, in accordance with an embodiment of the present invention.

Fig. 39 is a cross sectional view of blood col lection tube having the dynamic separator of Fig. 38 after centrifugation, in the second separation position.

Fig. 40 is a cross sectional view of blood collection tube having a dynamic separator in the first col lection position, in accordance with an embodiment of the invention.

Fig. 43 is a cross sectional view of a blood col lection tube having the dynamic separator of Fig. 40 after centrifugation, in the. second separation position.

Fig. 42 is a cross sectional view of a blood collection tube having a dynamic separator in the first coi leciion position, in accordance with an embodiment of the invention.

Fig. 43 is a cross sectional view of a blood col lection tube having the dynamic separator of Fig. 42 after centrifugation, in the second separation position.

Fig, 44 is a cross sectional view of an alternate embodiment of a dynamic separator in the first collection position, in accordance with an embodiment of the invention.

Fig. 45 is a cross sectional view of a dynamic separator, in accordance with an embodiment of the present invention.

Fig. 46 is a cross sectional view of an alternate embodiment of a dynamic separator of the present invention in the first collection position.

Fig. 47 is a cross sectional view of a tube containing a separator which functions by a combination of both dynamic and static modes in, a first col lection position, according to an embodiment of the invention.

Fig. 48 is a cross sectional view of the combination dynamic and static separator of Fig. 47 after centrifugation, in the second separation position.

Fig. 49 is a side view of an alternate embodiment of a combination dynamic and static separator in a first collection position, in accordance with an embodiment of the invention.

Fig. 50 is a side view of the combination dynamic and static separator of Fig. 49 after centrifugation, in the second separation position. Fig. 51 is a cross sectional view of the combination dynamic and static separator of Fig. 50 in the first collection position.

Fig. 52 is a cross sectional view of the combination dynamic and static separator of Fig. 50 in the second separation position.

Fig. 53 is a cross sectional view of a blood collection tube having an alternate embodiment of a separator, in accordance with an embodiment of the present invention.

Fig. 54 is a cross sectional view of the blood collection tube of Fig. 53 during centrifugation.

Fig. 55 is a cross sectional view of a blood collection tube having a separator, in accordance with an embodiment of the present invention.

Fig. 56 is a cross sectional view of a blood collection tube of Fig. 53, wherein the separator is in the second separation position.

Fig. 57 shows a bar chart of Platelet counts (PLT) for prototype tubes and a control tube after centrifugation at 1300 x g for 10 minutes, in accordance with an embodiment of the invention.

Fig. 58 shows a table of various clinical parameters for prototype tubes and a control tube, under various centrifuge conditions, in accordance with an embodiment of the in vention.

Fig. 59 shows a bar chart of Platelet count (PLT) results as shown in Fig. 58 for prototype tubes and a control tube after centrifugation at 1500 x g for 10 minutes, in accordance with an embodiment of the invention.

DETAILED DESCRIPTION OF THE INVENTION

It wi l l be readily understood that the components of the present disclosure, as general ly described and further il lustrated in the figures herein, could be arranged and designed in a wide variety of different configurations. Thus, the following more detai led description, as represented in the figures, is not intended to limit the scope of the disclosure, but is merely a representation of exemplary combinations of the components. A col lection container such as a blood col lection tube in accordance with an embodiment of the invention is shown in Figure 1 , The tube 10 contains an open upper end 12, a lower closed end 14, and sidewalls 16 having an inner wall 18 and an outer wall 20. A separator 22 of length L is fixed to the inner wall 18 within the container, thereby forming an upper 25 and lower reservoir 26.

The tube 10 is provided with a closure such as a pierceable closure or stopper 24, which may be pierced by the non-patient end of a blood collection needle such as a double- ended blood coliection needle commonly available in the art. The tube 10 may be evacuated, such that upon piercing by such a needle, blood is drawn from a patient's vasculature and into the tube 10. Detai ls of evacuated and non-evacuated blood col lection tubes and blood col iection are well known to those skil led in the art.

As noted above, fol lowing sample collection, the tube is centrifuged causing portions of the blood sample to migrate based on their relative densities or specific gravities.

Typically, upon sufficient centrifugation forces and duration, the blood sample separates into two phases of the blood sample, e.g., a generally acellular plasma and a cellular portion, typical ly i nclusi ve of erythrocytes (red blood cells), granulocytes, lymphocytes and monocytes as is commonly known in the art.

The various embodiments of the invention provide a separator in a blood col lection tube which provides an improved separation of a blood sample and an inert barrier which shows acceptable or no levels of assay interference and that confers therapeutic drug moni toring capabi lity when compared to a gel separation medium.

According to embodiments of the invention, the separator is typically a porous material made of e.g. an open cell foam, a foamed rubber, a fleece, a mat, a honeycomb-like material or the like. The separator may be a monolithic single piece of porous material or can be an component containing a porous material. The porous material is inert (e.g. chemically and biologically), which means that i t wi l l have no or minimal effect on the blood or plasma or serum sample which would resul t in an analysis which would differ beyond clinically acceptable limits from that made on the same sample if the porous material was not used. The pore size of the porous material separator is sized and configured such that all the components of whole blood (including the largest e.g. the cellular components desired to be removed from the upper reservoir) may pass through the pores of the separator into the lower reservoir. This includes by way of example a separator comprising a minimum pore size of e.g. 10 microns and in the range of 100 to 250 microns, preferably 190 to 220 microns. Thus, the porous material does not act as a filter, wherein a retentate and a filtrate are distinguished from each other. Examples of suitable materials are Basotect® V 3012 and Basotect® UF Melami ne based (e.g. a formaldehyde-melamine-sodium bisulfite copolymer) open cell foams available from BASF or foam S6050HY by KOEPP Schaum GmbH, Germany available in reticulated and non-reticulated form, or Polydamp® Hydrophobic Melamine (PHM) Foam available from Polymer Technologies, Inc. Newark, Delaware.

The tube 10 can be made of glass, plastic or other suitable materials. Some preferred materials used to manufacture blood col lection tube 10 include polypropylene, polyethylene, polyethyleneterephthalate, polystyrene, polycarbonate and cellulosics. More expensive plastics such as polytetrafluoroethylene and other fluorinated polymers may also be used. In addition to the materials mentioned above, examples of other suitable materials include polyolefins, polyamides, polyesters, silicones, polyurethanes, epoxies, acrylics, polyaerylates, polysulfones, polymethacrylates, PEEK, polyimide and fluoropolymers such as PTFE

Teflon®, FEP Teflon®, Tefzei®, polyvinylidene fluoride (PVDF) and perfluoroalkoxy resins. Glass products incl uding si lica glass may also be used to manufacture the collection devices. One exemplary glass product is PYREX® (avai lable from Corning Glass, Corning, New York). Ceramic collection devices can be used in accordance with embodiments of the invention. Cellulosic products such as paper and reinforced paper containers can also be used to form collection devices according to the invention.

The tube of the invention typical ly goes through additional processing steps either before or after the separator is inserted into the tube. For example, additives useful in blood or urine analysis, e.g., procoagulants or anticoagulants may be disposed Into the tube. As known in the art, blood analysis is often performed on serum, and procoagulants are typically used to enhance the rate of clotting. Such procoagulants include silica particles or enzyme cl ot activators such as elagic acid, fibrinogen and thrombin. If plasma is desired for analysis, an anticoagu l ant is general ly used to inhibit coagulation, such that blood cells can be separated by centrifugation. Such anticoagu lants include oxalates, citrate, EDTA, EGTA, BAPA, and BAPTA, heparin and Boophi lin. Additives are disposed in the containers in any suitable manner, liquid or solid, including dissolution in a solvent, or disposing in powdered, crystallized, or lyophilized form. It should be noted that a porous material (e.g. an open cell foam), may be used to cause the activation of the coagulation process instead of or together with the use of a procoagulant additive. The porous material is placed in the fluid path of blood collection, such that contact with the porous material enhances activation (intrinsic pathway) of platelets, thus accelerating clotting needed for serum col lection. The porous material may or may not need to act as a separator in such an embodiment.

Then, after any such additional additives are put into the tube, the tube (or group of tubes) is subjected to an evacuated chamber with a pressure below atmospheric pressure. A seal such as an elastomeric stopper or pierceable membrane is applied, and the tube is sterilized by a process such as irradiation (e.g., with cobal t 60 radiation), ethylene oxide gas exposure, or electron-beam exposure. (Note that several of these steps may be performed in an order other than that presented above).

In accordance with an embodiment of the invention, the blood collection tubes are capable of being formed in various shapes and in any desired size. For example, standard blood collection tubes with OD's of 13mm and 16 mm and lengths of 75mm, and 100mm are contemplated.

Figure 3 shows the axial separation position of a separator 22 along the longitudinal ax is of the tube after centrifugation such that the separator is located at or near the expected limit of the hematocrit content 34 i .e. the position at which the interface between the hematocri t (the cel l ular portion) and plasma or serum (non-cel lular liquid portion) occurs after centrifugation of a whole blood sample. For example, Dimension A (as shown in Figure 1 ) can be in the range of 10-60 mm, for example. In Figure 3 Dimension A is approximately 45 mm for a blood collection tube having an inner diameter (ID) of approximately 7.5mm, OD of 13mm, length of 100mm and a draw volume of 4.5 mis of blood.

Length L of a separator is typical ly proportional to the internal volume of the tube. For example, a separator length range of 6 to 50mm may be used for tube having an ID of approximately 7.5 mm, length of 100mm and a draw volume of 4.5 mls of blood.

The separators of various embodiments of the invention function by a dynamic mode, a static mode, or a combination of both dynamic and static modes, in order to form a barrier after centrifugation at the axial separation position along the longitudinal axis of the tube at which interface between the general ly acellular plasma and a cellular portion of the separated blood sample occurs.

A static separator or passive barrier is fixed at the axia! separation position prior to col lection or centrifugation and as such does not substantial ly move along the longitudinal axis of the blood collection tube during centrifugation. in contrast, to dynamic separators, the immobility of a static separator means that the overall density of the static separator may be independent of (and not intermediate of) the materials to be separated. Thus, the density of a static separator can be less than 1 .03 g/cc (for example Basotect® V 3012 foam has a density of 0.009 g/cc) or greater than 1 .09 g/cc and still function as a separator of whole blood.

A dynamic separator or active barrier moves during centrifugation from a first col lection position to the axial separation position along the tube length. This dynamic mode necessitates that the overall density of a dynamic separator typically has a specific density in the range of 1 .03 to 1.09 g/cc to be intermediate to that of the materials to be separated such as whole blood for example, in order to function.

Separators which function using a combination of dynamic and static modes will have a fixed static component or subcomponent and a dynamic component or sub component.

Figures 1 to 4 snow an embodiment of a static separator. The static separator 22 is preferably fixed to the inner wall 18 within the blood collection tube by for example a frictional, chemical and/or mechanical interaction or configuration such that the separator 22 does not substantial ly move in relation to the blood collection tube during centrifugation of a sample. For example, the separator is substantially stationary or immobile along the longitudinal axis of the blood collection tube during collection and separation of a sample. A chemical interaction includes for example adhesives, however care must be taken in the selection of a sui table adhesive which must be capable of adhering the separator to the inner wal l 18 while also remaining inert in the same manner as previously described for the porous material . A mechanical configuration includes for example using a friction force exerted between the inner wal l 18 and the separator 22 sufficient to prevent axial movement of the separator towards the bottom of the tube during centrifugation. An example includes using a separator wi th an outer diameter (OD) that is larger than the internal diameter (ID) of tube (e.g. a separator of OD 10mm may be used with a tube ID of 7.5mm) which may also have a modulus of elasticity sufficient to provide an axial force and/or coefficient of friction to resist or prevent axial movement as described above. Additional mechanical configurations may comprise interference engagement between the separator and a portion of the tube or structure attached or fi xed thereto, for example a separator may be placed in the blood collection tube mold prior to molding of the blood col lection tube, and the blood col lection tube then molded around i t. Other examples include the use of flanges or posts circumferentially spaced on the tube inner wall 18 on which the separator can rest or engage. Furthermore, the blood col lection tube may be formed such that the separator is integral with the blood collection tube.

Figure 2 shows an intimate contact around the entire circumference of the OD of the separator along its length (L) with the inner wall 1 8 of the blood collection tube such that there are effectively no gaps between separator and the inner wall. Thus, the inner wall 18 occludes a portion of the pores in the separator due to for example an interference fit between the separator 22 and the inner wal l 18 and a sample cannot flow from the upper reservoir 25 to the lower reservoir 26 or from the lower reservoir 26 to the upper reservoir 25 without passing through the separator.

Figure 3 shows the tube after collection and prior to centrifugation. During collection the whole blood sample 30 flows into the upper reservoir 25 and through the separator 22 into the lower reservoir 26 of the tube (thus the separator does not filter the blood) until the desired vol ume of blood has been col lected as dictated by the level of evacuation according to an embodiment of the invention. A densometric separation of the whole blood components occurs during centrifugation, in which al l species are free to move through the porous material until an appropriate equilibrium position is attained as dictated by the density of each component entity. Thus the cellular based components of the highest density (e.g. red blood cells and white blood cells) of whole blood pass through the pores of the porous material to the lower reservoir thereby formi ng the hematocrit region. In addi tion, the separator is positioned such that the lower density blood component species such as platelets have a centrifugation equi librium position winch is in inside the pores of the separator.

Figure 4 shows that after collection and centrifugation the various species in the hematocri t region 33 remain in their density equili brium position in the lower reservoir 26 and are effecti vely prevented from traveling above the separator 22 by gravity, density and the length of the tortuous flow path present in the porous material . In accordance with an embodiment of the i nvention, the species which are located within the separator are retained wi thi n the pores or captured by porous materia! and are prevented from flowing into the upper reservoir. Thus the porous material forms a passive barrier after centrifugation, which improves the separation of plasma from the remainder of the whole blood sample such as platelets, red blood cei ls, white blood cells.

It is generally desirable to direct blood into the blood collection rube such that the blood is able to travel beneath the separator, since serum tubes for some applications require the red blood cells to be below the separator (e.g., where the red blood cells clot and are to be separated from the serum). {This may be less important for plasma tubes.) Such an outcome may be difficult if the separator starts on the bottom of the tube.

Another embodiment of the static separator (as shown in Figs. 1 to 3) provides a solution for this problem by locating / applying the procoagulant (such as silica) in the lower reservoir 26 only (not shown) such that the collected red blood cells would only contact the procoagulant beneath separator 22.

Another solution to this problem is to use a dynamic separator which is designed to be secured at the top of the blood col lection tube, in a first collection position, such that the non- patient end of the needle can be inserted through the separator into the tube interior. The separator would thus be formed of a material pierceabie by such a needle, and of a size/shape to allow the needle to pass completely through. Securement of the separator at the top end of the tube could be attained by molding in a feature that mates with a corresponding feature of the tube closure, or by util izing a friction force /interference fit that holds the separator at the top of the tube prior to centrifugation, and the like. The forces experienced by the separator duri ng centri fugation overcome the securement forces thereby allowing the separator to launch and migrate down the tube to the second separation position.

Figures 5 to 46 show various dynamic separators, according to embodiments of the invention. The overal l density of a dynamic separator is typically in the range of 1.03 to 1 .09 g/cc to be intermediate to that of the materials to be separated such as whole blood for example, in order to effectively move during centrifugation to the axial separation position along the tube length.

These embodiments utilize a col lection container such as a blood collection tube 10, comprising an open end 12, a second end 14 and sidewalls 16 having an inner wall 18 and an outer wal l 20 therebetween, and a pierceabie closure 24 enclosing at least the open end. Figures 5 and 6 show a dynamic separator in accordance with an embodiment of the invention . Separator 122 is made from a porous material plug or cylinder 121 and at least one ballast weight/mass 123 attached or embedded within porous material plug 121. Figure 5 shows that during col lection of blood, the non-patient end of cannula 99 penetrates pierceable cap 24 and is embedded within separator 122. As described above the pore size of porous material plug 121 is such that the blood components can easily flow through the separator to the reservoir 13 without filtration of the unciotted blood components or a back flow due to a flow restriction occurring. The dynamic nature means that the overall density of a separator such as separator 122 has a speci fic density in the range of 1.03 to 1 .09 g/cc to be intermediate to that of the materials to be separated such as whole blood for example. Thus, each bal last weight/mass 123 is made from a high density material (e.g. glass, deliin, polystyrene) and is attached to or embedded in the porous material plug 121 (e.g. Basotect® V 3012 foam having a density of 0.009 g/cc) in order to obtain an overal l separator 122 specific density in the desired range. Prior to centrifugation separator 122 may be retained at the upper end 12 of tube 10 by securement to pierceable cap 24 (by for example a temporary adhesive bond), and/or inner wal l 18 (by for example a sufficient friction force).

Upon centrifugation and immersion of the separator 122 in the fluid, the lower density porous material plug 121 provides a buoyant upward force on the separator 122 relative to the fluid. Simultaneously, the higher density bal last weight(s)/mass 123 provide an axial force downward on the separator. The combined forces stretch and elongate porous material plug 121 axial ly, causing inward radial movement of the middle region. This radial movement pulls the porous material plug 121 out of contact with the inner wall 18 of the tube so that it is free to move axial ly. Alternatively, the porous material plug may not move radially inward, but allows for the centrifugal force to overcome the friction force, until the separator moves towards a position of equilibrium.

Therefore, multiple paths are developed both through the pores of porous material plug 121 and between the inner wall 18 of the tube and the OD of porous material plug 121 that permit the fl ow of the low-density serum component past the separator 122 as it migrates down the tube. Migration of the separator 122 terminates when it reaches the axial separation position between the lower density fluid component and higher density fluid or cel lular/solid components, equal to its overall density. Upon terminating centrifugation, porous material plug 121 expands to i ts undeformed shape, sealing against the inner wall 18 of the tube, thereby creating a barrier between the higher and lower density components of the sample fluid as shown in Figure 6. After centrifugation the various species in the hematocrit region 33 remain in their density equi librium position in the lower reservoir 26 and are prevented from traveling above the separator 1 22 by gravity, density and the length of the tortuous flow path present in the porous material. The species which are located within the separator 122 are retained within the pores or captured by porous material and are restrained from flowing into the upper reservoir. Thus, the porous material forms a passive barrier after centrifugation, which improves the separation of serum or plasma from the remainder of the whole blood sample such as platelets, red blood cel ls, white blood cells.

Figures 7 and 8 show another embodiment of a dynamic separator which is similar to that shown in Figures 5 and 6. Separator 132 is another dynamic separator having a porous material plug 13 1 and at least one bal last weight/mass 133 attached or embedded within porous material plug 131. The difference to that of the previous embodiment is that a recess 135 is present in the lower end of porous material plug 131 to provide a reduced cross section and assist allowing the non patient cannula to completely penetrate separator 132 when in the first col lection position. Thus, during col lection, needle cannula 99 penetrates pierceable cap 24 and separator 132 al lowing blood to flow directly into reservoir 13 of the tube through recess 1 35 as shown in Figure 7.

Figure 9 shows an embodiment of the invention in which a porous material dynamic separator is located adjacent a tube closure 224 of reduced cross sectional thickness when compared to conventional pierceable closure 24. The pierceable closure structure may be a foi l or a combination of a pierceable closure and foil as shown in US Patent 5,061 ,263 for example. Dynamic separator 142 has the same composition as separator 122 in Figure 5 and li kewise has overall separator specific density in the desired range. The reduced cross section of tube closure 224 together with the location of separator 142 adjacent tube closure 224 facili tates a greater depth of penetration of the non-patient cannula 99 either deep into separator 142 simi lar to that shown in Figure 5 or more preferably completely through the separator 142 during normal phlebotomy as shown in Figure 9.

Figures 10 and 1 1 show another embodiment of a dynamic separator. Dynamic separator 152 is made from a porous material plug 151 and a wiper 153. Wiper 153 has a disc portion 154 havi ng holes 155 or slits through the cross section and a taper section 156 (that tapers toward the upper end 12 of the tube in an opening fashion) around the outer circumference of disc portion 154, The disc portion 154 is bonded or attached to the lower surface of the porous material plug 151 . Wiper 153 is made from an elastomer (such as for example bromo butyl rubber or a thermoplastic elastomer) and fulfills the same function as the bal l ast weight/mass element in the previous dynamic separator embodiments in order to ensure that the overal l separator 152 specific density is in a desired range. The dynamic separator 1 52 is placed into the upper end 12 of tube 10, such that porous material plug 151 is projecting from the top of tube 12. Pierceable closure 24 is then inserted into and thereby seals upper end 12 in a manner that also compresses the porous material plug 151 to a reduced cross sectional thickness (L1 ) without moving the wiper 153. Wiper 153 is held in posi tion by either a friction force or a temporary adhesive bond between taper section 1 56 and inner wal l 1 8 of the tube 12.

On col lection a non patient cannula is able to penetrate deep into the porous material plug 15 ! such that blood will flow along the pores of porous material plug 151 , through the holes 155 of wiper 153, and into the reservoir 13 of tube 12. Upon centrifugation, the centrifugal forces stretch and elongate porous material plug 151 axially, causing inward radial movement such that the friction force between taper section 156 and inner wall 18 of the tube 12 is overcome and separator 152 begins to travel axially down the longitudinal, axis of tube 12. Taper section 156 can maintain radial engagement or move out of contact with the inner wall 18 of the tube 12 during movement. Separator 152 continues to migrate down the tube until the separation position is reached as determined by the overall separator density. On completion of centrifugation, porous material plug 151 expands to its undeformed shape and cross-sectional thickness (L2), thereby exerting taper section 156 to seal against the inner wail 18 of the tube, thereby creating another barrier in addition to the porous material plug 151.

Figures 12 and 13 show another embodiment of a dynamic separator. This embodiment comprises a separator 172, comprising a porous material plug 171 surrounded by a non-porous material (such as an elastomer) sleeve 173. The porous material plug 171 is preferably tapered from a larger diameter at the bottom to smaller diameter at the top, in a frusto-conical manner. A barrier cap 175 is secured to top of the porous material plug 171 which is made from a material prevents the porous material plug 171 from being penetrated by a cannula and forms a liquid seal with sleeve 173. The non-porous material of sleeve 173 has a specific gravity large enough to ensure the overall separator 172 specific density in the desired range, as wel l as providing sealing capability between the inner wall 18 within the blood collection tube and the separator 172. The porous material plug 171 is in intimate contact with the sleeve 173 and may be bonded together, or held together by, for example a mechanical means at the lower portion of separator 172, Prior to collection separator 172 is in a first position within the internal reservoir of the blood collection tube adjacent to or towards the open end of the blood collection tube and pierceabie closure. Upon collection, a non-patient needle 99 is inserted into and penetrates the pierceabie closure of the tube (not shown) and impacts the barrier cap 175, causing partial compression of the porous material plug 171 , breaking the fluid seal between barrier cap 175 and sleeve 173 thus allowing blood to flow both through porous material plug 171 and at the interface between the porous material plug 171 and sleeve 173, and into the lower chamber of the blood collection tube, Preferabl y, the force needed to compress the porous material plug 171 to a degree of compression substantial enough to al low for blood to flow through the separator 172 is less than the friction force between the sleeve 173 and the inner wall 18 of the tube 10.

During centrifugation, the magnitude of the barrier cap 175 specific density causes the porous material pl ug 171 to deform to al low the flow of blood components of lower density to flow through and above the separator 172. After centrifugation, porous material plug 171 recovers to the original undeformed state such that barrier cap 175 reforms a fluid seal with sleeve 173 to form a barrier at the second separation position between the cel lular and non- cel lular components of blood,

In an alternate embodiment the barrier cap may not completely occlude the top of the porous material plug such that the barrier cap does not form a complete fluid seal with sleeve in the undeformed state and a tortuous flow path would exist through the separator.

Figures 14 and 15 show an embodiment of the invention in which a cylindrical funnel structure 186 Is engaged into the pierceabie closure 324 of the blood collection tube 10, and at least partially extends into or completely through the length of dynamic separator 182 when dynamic separator 182 is positioned in a first position substantially adjacent pierceabie closure 324 (See Figure 14a). Dynamic separator 182 has, for example the same composition as separator 122 in Figure 5. A slit or small opening through the cross section and along the center a x is of the porous material plug, allows for cylindrical structure 186 to at least partially, preferably completely, extend into the separator 182. Separator 182 is retained on cylindrical structure 186 by friction or by a teniporar)' adhesive bond. During collection, a non-patient needle 99 is inserted into the pierceabie closure 324. The tip of the non-patient need!e protrudes through the pierceable closure 324, interna! to the cylindrical structure 186, preferably through or bypassing separator 182 depending on the depth of penetration of the cylindrical structure 186 into separator 1 82. During centrifugation, the friction forces between the cylindrical structure 186 and the separator 182 are low enough to allow separator 1 82 to launch from the first position and to the second separation position, wherein the separator 182 overlays the partially formed cellular material of the sample (See Figure 14b).

A fu l ly cored inner section 187 of the porous material plug along the center axis can be removed from the separator in place of a slit. However the removal of a core from the separator 182a requires that the inner wal l 318 of the tube 10 to be tapered or comprise ramps that provide for tapering from a larger inside diameter to a smaller inside diameter when moving from the open end to the second end of the tube. Thus, during centrifugation, as the separator 182a moves from the first position to the second separation position (See Figure 15a), the taper of the sidewal! of the tube provides for radial compression of the separator, wherein the slit or cored opening compresses, thereby restricting flow of cellular material through the separator to the pore structure of the porous material only (See Figure 15b).

Figures 16 to 20 show two versions of an embodiment of a dynamic separator. Figure 17 shows a separator 192a in which a ballast mass 193a has a conical shaped, lumen 195a therethrough and is enclosed by a cylindrical sleeve of porous material 191 a which seals against the i nner waii 18 of blood coi lection tube 10. Upon collection a non-patient needle 99 of a blood col lection need le penetrates the pierceable closure 24 and aligns with conical lumen 195a, blood then funnels through the separator 192a via !umen 195a, and into the bottom section of the blood collection tube. Blood may transfer through the base of separator 192a, by mechanisms such as fluid pressure causing temporary distortion for blood to transfer, or a one-way valve (not shown) such as a duckbill valve located at the base of the conical lumen 195a. Blood is then forced to pass through the cylindrical sleeve of porous materia! 191 a during centrifugation as the one way valve mechanism prevents any back flow of blood up the conical l umen 195a.

Figure 18 shows a separator 192b in which an annular ballast mass 193b is attached to the upper surface of a porous material plug 191 b which has a conical shaped lumen 195b extending almost completely through. Upon coilection a non-patient needle 99 of a blood col lection needle penetrates the pierceable closure 24 and aligns with conical lumen 195b, the majority of blood flow then occurs through the separator 192b via lumen ! 95b, and into the bottom section of the blood collection tube. Blood may also transfer through separator 192b by diffusion through the pores of the porous material plug 191 b.

The dynamic separators 192a and 192b have an overall separator specific density in the desired range to ensure movement of the separator under centrifugation forces from a first position (Figure 19) near the open end and pierceable closure to the second separation position (Figure 20).

Figures 21 to 23 show a di fferent embodiment of a dynamic separator which uses a rotational movement to form a barrier during separation. Separator 212 is made from a porous material plug 21 1 and at least one bal last weight/mass 213 attached to the porous material pl ug 21 1 . During collection, separator 212 is located at a first position in the tube wherein ballast mass 213 is attached to the side of porous material plug 21 1 and thus interposed between inner wal l 18 of the blood collection tube 10 and porous material plug 21 1 (see Figures 23 and 24). Therefore col lected blood can flow through the separator 212 via flow channels 215 and 216. The dimensions W and Y of porous material plug 21 1 (when undeformed) are greater than the internal diameter of blood collection tube 10 such that it will not fit unless compressed into the tube opening, wherein the friction and radial compression forces between the separator 212 and Inner wal l 18 maintain the separator 212 in the first collection position , The off-center location of ballast mass 213 causes the center of mass to be offset from the centroid of the separator 212. Thus, during centrifugation a moment is created which overcomes the forces holding separator 212 in the first collection position, which al lows for rotation of the separator 212, wherein the mass 213 moves first and then the porous material pl ug 21 1 fol lows, thereby rotating the separator 212 by 90° into a second separation position (See Figure 23). Once the separator is in the second separation position, there is intimate contact around the entire circumference of the outer diameter of the porous material pl ug 21 1 along dimension W with the inner wall 38 of the blood collection tube such that there are no gaps between porous material plug 21 1 and the inner wail 18, wherein the friction and radial compression forces generated due to the magnitude of dimension Y between the separator 212 and inner wal l 18 maintain the separator 212 in the second separation position.

Figures 24 to 30 show various embodiments of a three part dynamic separator. Figure

24 shows a separator 222 comprising a porous material cylinder 221 , a funnel shaped valve body 223, and a di verier val ve 225 that may resemble a valve pin sealing element or arrangement. Porous material cylinder 221 surrounds the central portion of val ve body 223 and contacts inner wai l 18 of blood col lection tube 10. On col lection the separator 222 is in a first position adjacent tube closure 24. Lumen 227 comprises a funnel shaped upper end that guides blood drawn through tube closure 24 to a channel 228 axially oriented with the diverter valve 225, such that blood flows through separator 222 and exits out of diverter valve 225, Separator 222 is retained in the first position via securement of the top of the valve body 223 to the tube closure and/or a friction force between the outer surface of porous material cylinder 221 on the inner wall 1 8 of blood col lection tube 10.

Upon centrifugation, separator 222 moves downwards to the second separation position (See Figure 26) as the overall separator specific density in the desired range as previously described. Upon arriving at the separation position diverter valve 225 hits either or both of a cel lular portion of the blood sample, or a feature of the tube such as the bottom of the tube, causing the diverter valve 225 to move to the closed position (See Figure 29) which blocks channel 228 such fluid flow from beneath to above separator 222 can only occur by passing through porous material cyl inder 221 either between inner wall 18 and the outer surface of val ve body 223 or through one of the windows 229 in the sidewall of lumen 227. A number of projections 230 from the diverier valve 225 externally engage external portions of the funnel in a cam-follower type of arrangement. The diverter valve 225 is designed to not re-open once i t has closed under normal operating conditions unless an external manual force is applied to both the valve body 223 and the diverter valve 225.

Figures 28 to 30 shows addi tional embodiments (225a, 225b, 225c) of diverter valve design which may have a concave lower surface 225a, an extended length and a concave lower surface 225b, or an extended length and a spherical ball with a flange 225b wherein the OD of the flange is equal to the internal diameter of blood col lection tube 10 a point towards the lower end 14.

Figures 3 1 to 33 show another embodiment of a dynamic separator. Separator 232 has a porous material plug 231 and at least one ballast weight/mass (not shown) attached to or embedded within porous material plug 231 , in addition a flexible and liquid permeable inner container 235 is affixed to the separator 232. inner container 235 has an open top which is attached to the OD of porous material plug 231 , a closed bottom end and a flexible col lapsi ble sidewall extending therebetween. An additive such as e.g. procoagulant or anticoagulant can be spray dried inside inner container 235. Prior to and upon collection separator 232 is located at a first collection position adjacent the pierceable closure 24 (with the inner container 235 hanging beneath) so that a non-patient needle will either completely or deeply penetrate separator 232 to deliver whole blood 30 into the inner container 235 and mix with the addi tive present (see Figure 32).

Upon centrifugation, separator 232 moves down the longitudinal axis of tube 10 under the relative centrifugal force whilst compressing the inner container 235, resulting in the expulsion of the fluid component of the blood through the until such time that no more fluid can be expressed and the separator 232 rests at the second separation position on top of the hematocrit region 33 or cel lular mass resulting in the separation of serum or plasma 32 from the cellular components of whole blood.

One advantage of this embodiment of collecting blood into a permeable inner container is the minimization or el imination adherence of blood cells, red cel ls in particular to the tube wal l which can result in interferences with a number of diagnostic tests. It may also el imi nate the need of using surfactants on the inner surface of the collection tube for mi nimizing red cei l adherence.

Figures 34 to 46 show dynamic separators having a porous material plug and various embodiments of bal last weights/ masses. Figure 34 shows a separator 242 comprising a porous material plug 241 and a cylindrical ring ballast mass 243 circumscribing the porous material pl ug 241 . The ring 243 provides density and mass to enable the movement of the separator system under centrifugal forces against the forces of buoyancy and friction between the open-cel l foam and the sidewal l of the tube. In one embodiment, the porous material plug has a diameter larger than that of the internal diameter of the tube. The ring 243 comprises an outside diameter smaller than the internal diameter of the tube, and the inside diameter of the ring causes "mushrooming" of the porous material plug, thereby establishing interference engagement wi th the inner wall 3 8 of the tube. Figure 35 shows another embodiment in which the porous material plug 241 a may be formed with a cutout portion 246 such that the ring 243 surrounds the porous material plug 241 , without causing "mushrooming" of the porous material plug to engage the inner wal l 18 of the tube.

Figures 36 to 39 show two embodiments of a dynamic separator 252 comprising a porous material plug 251 and one or more bead ballast masses 253. The bead ballast masses 253 are affixed to the porous material plug 251 via friction fit, interference fit, and/or chemical bonding. The beads may be affixed to the top surface of porous material plug 251 (Figures 36 and 37) or sandwiched between two foam portions 251 a 251 b, (Figures 38 and 39) which in some cases may involve foams of differing diameters, material, or pore size.

Figures 40 and 41 show a dynamic separator having a ballast weight/ mass ring 263 positioned on top of the porous material pl ug 261 via friction fit, interference fit, and/or chemical bonding. The ballast mass ri ng 263 may comprise an annular hole, such that the ring does not interfere with the non-patient needle on collection. Porous material plug 261 may also have an annular recess on the upper surface (not shown) to receive ring 263 such that the top surface of the separator is flat.

Figures 42 and 43 show a dynamic separator 272 positioned directly inside pierceable closure 724 of the tube, and mechanical ly or chemically engaged or interfaced to the pierceable closure 724 and not the inner wall 18 of the tube 10. The engagement of separator 272 to the underside of the pierceable closure 724 at the first collection position in this embodiment provides for the possibility of the top surface of the separator being devoid of ceils resting on it upon the launch of the separator 272 during centrif ligation.

Figures 44 and 45 show more embodiments of a dynamic separator directly mechanically or chemically engaged or interfaced to the pierceable closure but also in contact with the inner wal l of the tube. Separator 282 has a feature 285 that mates with a

corresponding feature 287 of the tube closure 824 to retain separator 282 in the first collection position. However centrifugal forces can override the retention features 285 and 287 such that separator 282 may migrate to the second separation position. Figure 45 shows a separator 292 having a first upper porous materia! section 294 and a lower porous material plug section

291 with a plurality of ballast mass beads 293 sandwiched between the sections. Separator

292 is retained in the first collection position by mating features 295 and 297. During centrifugation first upper porous material section 294 may remain engaged to the bottom portion of the pierceable closure 924, while lower porous material plug section 291 and the bal last mass beads 293 launch to act as a dynamic separator. In an alternate embodiment, the first upper porous material section or lower porous material plug section may be partial ly perforated such that it tears to facilitate the launch of the dynamic separator portion.

Figure 46 shows an embodiment of a dynamic separator 312 which utilizes spikes 313 through portions of a porous material pl ug 31 1 , as a means of attachment to the lower surface of pierceable closure 324 of the blood col lection tube 10 to retain separator 312 in the first collection position , The spikes 313 can also function as the ballast mass for the separator 312 or a dedicated bal last mass can be present in addition to the spikes 313 in order to give a separator overall density in the desired range. In embodiments where the spikes 313 also function as the bal last mass, the forces required to remove the spikes 313 from the porous material plug 3 1 1 are larger than the force required to remove the spikes 313 from the pierceable closure 324,

Figures 47 to 56 show a number of separator embodiments which function using a combination of dynamic and static modes. Each embodiment has a fixed static

component/sub assembly and a dynamic component/sub assembly.

Figures 47 and 48 show an embodiment of a separator 412 which has a static component 425 and a dynamic component 426. Static component 425 comprises a cylinder of a porous materia! having a top and bottom end and a lumen 430 along the center axis to provide for a flow path coaxial with the axis of the outside diameter of the cylinder, and the tube. Static component 425 is preferably fixed to the inner wall 1 8 within the blood collection tube by for example a factional, chemical and/or mechanical interaction or configuration such that the static component 425 does not substantially move in relation to the blood collection tube during centrifugation of a sample. Dynamic component 426 comprises a separate and detached pl ug 43 i that shares a similar diameter with that of the static component 425 and a bal last mass 433 that fits wi thin the inside diameter of lumen 430 in the static component

425. Bal last mass 433 such as a glass bead may be attached via adhesive to plug 431. Plug 431 may be made from a porous material such as open cell foam or a non-porous material such as a closed cell foam, or polymer. The overall density of dynamic component 426 is in the specified range for a dynamic separator.

Prior to and upon col lection, dynamic component 426 is secured to inner wall 18 by an adhesive above static component 425 in the first col lection position. The tip of a non- patient needle protrudes through the pierceable closure and blood flow driven by the pressure conditions of the evacuated tube allows for blood to be drawn around dynamic component

426, and generally through the lumen 430 of static component 425. Upon centrifugation, the cel lular material of the blood is driven below and into static component 425, and finally, dynamic component 426 iaunches from the first col lection position (Figure 47) and moves to the second separation position (Figure 48), wherein the plug 431 overlays the top end of static component 425 with ballast mass 433 occl uding lumen 430 to form separator 412. Plug 431 may form a liquid seal with the inner wall 18 if made from a non-porous material or a greater length of torturous path barrier if formed from a porous material.

A laminated sheet of material may be used to adhere plug 431 to inner wall 18 instead of adhesive. However the strength of the bond between laminated sheet and inner wall 18 has to be greater than that between plug 43 1 and laminated sheet such that the laminated sheet remains on inner wall 18 after the launch of dynamic sub-component 426.

Figures 49 to 52 show an embodiment of a separator 512 which has a static component 525 and a dynamic component 526 similar to that shown in Figures 47 and 48. The only difference to that of the previous embodiment is that a dynamic component 526 is unitary to static component 525, such that plug 531 is connected by a living hinge to static component 525. Therefore prior to and upon collection ballast mass 533 rests at the top of lumen 530 i n the static component 525 causing plug 53 Ϊ to be raised in a partially open manner, in the first col lection position (See Figures 49 and 51 ). Thus, col lected blood driven by the pressure conditions of the evacuated tube flows around plug 531 and bal last mass 533, and general ly through the l umen 530 of the static component 525. Upon centrirugation, the cellular material is driven below and into the static component 525, and final ly, the dynamic component 526 pivois from the first position and moves to the second separation closed position (See Figures 50 and 52), wherein the plug 531 overlays the top end of the static component 525 and bal last mass 533 occludes lumen 530. Plug 531 may form a liquid seal with the inner wal l 18 if made from a non-porous material or a greater length of torturous path barrier if formed from a porous material .

Figures 53 and 54 show an embodiment of a separator 612 comprising a porous material plug 61 1 which is suitable for use as a dynamic or static separator. The upper surface 614 of separator 612 is concave, with the concavity having a funnel shaped profile. This may provide for the separator to be used in fixed-angle centrifuges, wherein the tube may rotate about the axis of a centrifuge's rotor in a manner where the axis of the tube is not completely perpendicular to the centrifuge (See Figure 54).

Separator 612 is affixed to inner wall 18 when used as a static separator. A ballast mass is attached to porous material plug 61 1 for use as a dynamic separator in order to provide the required overall specific densi ty to allow for movement during centrifugation to the separation position.

The advantage of the concave profile for the upper surface relates to utilizing a minimum length of porous material needed to provide for an effective barrier, that if parallel on both upper and lower surfaces sides l ike a uniform plug of foam, would otherwise not be able to be used with fixed angle centrifuges without providing for a larger thickness such that the cel l/cel l -free interface would extend through the barrier (and normal to the axis of centrifugation) in both fixed and swing bucket centrifuges.

Figures 55 and 56 show an embodiment of a separator 712 comprising a porous material plug 71 1 which is suitable for use as a dynamic or static separator. Separator 712 comprises a porous material plug 71 1 and a higher density cylindrical straw like structure 713. Cylinder 7 13 is as long if not longer than the porous material plug 71 1 , and interposed between the inner wall 18 of the blood col lection tube 10 and the sidewall of the porous material plug 71 1 to be held in an interference engagement. Upon collection separator 712 is intermediately positioned inside the blood col lection tube 10, such that blood when introduced through the pierceable closure via a non-patient needle, blood rests on the top of the separator 712, and flows through the lumen 715 of cylinder 714 under a combination of vacuum (that diminishes while fill ing) and gravity driving forces. Upon centrifugation, cylinder 7 13, being of a higher density and having less frictional resistance between the porous material p l ug 71 1 and inner wall 18 launches downward to the bottom of the blood col lection tube 10, thereby allowing porous material plug 71 1 to function as the barrier structure with i ntimate contact with inner wal l 18 establ ished around the entire outer circumference.

In static embodiments porous material plug 71 1 remains in the separation position and does not substantial ly move during centrifugation. In dynamic embodiments, the porous materia! plug 71 1 moves before and after the cylindrical structure 713 launches from the porous material plug 71 1. An additional bal last mass (not shown) adhered to the porous material plug 7 1 1 causes the separator to finally position itself at the cell / cel l-free interface.

In an al ternative dynamic embodiment the cylindrical structure is disposed in a fully cored inner section of the porous material plug along the center axis. However the removal of a core from the porous material pl ug requires that the inner wal l of the blood col lection tube to be tapered or comprise ramps that provide for tapering from a larger inside diameter to a smaller inside diameter when moving from the open end to the second end of the tube. Thus, during centrifugation, (after launch of the cylindrical structure) as the separator moves from the first posi tion to the second separation position, the taper of the sidewall of the tube provides for radial compression of the separator, wherein the cored opening compresses, thereby restricting flow of cel lular material through the separator to the pore structure of the porous material only.

EXAMPLES

The fol l owing examples are intended to i ll ustrate embodiments of the invention and are not intended to l i mit the invention.

Example 1

Initial Eval uation of Open Cel l Foam

Prototype tubes were fabricated with three types of foam (Basotect® V3012 and Basotect® UF Melamine based open cell foams available from BASF and PoJydamp® Hydrophobic Melamine Foam PHM foam available from. Polymer Technologies Inc) cut into 10mm OD circular tubes and inserted into unevacuated Lithium-Heparin tubes. These prototypes included two different lengths (L= 25 or 50mm) of separator and two positions (A= 0 or 12 mm) tubes. Citrated sheep' s blood (5 mis) was dispensed into each of the lubes. The tubes were then centrifuged at 1300 x g for 10 minutes and the separator performance was then eval uated .

Whole blood was observed in the Sower reservoir for each type of foam prior to centrifugation. A barrier between hematocri t and plasma was formed by each type of foam, however the positioning of the foam separator was critical to barrier formation as each foam located at the bottom of the tube (A=0) did not form a barrier. In addition a separator length L of 50mm at a position of A= 12mm prevented the addition of 5mls of blood to the tube.

Example 2

First Evaluation of Open Cell Foam using Evacuated Tubes with Human Blood

Method; Prototype tubes were fabricated with two types of foam (Basotect® V 3012 and Basotect® UF Melamine based open cell foams available from BASF) cut into 10mm OD circul ar tubes and inserted into Lithium-Heparin tubes of approximately 7.5mm ED and 13mm OD at a fixed position approximately 45 mm from the bottom of the tube (the expected position of the plasma/hematocrit interface). These prototypes included two different lengths (L= 6 or 12mm) of separator. The prototype tubes were then evacuated to give a 4.5ml fil l . The prototype tubes and a control (Plasma Separation Tube (PST) Gel Tube from Becton Dickinson and Company) were then fil led using an indirect blood draw procedure from 5 subjects and centrifuged at 1300 x g for 10 minutes. The tubes were then compared visual ly and for a range of cl inical chemistry parameters.

Results; Foam integrity was unaffected by evacuation and the separator in each prototype tube remained immobile upon centrifugation. All the prototype separators formed a barrier between the plasma and hematocrit. Some hemolysis was also observed within the L=i2mm Basotect© V 3012 foam separator and within the gel layer of the control tube. No apparent difference observed between the prototype foam separator tubes and the Gel "Control" tube for CA (calcium), MG (magnesium), K+ (potassium), CHOL (cholesterol), ALB (albumin) or Folate within each subject. A slightly high bias was observed for LDH (lactate dehydrogenase) from tubes with foam separator, however, the results were within expected normal range.

Example 3

Second Eval uation of Open Cel l Foam using Evacuated Tubes with Human Blood

Method; Prototype tubes were fabricated with two types of foam (Basotect® V 3012 (W) and Basotect® UF (G) Melamine based open cell foams available from BASF) cut into 10mm OD circular tubes, with a length L= 12mm, and inserted into Lithium-Heparin tubes of approximately 7.5mm ID and 13mm OD at a fixed position approximately 45 mm from the bottom of the tube (the expected position of the plasma/hematocrit interface). The prototype tubes were then evacuated to give a 4.5ml fill . The prototype tubes and a control (Plasma Separation Tube (PST) Gel Tube from Becton Dickinson and Company) were then filled using an indirect blood draw procedure from 5 subjects and centrifuged at 1300 x g for 10 minutes. The tubes were then compared visually and evaluated for a range of clinical chemistry parameters. In addition cel l counts were performed using the Beckman Coulter LH750 analyzer on each tube to evaluate separation performance. In addition, each tube was inverted 8 times 60-90 minutes after centrifugation and a second set of cell counts conducted to eval uate the barrier properties of each separator. Results; No apparent differences were observed between the foam separator tubes and Gel "Control" tubes for White Blood Cell (WBC), Red Blood Cell (RBC), Hemoglobin (HGB) and Platelet (PLT) counts within a given subject,

Figure 57 shows that mixing of separated plasma by inversion of the tube 8 times after centrifugation resulted in. elevated platelet counts when compared to the initial post centrifugation platelet count. However, a simi lar increase in platelet counts was observed in the foam separator tubes and the gel control tubes.

Within a subject, no apparent differences were observed between the Foam Separator tubes and the Gel Control tubes for GLUC (glucose), BUN (blood urea nitrogen), CREA (creatinine), K+ (potassium), CL (ch loride), Fe (iron), Ca (calcium), Mg (magnesium), Phos (phosphate), Choi (cholesterol), Trig (triglycerides), Uric (uric acid), TP (total protein), ALB (albumin), AST (aspartate aminotransferase), ALT (alanine aminotransferase), LD (lactate dehydrogenase), GGT (gamma glutamyl transferase), ALKP (alkaline phosphatase) or TBEL (total bilirubin).

Example 4

Effect of Centrifugation Parameters on Residual Ceil Counts

Method; Prototype tubes were fabricated with a separator made from Basotect® V 3012 (Melamine based open cell foam available from BASF) cut into 10mm OD circular tubes, with a length L- 12mm, and inserted into Lithium-Heparin tubes of approximately 7.5mm ID and 13mm OD at a fixed position approximately 45 mm from the bottom of the tube (the expected position of the plasma/hematocrit interface). The prototype tubes were then evacuated to give a 4.5ml fi ll . The prototype tubes for 2 of the subjects (8 and 9) to be tested were steri lized using irradiation. The prototype tubes and a control (Plasma Separation Tube (PST) Gel Tube from Becton Dickinson and Company) were then filled using an indirect blood draw procedure from 10 subjects. The prototype tubes were then centrifuged at 1500 x g, 3000 x g, or 4500 x g for 3, 5 or 10 minutes while the control gel tubes were centrifuged at 1500 x g for 10 minutes. Cel l counts using the standard procedures were then conducted to eval uate separation performance under each of the centrifuge conditions. In addition, each tube was inverted 10 times 60-90 minutes after centrifugation and a second set of cei l counts conducted to evaluate the barrier properties of each separator. Results; The integrity of ai l the foam separators were unaffected by evacuation. Upon centrifugation, all prototype tubes formed good barriers.

A summary of the cel l count resu lts is shown in Figure 58. Overall WBC, RBC and HGB counts did not show a significant difference across all the iterations of testing. However the results did show that at a given centrifugation speed, the longer the centrifugation time, the fewer number of contaminating Platelets present in the supernatant plasma for the foam separator tubes. However, centrifugation for 3 minutes for 1500 x g may not be enough to gi ve plasma of the requisite purity. The results suggest that not only the foam separator works as an effective barrier, but may also as a capture for the residual platelets in its infrastructure at the plasma- foam interface. Irradiation of the tubes (steri lization) does not appear to affect the performance of the foam separator tube.

Figure 59 shows a comparison of the platelet counts at the BD recommended centrifugation parameters ( 1500 x g for 10 minutes) for gel tube centrifugation with foam separator tubes. It can be seen that immediately fol lowing centrifugation, the PLT levels in the foam separator tubes are equivalent to the gel control tubes. It should be noted that subjects 5 through 10 were actual ly analyzed 24 hours after initial draw due to reagent availability. Following 10 inversions of the tubes, the PLT counts in the foam separator tubes increased slightly (maximum = 33) compared to the Gel Control Tube (min = 80, max = 190). These results show a better performance of the foam separator tubes compared to the gel control tubes.

This improvement in separation of the foam separator over the gel separator can be explained by location of platelets after centrifugation. In the foam separator tubes the platelets are trapped or captured within the pores of the separator after centrifugation, thus the 10 inversions of the tubes fai ls to cause the platelets to flow out of the separator. In the gel separator tube, platelets may be present on the upper surface of the gei barrier after centrifugation as a result of having a lower density than the gel or due to premature barrier formation of the gel (before complete separation), thus the 10 inversions of the gel tubes causes platelets to mix with the plasma, giving a lower separation performance.

Example 5

Recovery of Digoxin spiked into an Open Cel l Foam Separator Prototype tubes were fabricated with a separator made from Basotect© V 3012 (Melamine based open ceil foam avai lable from BASF) cut into 10mm OD circular tubes, with a length L= 12mm, and inserted into Lithium-Heparin tubes of approximateiy 7.5mm ID and 13mm OD at a fixed position approximately 45 mm from the bottom of the tube (the expected position of the plasma/hematocrit interface). Digoxin High Calibrator (0.5 mls) was spiked onto the foam. The prototype tubes were then evacuated to give a 4.5ml fill. The tubes were fi l led using an indirect draw procedure from 1 subject, mixed and centrifuged at 1500 x 'g' for 10 minutes. The Digoxin High Calibrator was assayed on the Elecsys neat, 1 : 10 dilution and 1 :20 di lution. The prototype tubes were also assayed for Digoxin on the Elecsys 2010 in triplicate 4 hours after the sample draw.

The results for the Digoxin High Calibrator are shown in Table 1 :

Table 1

<img file="WO2011069145A2_D0001.tif" /> The Calibrator average result is 4.49 ng/ml; therefore 2.24 ng of Digoxin was added to the foam separator. Since the blood sample volume was 4.5ml, then 2.24 ng of Digoxin was distributed into this volume, thus giving a concentration of 0.499 ng/ml Digoxin in the tube.

Table 2 shows the concentration of the Digoxin within the samples recovered from the prototype tubes.

Table 2

<img file="WO2011069145A2_D0002.tif" /> Therefore the overal l average recovery for both the samples (same subject) was 0.504 ng/ml . Thus, Digoxin was quantitatively recovered from the two prototype foam separator tubes.

Other embodiments of the invention wi l l be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein.