Plasma / cell concentrator device and method

Abstract

A plasma / cell concentration device and method for concentrating components from a fluid are described. Generally, a predetermined amount of fluid is forced from a first reservoir to a second reservoir through a fluid channel, and a predetermined amount of desiccant can be introduced for mixing with the fluid to produce a mixture. The mixture of fluid and desiccant can be passed between the first and second reservoirs until one or more components from the fluid are absorbed by the desiccant. After mixing, the desiccant can be prevented from migrating into the removal channel, and the fluid can be removed from a removal channel in fluid communication with the fluid channel.

Description

[Technical field]

[0001] The present invention relates to an apparatus and method for concentrating blood components, and more particularly to an apparatus and method for efficiently concentrating macromolecular plasma components and / or cells from plasma with or without leukocytes or from platelet-rich plasma without the need for centrifugation. [Background technology]

[0002] Blood can be fractionated and different fractions of blood can be used for different medical needs. For example, anemia (low amount of red blood cells) can be treated with transfusions of red blood cells. Thrombocytopenia (low platelet concentration) can be treated with transfusions of platelet concentrates.

[0003] The deposition of the various blood cells and plasma is based on the differences in the specific gravity of the cells and the viscosity of the medium. When the deposition reaches equilibrium, the heaviest ones will eventually deposit at the bottom, and the lightest ones will rise to the top. Under the influence of gravity and centrifugal forces, blood naturally deposits into three layers. At equilibrium, the top, low-density layer is a straw-colored, transparent fluid called plasma. Plasma is an aqueous solution that contains salts, metabolites, peptides, and many proteins, ranging from small (insulin) to very large (complement components). Plasma itself has limited medical use, but it is further fractionated to obtain proteins that are used, for example, in the treatment of hemophilia (factor VIII) or as a hemostatic agent (fibrinogen). Most of the plasma proteins and platelets in whole blood are in the plasma after gentle centrifugation, so the concentration of platelets in the plasma is increased and suspended in the supernatant plasma, and this component is called platelet rich plasma (PRP). The top layer after centrifugation typically contains only plasma proteins and is commonly referred to as Platelet-Poor Plasma (PPP) because platelets are absent or in low numbers as a result of the "hard spin".

[0004] The bottom dense layer is a deep red viscous fluid made up of nucleated red blood cells (RBCs) specialized for oxygen transport. The red color is due to the high concentration of chelated iron or heme, which causes the high specific gravity of red blood cells. Blood type-matched packed red blood cells are useful in treating anemia due to bleeding or other causes. The relative amount of red blood cells in whole blood is called the hematocrit, and in normal humans it ranges from about 38% to about 54%.

[0005] The middle layer is the smallest layer and appears as a thin white band above the red blood cell layer and below the plasma, and is called the buffy coat. The buffy coat itself has two main components: white blood cells, which have a nucleus, and non-nucleated bodies called platelets. White blood cells confer immunity and also contribute to debris collection. Platelets seal blood vessel breaks to stop bleeding and deliver growth and wound healing factors to wound sites. The buffy coat can be separated from whole blood by "hard spinning" the blood. In a "hard spin", whole blood is spun hard and long enough to deposit platelets from the plasma onto the red blood cell pack and to allow white blood cells to permeate through the red blood cell pack to the red blood cell / plasma interface.

[0006] In order to separate a given volume of blood into its constituent components, it is necessary to separate and concentrate the components, generally using a centrifuge device. Other methods for concentrating blood components, particularly plasma macromolecular components, and / or platelets, and / or white blood cells, include devices that employ scavenging hollow fiber filtration to remove water and low molecular weight solutes from plasma or PRP. Other devices use dry size-exclusion chromatography beads to adsorb water and low molecular weight solutes in the centrifuge tube. These devices include a paddle or rake to agitate the plasma / bead mixture during concentration, and then the beads are centrifuged into the lower or distal compartment of the centrifuge tube (or drum) through a frit (e.g., a filter with well-known porosity made of sintered particles of glass, ceramic, etc.) that retains the swollen beads while allowing the concentrated product to pass through.

[0007] Yet another alternative is a device that uses beads contained within a syringe in combination with an agitator such as one or more metal BBs to facilitate mixing of the beads with the plasma, followed by squeezing the concentrated product through a bead-retaining frit or centrifugation through a bead-retaining frit.

[0008] In all of these methods, some means of sweeping or agitation is usually required to prevent the formation of a gel polarizing film on the surface of the beads or fibers. In the case of hollow fibers, this is accomplished by circulating plasma through the fibers during filtration to clean the membrane surface; in the case of centrifuges, agitation is accomplished with a paddle or rake; in the case of bead-loaded syringes, agitation is accomplished by shaking the syringe so that the BB stirs the bead / plasma suspension.

[0009] Hollow fiber membrane cartridges are generally expensive, and methods using hollow fiber membrane cartridges require circulation pumps and tubing circuits.When using chromatography beads, the beads need to be mixed quickly after contact with plasma to prevent gel polarization, which can deposit large molecules on the bead surface and clog the pores of the beads, and to prevent beads from clumping due to accumulation of high concentrations of proteins on the bead surface. Summary of the Invention [Problem to be solved by the invention]

[0010] Thus, a need remains for a simple and effective method and device for effectively concentrating macromolecular plasma components and / or cells from plasma or PRP, with or without leukocytes, without the need for centrifugation. [Means for solving the problem]

[0011] The present invention relates to an apparatus and method for efficiently concentrating macromolecular plasma components and / or cells from plasma or PRP with or without leukocytes without the need for centrifugation. One embodiment of the apparatus can utilize a first syringe fluidly coupled to a second syringe via a manifold. The manifold also includes receiving channels for temporarily or permanently securing the tips of each syringe. Each syringe can be directly integrated with the manifold or can optionally include non-standard connections to ensure that one or both of the syringes cannot be inadvertently attached or detached from an inappropriate port.

[0012] The fluid channel may extend through the manifold and fluidly couple the opening of the first syringe to the opening of the second syringe, and the removal channel may extend through the manifold and fluidly couple to the fluid channel. To prevent the release of fluid from the fluid channel, the removal channel may incorporate an actuatable valve, configurable between an open and a closed configuration, for example, automatically or manually via a stopcock. A filter or frit may also be incorporated into the manifold at the inlet of the removal channel to filter select components passing from the fluid channel to the removal channel.

[0013] The fluidic channels can optionally incorporate a number of protrusions or elements, such as vanes, spikes, crosshairs, etc., to promote effective mixing of the fluids as they pass through the fluidic channels. Additionally, a predetermined amount of dry beads or elements can also be added directly into the fluidic channels or into one or both of the syringes.

[0014] A predetermined amount of plasma or PRP to be concentrated may be introduced into the first or second syringe (or both), and the syringes may be coupled to corresponding ends in the manifold such that their openings are located at each end of the fluidic channel. With a predetermined amount of dry beads held in the fluidic channel and / or held in one or both of the syringes, the respective plungers may be alternately actuated to push the plasma or PRP back and forth between the syringes through the fluidic channel. To prevent leakage of the plasma or PRP from the fluidic channel as it is circulated between the syringes, the withdrawal channel may be held closed. As the plasma or PRP is circulated between the syringes through the fluidic channel, the predetermined amount of beads may mix thoroughly with the plasma or PRP and absorb various components in the plasma or PRP, such as water, electrolytes, and small proteins, so that the plasma or PRP is concentrated, leaving a concentrate of platelet-rich plasma. As the mixture of beads and plasma or PRP circulates back and forth between the syringes, the syringe connections to the manifold may be integral to ensure a fluid-tight connection, and the inner diameter of the syringe coupling may taper internally at the proximal end of the coupling to reduce the possibility of clogging from bead agglomerates that may form due to gel polarization.

[0015] Once the plasma or PRP is sufficiently concentrated, a valve along the removal channel may be opened to allow removal of the concentrated product passing through the fluidic channel. A filter or frit may prevent beads and other components from entering the removal channel, so that only the concentrated product passes through the removal channel and into a reservoir, such as a collection syringe. As the mixture of beads and plasma or PRP continues to circulate through the fluidic channel, the removal channel may be opened to allow removal of the concentrated product. This ensures that the concentrated product is collected uniformly from the mixture. Additionally, sweeping the bead surface during product extraction may further ensure that residual water absorption does not cause gel polarization and loss of macromolecular components, or clumping, if the beads are not yet fully saturated.

[0016] Alternatively, both plungers on each syringe may be depressed simultaneously to generate pressure to force the concentrated product out of both syringes and through an outlet channel for collection with the valve open. In either case, a filter or frit may prevent beads from migrating into the outlet channel, allowing only the concentrated product to pass through.

[0017] Alternatively, additional outlet channels and ports may be provided to allow material to be introduced without passing through a filter or frit. As material is introduced from the manifold, it may mix with the beads in the manifold and the beads may be swept into one or both of the syringes with their respective plungers partially extended.

[0018] These beads can be made of any number of materials configured to act as concentrators when in contact with and mixed with plasma or PRP, selectively absorbing certain components such as water, electrolytes, small proteins, etc., leaving a concentrated product for collection. Examples of various materials from which beads can be formed can include various polymers, metals, minerals, polysaccharides, silica gels, ceramics, glass, etc., any material suitable for processing biomaterials.

[0019] In another aspect, the manifold includes a manifold body including ports disposed at opposite ends of the manifold body, each of which can be fluidly connected to one another via a fluid channel extending between the ports through the manifold body. The removal channel can extend from the fluid channel through a filter or frit positioned to prevent or inhibit beads or other components from passing through the removal channel.

[0020] Any number of configurations are possible, such as two in-line syringes with a spring-like biasing element attached to one of them, with mixing achieved by alternately applying and releasing pressure to only one of the syringes. The two syringes do not need to be parallel, but may be connected to one another via adjacent connecting manifolds at any angle. A second, improved syringe may be coupled to an opposing port of the manifold body rather than having a plunger, and the second syringe may include a biasing element therein anchored between the moveable plunger and an anchor platform that may be rigidly positioned relative to the body of the syringe.

[0021] In yet another example, a volume of plasma or PRP may be concentrated by connecting a first syringe to a second syringe through a manifold containing a filter or frit, the first syringe optionally being preloaded with a volume of beads, and the second syringe containing the volume of plasma or PRP to be concentrated. Alternatively, in another embodiment, the volume of beads may be preloaded within the manifold itself. The manifold or second syringe may include a valve, such as a stopcock, that is initially closed to prevent communication between the first and second syringes when initially coupled to the manifold.

[0022] With the plunger of the first syringe initially fully depressed, the plunger may be drawn proximally with the valve closed, creating a vacuum force within the first plunger. This may also allow a predetermined amount of entrapped beads to disperse within the syringe prior to contact with the fluid contained within the second syringe. The first plunger may be locked in its retracted position by a locking mechanism, and the valve may be opened to allow the vacuum within the first syringe to draw fluid from the second syringe through the manifold. This sudden diffusion of fluid into the first syringe helps minimize the possibility of beads clumping within the first syringe upon initial fluid contact, and may also facilitate rapid introduction of fluid after the valve is opened. Once fluid has been introduced into the first syringe, the locking mechanism may be released (e.g., by twisting the plunger rod relative to the syringe body) to allow circulation of the slurry between the two syringes.

[0023] An alternative manifold body may have a first port and a second port disposed along a first surface of the manifold such that both are adjacent to each other and fluidly connected via a curved or angled fluid channel. An optional chamber may be incorporated between the fluid channels to hold a quantity of beads during the initial mixing stage between the syringes. In addition, a filter or frit well may be further configured to extend from the chamber to hold the frit within the chamber and also to allow a removal syringe to be coupled to the manifold body. Since the ports are disposed along a common plane of the manifold body, the syringes may be at any number of angles relative to each other and to the manifold body. Additionally, this embodiment may be utilized with any of the devices and methods described herein.

[0024] In yet another embodiment for recovering a predetermined amount of concentrated product, rather than removing the concentrated product directly from the manifold, a combination can be used in combination with a centrifuge. The combination can have a housing with an inlet port located at an upper portion of the housing through which the mixed slurry is introduced. The outlet port can be located at a lower portion of the housing, and a frit can be located within the housing to separate the interior volume into an upper portion and a lower portion.

[0025] Once the mixed slurry is introduced through the ports into the top of the housing, both ports may be capped or closed and the entire joint subjected to centrifugation to drive the fluid from the top, through the frit and into the bottom of the housing. Once all of the concentrated product has been separated into the bottom and the beads remaining in the top above the frit, the removal port may be opened to remove the concentrated product from the housing.

[0026] In one embodiment of a method for concentrating a component from a fluid, the method generally includes forcing a volume of fluid from a first reservoir through a fluidic channel to a second reservoir and introducing a volume of a desiccant to mix with the fluid. The mixture of fluid and desiccant can be passed between the first and second reservoirs until one or more components from the fluid are absorbed by the desiccant. After mixing, the fluid can be removed through an output channel in fluid communication with the fluidic channel while preventing the desiccant from migrating into the output channel.

[0027] In one embodiment of a device for concentrating a component from a fluid, the device may generally include a connector having a first port and a second port connected via a fluidic channel, a first reservoir having a variable pressure attachable to the first port, and a second reservoir having a variable pressure attachable to the second port. The removal channel may be in fluid communication with the fluidic channel, and the filter may be positioned to prevent passage of the desiccant from the fluidic channel to the removal channel. [Brief description of the drawings]

[0028] [Figure 1] FIG. 1 is a perspective view of a device that can be used to effectively concentrate plasma components and / or cells without the need for centrifugation. [Figure 2A] FIG. 2A is a perspective view of another embodiment of a manifold that can be used in conjunction with a conventional or modified syringe. [Figure 2B] FIG. 2B is a side view of another embodiment of a manifold that can be used in conjunction with a conventional or modified syringe. [Figure 2C] FIG. 2C is a bottom view of another embodiment of a manifold that can be used in conjunction with a conventional or modified syringe. [Figure 2D]FIG. 2D is a top view of another embodiment of a manifold that can be used in conjunction with a conventional or modified syringe. [Figure 3A] 3A-3E are perspective views showing one example for concentrating plasma or PRP. [Figure 3B] 3A-3E are perspective views showing one example for concentrating plasma or PRP. [Figure 3C] 3A-3E are perspective views showing one example for concentrating plasma or PRP. [Figure 3D] 3A-3E are perspective views showing one example for concentrating plasma or PRP. [Figure 3E] 3A-3E are perspective views showing one example for concentrating plasma or PRP. [Figure 4A] 4A-4D are perspective views showing another embodiment for concentrating a volume of buffy coat. [Figure 4B] 4A-4D are perspective views showing another embodiment for concentrating a volume of buffy coat. [Figure 4C] 4A-4D are perspective views showing another embodiment for concentrating a volume of buffy coat. [Figure 4D] 4A-4D are perspective views showing another embodiment for concentrating a volume of buffy coat. [Diagram 5] FIG. 5 is a perspective view of an alternative embodiment of a single syringe pressure application and release device. [Figure 6] FIG. 6 is a perspective view of another example device that can utilize the vacuum force generated in one of the syringes to rapidly mix the beads with the plasma or PRP. [Figure 7A] FIG. 7A is a partial cross-sectional side view showing a manifold that can fluidly couple two syringes to one another at various angles. [Figure 7B] FIG. 7B is a partial cross-sectional perspective view showing a manifold that can fluidly couple two syringes to one another at various angles. [Figure 8]FIG. 8 is a perspective view showing a coupling that can be used in combination with a centrifuge. [Figure 9A] 9A-9C show an example of a method in which a volume of plasma or PRP is mixed with a desiccant and centrifuged to obtain a concentrated product. [Figure 9B] 9A-9C show an example of a method in which a volume of plasma or PRP is mixed with a desiccant and centrifuged to obtain a concentrated product. [Figure 9C] 9A-9C show an example of a method in which a volume of plasma or PRP is mixed with a desiccant and centrifuged to obtain a concentrated product. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0029] A perspective view of an apparatus according to one embodiment is shown in FIG. 1, which illustrates an assembly 10 that can be used to effectively concentrate one or more plasma components and / or cells from plasma or PRP without the need for centrifugation. The assembly 10 may concentrate components and / or cells that may or may not include white blood cells. As shown, a first reservoir or syringe 22 is fluidly coupled to a second reservoir or syringe 26 via a manifold 12 that also includes a receiving channel for temporarily or permanently securing the tip of the corresponding syringe 22, 26 to the manifold 12. Although syringes are shown and described, alternative fluid container or reservoir structures may be used instead. Each syringe 22, 26 may be directly integrated with the manifold 12 or may be attached to an optional non-standard connection to prevent inadvertent attachment and detachment of one or both of the syringes 22, 26 to the wrong port.

[0030] A fluid channel 16 may extend through the manifold 12 to fluidly couple the openings of a first syringe 22 and a second syringe 26, and an output channel 18 may extend through the manifold 12 to fluidly couple to the fluid channel 16, which may have a diameter of, for example, 5 mm, but may range from, for example, 3 to 10 mm. A openable / closable valve 20, which may be configured, for example, automatically or manually via a stopcock, may be integrally incorporated with the output channel 18 to prevent the release of fluid from the fluid channel 16. A filter or frit 14 may also be incorporated into the manifold 12 at the inlet of the output channel 18 to filter selected components passing from the fluid channel 16 to the output channel 18.

[0031] The fluidic channels 16 can optionally incorporate a number of protrusions or elements, such as vanes, spikes, crosshairs, etc., to promote effective mixing of the fluids as they pass through the fluidic channels 16. Additionally, a quantity of dry beads or elements can also be added directly into the fluidic channels 16 or into one or both of the syringes 22, 26. With the quantity of dry beads or elements contained directly into the fluidic channels 16 or into one or both of the syringes, the device can optionally be moved, such as tapped or shaken, while held in a horizontal configuration to promote dispersion of the quantity of dry beads on the bottom of the connecting channels 16 or within the syringes 22, 26 prior to introducing the material to be concentrated. This can reduce the likelihood of clumping immediately after introducing the plasma or PRP into contact with the beads.

[0032] A predetermined amount of plasma or PRP to be concentrated may be introduced into the first syringe 22 or the second syringe 26 (or both), which may be coupled to corresponding ends in the manifold 12 such that their openings are located at each end of the fluidic channel 16. With a predetermined amount of dry beads held within the fluidic channel 16 and / or held within one or both of the syringes 22, 26, the respective plungers 24, 28 may be alternately actuated to push the plasma or PRP back and forth between the syringes 22, 26 through the fluidic channel 16. As the plasma or PRP is circulated between the syringes 22, 26, the withdrawal channel 18 may be held closed to prevent leakage from the fluidic channel 16. As the plasma or PRP circulates between syringe 22 and syringe 26 via fluid channel 16, a quantity of beads may thoroughly mix with the plasma or PRP and absorb various components in the plasma or PRP, such as water, electrolytes, and small proteins, thereby concentrating the plasma or PRP and leaving a concentrate of platelet rich plasma. As the mixture of beads and plasma or PRP circulates back and forth between syringe 22 and syringe 26, the syringe connection to manifold 12 may be integral to ensure a fluid tight connection, and the inner diameter of the syringe connection may taper internally at the proximal end of the connection to reduce the possibility of clogging due to bead agglomerates that may form due to gel polarization, for example, a diameter (2-6 mm) that is relatively smaller than the syringe inner diameter but relatively larger than a standard syringe inner diameter.

[0033] Once the plasma or PRP is sufficiently concentrated, a valve 20 along the removal channel 18 may be opened to allow removal of the concentrated product passing through the fluidic channel 16. The filter or frit 12 may prevent beads and other components from entering the removal channel 18, so that only the concentrated product passes through the removal channel 18 and into a reservoir such as a collection syringe. As the mixture of beads and plasma or PRP continues to circulate through the fluidic channel 16, the removal channel 18 may be opened to allow removal of the concentrated product. This ensures that the concentrated product is collected uniformly from the mixture. Additionally, sweeping the bead surfaces during product extraction may further ensure that residual water uptake does not cause gel polarization and loss of macromolecular components, or clumping, if the beads are not yet fully saturated.

[0034] Alternatively, pressure may be generated by simultaneously depressing the corresponding plungers 24, 28 of both syringes 22, 26 to force the concentrated product out of both syringes 22, 26 and through the withdrawal channel 18 for collection with the valve 20 open. In either case, a filter or frit 14 may prevent beads from migrating into the withdrawal channel 18, allowing only the concentrated product to pass through.

[0035] Alternatively, additional withdrawal channels and ports may be provided to allow material to be introduced without passing through the filter or frit 14. As material is introduced through the manifold 12, it may mix with the beads contained therein and be swept into one or both of the syringes 22, 26 to force the corresponding plungers 24, 28 to partially expand.

[0036] These beads can be made of any number of materials configured to act as concentrators when in contact with and mixed with plasma or PRP, selectively absorbing certain components such as water, electrolytes, small proteins, etc., leaving a concentrated product for collection. Examples of various materials from which beads can be formed can include various polymers, metals, minerals, polysaccharides, silica gels, ceramics, glass, etc., any material suitable for processing biomaterials.

[0037] Examples of polymers include, for example, dextranomer, polystyrene, polyethylene, polyvinyl chloride, polypropylene, polyacrylamide, etc. Examples of metals include, for example, titanium, etc. Examples of minerals include, for example, zeolite, corundum, quartz, etc. Examples of polysaccharides include, for example, alginate gel, starch, dextran, agarose, etc.

[0038] In some variations, the beads may be conjugated to an activator, such as an antibody, such as immunoglobulin g.

[0039] Since the beads are used to induce changes in biological material to concentrate plasma or PRP, the size and composition of the beads can vary depending on the material of the beads used and the biological material to be concentrated. In one example, the beads can be made of polyacrylamide material (e.g., Bio-Rad P6 chromatography beads with a nominal molecular weight cutoff (NMWL) of 1000 to 6000), and each can be sized to have a dry bead diameter of, for example, 45 to 90 micrometers. Thus, for a given amount of plasma or PRP to be concentrated, such as about 10 ml, the volume can be anywhere from, for example, 5 to 30 ml, the weight of the beads used can be anywhere from, for example, about 200 to 5,000 mg, but can be about 1,000 mg, and the ratio of beads to plasma or PRP can be anywhere from, for example, 0.04 to 0.15, but can be about 0.1, to effectively separate the desired components.

[0040] Additionally, the number of cycles between syringe 22 and syringe 26 can vary depending on the beads and the biological material to be concentrated. In one embodiment, the plasma or PRP can be circulated for, for example, 10 to 50 cycles to thoroughly mix the beads with the plasma or PRP. Alternatively, the circulation can be timed to allow sufficient mixing, for example, ranging from 30 seconds to 2 minutes or more. Additionally, because the diameter of fluid channel 16 is relatively small compared to the size of the beads, clumps formed due to incomplete mixing can be broken apart by turbulence and shear as the clumps pass through fluid channel 16. Additionally, circulating back and forth between syringe 22 and syringe 26 can promote efficient mixing of the beads with the plasma or PRP, producing a more uniform slurry with less foaming.

[0041] 2A-2D are perspective, side, bottom, and top views, respectively, of a manifold according to another embodiment that may be used in combination with a conventional or modified syringe. Manifold 30 includes a manifold body 32 including ports 34, 36 disposed at opposite ends of manifold body 32, each of which may be fluidly connected to one another via a fluid channel 38 extending through manifold body 32 between ports 34 and 36. Extraction channel 40 may extend from fluid channel 38 through a filter or frit 42 positioned to prevent or inhibit beads or other components from passing through and entering extraction channel 40.

[0042] Although the outtake channel 40 is shown extending transversely to the fluid channel 38, the outtake channel 40 may alternatively be inclined at a non-standard angle relative to the fluid channel 38. Additionally, while the assembly 30 can be used in combination with conventional syringes, in other embodiments the ports 34, 36 can be adjusted or modified to temporarily or permanently engage syringes of particular configurations.

[0043] Example 1

[0044] In one example of a test that was performed, as shown in the perspective view of Figure 3A, a first volume 50A of citrated bovine PRP was introduced directly into a first syringe 22 coupled to port 36, and the outlet of the withdrawal channel 40 was sealed by a cap. A second syringe 26 with its plunger 28 fully depressed was coupled to port 34, and a predetermined amount of enrichment beads 52 was preloaded into the fluidic channel 38 contained within the manifold body 32 prior to attachment of the first syringe 22 loaded with PRP.

[0045] Depression of first plunger 24 forces a first volume 50A of PRP into fluid channel 38 and into contact with beads 52 contained therein. As more first volume 50A is introduced, the PRP and beads begin to mix by being forced into the opposing second syringe 26, as shown in the perspective view of FIG. 3B, and second plunger 28 can be forced to expand to entrain a second volume 50B.

[0046] The gentle shuttle of the PRP between the syringes 22 and 26 through the manifold body 32 may be maintained for a period of time, for example, about 30 seconds, to allow the beads 52 to fully absorb water from the plasma. Once mixing is complete, the concentrated mixture may appear as a uniform slurry with no visible clumps of beads. As shown in the perspective view of FIG. 3C, the cap (and valve, if any) may be removed and a collection syringe 54 may be attached to the withdrawal port 40 with its plunger fully depressed. This circulation may continue between the first syringe 22 and the second syringe 26 while the concentrated product 56 is forced through a filter or frit 42 that prevents the passage of beads by moderate pressure generated by the flow restriction and resistance of the syringes 22, 26. This pressure may force the concentrated PRP through the bead-retaining filter or frit 42 and into the collection syringe 54, as shown in the perspective view of FIG. 3D.

[0047] When most of the concentrated fluid 56 had been squeezed through the filter or frit 42 and into the collection syringe 54, the remaining bead pack 52 appeared uniform and white, as shown in the perspective view of FIG. 3E, indicating that the mixing and collection maintained the uniformity of the slurry and that the concentrated fluid 56 had been nearly completely transferred to the collection syringe 54.

[0048] Example 2

[0049] In another example of a test that was performed, a predetermined amount of buffy coat 60A consisting of plasma, platelets, white blood cells, and a small amount of red blood cells was introduced into a first syringe 22 coupled to an opposite port 36 and a second syringe 26 coupled to a port 34 with its plunger 28 fully depressed, as shown in the perspective view of FIG. 4A.

[0050] A predetermined amount of beads 52 was preloaded into the first syringe 22 along with a predetermined amount of buffy coat 60A, although beads 52 may alternatively be preloaded into the fluid channel 38 of manifold body 32. As shown in the perspective view of FIG. 4B, the predetermined amount of buffy coat could then be circulated between syringes 22, 26 via fluid channel 38 to allow for thorough mixing of beads 52 between the predetermined amounts of volumes 60A, 60B. To prevent agglomeration of beads 52 (e.g., upon initial contact with fluid), volumes can be rapidly circulated between syringes 22 and 26, especially when beads 52 are preloaded directly into one of the syringes.

[0051] After a period of circulating the mixed slurry back and forth between syringe 22 and syringe 26, for example about 30 seconds, collection syringe 62 was attached to outlet port 40, and concentrated product 64 was expressed through bead retaining filter or frit 42 and into collection syringe 62 by continuing to circulate the slurry back and forth between in-line syringe 22 and in-line syringe 26, as shown in the perspective view of FIG. 4C.

[0052] After the concentrated product 64 has been collected, the remaining bead packs 52 may appear uniformly and slightly pink from traces of red blood cells in the small amount of residual interstitial fluid between the packed beads 52. Figure 4D is a perspective view showing how the concentrated product 64 may be collected while the beads 52 are retained within the first syringe 22 and / or second syringe 26.

[0053] As shown in both examples above, in the concentration of PRP and buffy coat, the ratio of the recovered amount to the initial input amount was consistent with the amount of water that would be absorbed by the amount of beads 52 employed, indicating that cells and macromolecules were effectively concentrated. In another preliminary test, cell counting was performed and a relatively high cell recovery rate was confirmed.

[0054] Form by example

[0055] Any number of configurations are possible, such as two in-line syringes with a spring-like biasing element attached to one of them, with mixing achieved by alternately applying and releasing pressure to only one of the syringes. The two syringes do not need to be parallel, but may be connected to one another via adjacent connecting manifolds at any angle. The perspective view of FIG. 5 shows syringe 22 coupled to a manifold body 70 that includes a filter or frit 72. A second modified syringe 78 may be coupled to an opposing port of manifold body 70 rather than having a plunger, and the second syringe 78 may include a biasing element 80 therein anchored between a movable plunger and an anchor platform 82 that may be rigidly positioned relative to the body of the syringe 78.

[0056] The biasing element 80 may maintain the plunger in a compressed state such that the plunger is biased to push away from the anchor platform 80. In this configuration, the first syringe 22 may contain beads 52 as shown, or the manifold body 70 may contain beads 52. In either case, plasma or PRP may be introduced into the first syringe 22 and the plunger 24 may be depressed to force the mixture of beads 52 and plasma or PRP through the fluid channels of the manifold body 70 and into the second syringe 78. The plunger 24 of the first syringe 22 may be released so that the biasing element 80 automatically presses the plunger in the second syringe 78 to force the mixture back through the manifold body 70 and into the first syringe 22. This process may be repeated until the fluid is sufficiently concentrated, as described herein. The stopcock 76 or valve (if provided) may be opened to allow the concentrated product to pass through the filter or frit 72 , through the withdrawal channel 74 and into an attached collection syringe 86 .

[0057] As shown in the perspective view of FIG. 6, in yet another embodiment, a volume of plasma or PRP may be concentrated by connecting a first syringe 22 to a second syringe 88 through a manifold 70 containing a filter or frit 72, as described above. The first syringe 22 may optionally be preloaded with a volume of beads 52, and the second syringe 88 may contain a volume of plasma or PRP 89 to be concentrated. In another embodiment, the volume of beads 52 may instead be preloaded within the manifold 70 itself. The manifold 70 or second syringe 88 may include a valve 91, such as a stopcock, that is initially closed to prevent communication between the first syringe 22 and the second syringe 88 when initially coupled to the manifold 70.

[0058] With the plunger of the first syringe 22 initially fully depressed, the plunger may be pulled proximally with the valve 91 closed such that a vacuum force is created within the first plunger 22. This may also disperse the entrapped amount of beads 52 within the syringe 22 prior to contact with the fluid contained within the second syringe 88. The first plunger may be locked in its retracted position with the locking mechanism 93, and the valve 91 may be opened such that the vacuum within the first syringe 22 draws fluid 89 from the second syringe 88 through the manifold 70. This sudden diffusion of fluid 89 into the first syringe 22 helps minimize the possibility of beads 52 clumping within the first syringe 22 upon initial fluid contact and may also facilitate rapid introduction of fluid 89 after the valve 91 is opened.

[0059] Once fluid 89 is introduced into the first syringe 22, the locking mechanism 93 is released (e.g., by twisting the plunger rod relative to the syringe body), allowing circulation of the slurry between the two syringes 22, 88. As described above, a collection syringe can then be coupled to the manifold 70 for collection. Similarly, the syringes 22, 88 can be attached via a straight connector or a connector that allows the two syringes to be adjacent to each other at any angle.

[0060] 7A and 7B are partial cross-sectional side and perspective views illustrating a manifold 90 that can fluidly couple two syringes to one another at various angles. As shown, a manifold body 92 may have a first port 96 and a second port 98 disposed along a first surface of the manifold 92 such that both are adjacent to one another and fluidly connected via curved or angled fluid channels 100, 102. An optional chamber 94 may be incorporated between the fluid channels 100 and 102 to hold a predetermined amount of beads 52 during the initial mixing stage between the syringes. Additionally, a filter or frit well 104 may be further configured to extend from the chamber 94 to hold a frit within the chamber 94 and also to allow for coupling of a removal syringe to the manifold body 92.

[0061] Because the ports 96, 98 are disposed along a common surface of the manifold body 92, the syringes can be at any number of angles relative to each other and to the manifold body 92. Additionally, this embodiment can be utilized with any of the devices and methods described herein.

[0062] Additionally, in any of these embodiments described herein, the product collection syringe (or other collection vessel, such as a centrifuge tube integral with the frit on the distal side of the receiving space) may be centrifuged to centrifuge the concentrate through the frit and separate it from the bead pack. This may have the advantage of forcing any interstitial fluid between the beads out of the bead pack, improving volumetric recovery, but at the expense of an extra transfer step and centrifugation.

[0063] As a further alternative to applying pressure to the slurry to recover the concentrated product, for example, a stream of air can be used to blow the concentrated product out of the bead pack through a frit. This requires a pressurized air source to achieve good volumetric recovery. Additionally, foaming can occur if not done carefully, potentially damaging cells and plasma proteins.

[0064] With regard to volume recovery, it should be noted that when the beads are densely packed, approximately 1 / 3 of the total pack volume is interstitial space. This volume may not be recovered if the concentrate is pressed out. Regardless of the method used to recover the concentrate (pressure or centrifugation), the magnification may be limited to approximately 3x due to the difficulty of effectively mixing slurries with a volume ratio of (fully swollen) beads to residual interstitial fluid greater than 2. Reasonable volumetric recoveries of product harvested by pressure may be, for example, on the order of 1.5x to 2x.

[0065] In yet another embodiment for recovering a predetermined amount of concentrated product, rather than removing the concentrated product directly from the manifold, a coupling 110 as shown in Figure 8 can be used in combination with a centrifuge. The coupling 110 can have a housing 112 with an inlet port 114, such as a Luer connector, disposed at an upper portion of the housing 112 through which the mixed slurry is introduced. An outlet port 116, such as a Luer connector, can be disposed at a lower portion of the housing 112, and a frit 118 can be disposed within the housing 112 to separate the interior volume into an upper portion 120 and a lower portion 122.

[0066] Once the mixed slurry has been introduced into the upper portion 120 of the housing 112 through port 114, both ports 114, 116 may be capped or closed and the entire joint 110 may be subjected to centrifugation to drive the fluid from the upper portion 120, through the frit 118 and into the lower portion 122 of the housing 112. Once all of the concentrated product has been separated into the lower portion 122 and the beads remaining in the upper portion 120 above the frit 118, the removal port may be opened to remove the concentrated product from the housing 112.

[0067] In one use of the housing 120, the manifold 12 may be coupled to each syringe 22, 26 to initially mix a predetermined amount of plasma or PRP with a predetermined amount of dry beads that may freely intermix with the amount of plasma or PRP as they pass between the syringes 22, 26, as described above and shown in FIG. 9A. The resulting slurry or mixture 130 of plasma or PRP and dry beads may be disconnected from the manifold 12 and then transferred to one of the syringes 26, which may then be fluidly coupled to the inlet port 114. As shown in FIG. 9B, the slurry or mixture 130 from the syringe 26 may then be transferred through the inlet port 114 and into the upper portion 120 of the housing 112.

[0068] Once the slurry or mixture 130 has been sufficiently displaced, the syringe 26 may be disconnected from the input port and the housing 112 may be centrifuged to drive the slurry or mixture 130 from the upper portion 120, through the frit 118 and into the lower portion 122 of the housing 112. The frit 118 may retain the dry beads that have absorbed one or more components from the plasma or PRP, thereby retaining the resulting concentrated product 134 within the lower portion 122. The additional syringe 132 may be fluidly coupled to a removal port 116 for removing the concentrated product 134 from the lower portion 122 into the additional syringe 132, as shown in FIG. 9C.

[0069] The above disclosed apparatus and methods are not limited to the individual embodiments shown or described, but may include combinations incorporating individual features among different variations. Modifications of the above-described assemblies and methods for implementing the invention, combinations between different aspects that can be implemented, and variations of aspects of the invention that are obvious to those skilled in the art are intended to be within the scope of the claims.

Claims

1. a connector in fluid communication with the fluid channel; a first reservoir having a variable pressure; a second reservoir having a variable pressure, the first reservoir being fluidly coupled to the second reservoir via the connector while being maintained at an angle relative to the second reservoir, the connector being sized to allow flow of desiccant between the first reservoir and the second reservoir; a biasing element disposed in-line with the second reservoir and configured to exert pressure against a plunger within the second reservoir; an outflow channel in fluid communication with the fluid channel; a filter positioned within the connector to prevent passage of the desiccant from the fluid channel to the removal channel; An apparatus for concentrating a component from plasma or PRP comprising:

2. The apparatus of claim 1 , wherein the connector comprises a manifold body.

3. The device of claim 1 , wherein the first reservoir and the second reservoir each comprise a syringe having a plunger.

4. The apparatus of claim 1 , further comprising a valve in communication with the removal channel.

5. The device of claim 1 , wherein the filter comprises a frit.

6. The device described in claim 1, wherein the desiccant comprises a predetermined amount of desiccant beads for introduction into the first reservoir or the second reservoir.

7. The device of claim 1, wherein the biasing element comprises a spring positioned to apply pressure against the plunger in the second reservoir.

8. The device of claim 1 , wherein the first reservoir further comprises a locking mechanism configured to maintain a position of a plunger within the first reservoir to generate a vacuum force.

9. pumping a volume of fluid comprising plasma or PRP from a first reservoir through a fluid channel to a second reservoir; passing the fluid and a quantity of the desiccant between the first reservoir and the second reservoir until one or more components from the fluid are absorbed by the desiccant; preventing migration of the desiccant into an extraction channel in fluid communication with the fluid channel and removing the fluid from the extraction channel; A method for concentrating a component from plasma or PRP comprising:

10. 10. The method of claim 9, wherein the one or more components absorbed by the desiccant from the volume of fluid comprises water, electrolytes, and / or proteins.

11. 10. The method of claim 9, wherein pumping the amount of fluid comprises transferring the amount of fluid from a first syringe to a second syringe through the fluid channel formed through a manifold.

12. The method of claim 9 , wherein passing the fluid and the amount of desiccant comprises introducing desiccant beads.

13. 10. The method of claim 9, wherein passing the fluid and the amount of desiccant comprises passing the fluid for between 30 seconds and 2 minutes.

14. The method of claim 9 , wherein removing the fluid comprises opening a valve such that the removal channel is in fluid communication with the fluid channel.

15. 15. The method of claim 14, wherein preventing migration of the desiccant comprises passing fluid through a filter or frit to inhibit the desiccant from entering the extraction channel.

16. The method of claim 9 , further comprising collecting the fluid from the removal channel via a collection syringe.

17. 10. The method of claim 9, wherein removing the fluid comprises removing the fluid from the removal channel while the fluid and the desiccant are passing between the first reservoir and the second reservoir.

18. The method of claim 9 , wherein removing the fluid comprises removing from the removal channel by inducing pressure in each of the first and second reservoirs.

19. 10. The method of claim 9, wherein the second reservoir includes a biasing member contained therein that exerts a biasing force against a plunger within the second reservoir.

20. 10. The method of claim 9, wherein pumping the volume of fluid comprises inducing a vacuum in the second reservoir to draw the volume of fluid into the first reservoir.

21. 10. The method of claim 9, wherein the step of pumping the quantity of fluid further comprises the step of retaining the desiccant in a manifold disposed between the first reservoir and the second reservoir prior to transferring the fluid and the quantity of desiccant between the first reservoir and the second reservoir.

22. 10. The method of claim 9, wherein pumping the quantity of fluid further comprises retaining the desiccant in the first reservoir or the second reservoir prior to transferring the fluid and the quantity of desiccant between the first reservoir and the second reservoir.

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