WO2011119535A1

WO2011119535A1 - Filtration systems, methods, and devices to derive component fractions from whole blood

Info

Publication number: WO2011119535A1
Application number: PCT/US2011/029324
Authority: WO
Inventors: Edward F. Leonard
Original assignee: The Trustees Of Columbia University In The City Of New York
Priority date: 2010-03-22
Filing date: 2011-03-22
Publication date: 2011-09-29

Abstract

Filtration systems, methods, and devices disclosed herein can be used to differentiate among cell types in whole blood, in particular, to separate platelets and/or plasma from erythrocytes and/or leukocytes in the blood. By using one or more filters, platelets and/or plasma may be allowed to pass through the filter while leaving other blood cells behind. A second filter can be used, simultaneously or sequentially, to further separate the platelets from plasma. The filters may be oriented such that direction of gravity encourages components away from the discriminating filter so as to minimize blockage. The filters can form a part of the walls of microchannels, whereby the passage of fluid acts to sweep components from the filter surface. Clinically relevant quantities of platelets can thus be isolated for use, for example, in emergency medical situations.

Description

FILTRATION SYSTEMS, METHODS, AND DEVICES TO DERIVE COMPONENT FRACTIONS FROM WHOLE BLOOD

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application claims the benefit of U.S. Provisional Application No.

61/316,293, filed March 22, 2010, which is hereby incorporated by reference herein in its entirety.

STATEMENT REGARDING FEDERALLY-SPONSORED RESEARCH

This invention was made with U.S. government support under project no. 5 R21

HL088162-02, awarded by the National Institutes of Health, National Heart, Lung, and Blood Institute. The U.S. government has certain rights in the invention.

FIELD

The present disclosure relates generally to blood processing technology, and, more particularly, methods, devices, and systems for filtration of whole blood in order to derive component fractions thereof.

BACKGROUND

Blood components are often necessary to treat injured victims in the field. The life- saving properties of platelets, however, come with a short time window. Efforts to expand the usable life of this important blood component have thus far met with very little success. Fresh platelets, used within a few days of donation, are the most effective; however, the logistics associated with maintaining an adequate supply to remote locations or in emergency situations. The ability of field hospitals or triage centers to process their own whole blood into familiar components may enable more immediate life saving aid to injured victims. Blood component separation for the purpose of transfusions, are conducted primarily on instruments which uses a centrifuge to separate the main components of blood.

Centrifugation makes use of the varying densities of red blood cells (RBC), platelets and plasma to concentrate them into localized regions within the centrifuged volume. During centrifugation, RBC will migrate in the radial direction to the outermost region of centrifuge, platelets will migrate and collect in the intermediate regions of the blood volume and cell free plasma will collect in the region closest to the center of the centrifuge. The volumes within these separated regions are collected to have RBC (up to 70% hematocrit), platelet rich plasma and cell free plasma.

Centrifugation has proven to be a very reliable way to separate blood components with minimal damage. Therefore, it is the main process used by modern blood banks.

However, the size, process time and cost of these instruments make them impractical for certain applications where the separation needs to be carried out in the field where space and power sources may be limited. The main drawbacks of commercially available blood component separators can include processing times, which can take multiple hours to complete, high power consumption to operate the centrifuge, and a size and weight comparable to small refrigerators that inhibits portability.

SUMMARY

Filtration systems, methods, and devices disclosed herein can be used to differentiate among cell types in whole blood, in particular, to separate platelets and/or plasma from erythrocytes and/or leukocytes in the blood. By using one or more micropore or nanopore filters, platelets and/or plasma may be allowed to pass through the filter while leaving other blood cells behind. A second micropore or nanopore filter can be used, simultaneously or sequentially, to further separate the platelets from plasma. The filters may be oriented such that direction of gravity encourages components away from the discriminating filter so as to minimize blockage. The filters can form a part of the walls of microchannels, whereby the passage of fluid acts to sweep components from the filter surface. Clinically relevant quantities of platelets can thus be isolated for use, for example, in emergency medical situations.

In embodiments, a method of filtration to derive component fractions from whole blood, can include passing a stream of whole blood across a first face of a first filter so as to produce a filtrate stream on a second opposite face of the first filter. The filtrate stream can be substantially free of erythrocytes. The method can also include passing the filtrate stream across a first face of a second filter so as to produce a substantially cell-free plasma stream on a second opposite face of the second filter.

In embodiments, a device for filtration of whole blood to derive component fractions thereof can include first through third microchannels, a first filter, and a second filter. The first filter can be arranged between the first and second microchannels. The first filter can be constructed to prevent the passage of erythrocytes therethrough. The second filter can be arranged between the second and third microchannels. The second filter can be constructed to prevent the passage of platelets therethrough. The first microchannel can be configured to accept a flow of whole blood therein.

In embodiments, a device for filtration of whole blood to derive component fractions thereof can include one or more first filters and one or more second filters. Each first filter can have pores with diameters between 2μιη and 5μιη. The one or more first filters can have a filtration area of at least 200 cm . The one or more second filters can each have pores with diameters between 0.2μιη and Ι .Ομιη. The one or more second filters can also have a filtration area of at least 200 cm . Each first filter can be arranged with respect to a corresponding one of the second filters such that a filtrate of each first filter is incident on a filtering surface of the corresponding one of the second filters. Each first filter can also be constructed such that platelets in whole blood pass through the first filter when the whole blood is incident on a respective filtering surface of said each first filter. Each second filter can also be constructed such that platelets do not pass through the second filter when the platelets are incident on the respective filtering surface of said each second filter.

In embodiments, a method of separating platelets from whole blood can include passing blood with platelets over a first flat filter. The first flat filter can have a smooth surface and pores of uniform diameter. The pores can be sized to permit the passage of platelets therethrough but to block the passage of erythrocytes. The method can further include sweeping erythrocytes from the first flat filter smooth surface by maintaining a sufficient shear rate of the blood over the first flat filter smooth surface and recovering an erythrocyte-depleted filtrate from the first flat filter. The filtrate can be flowed over a second flat filter. The second flat filter can have a smooth surface and pores of uniform diameter. These pores can be size to block the passage of platelets. The method can further include sweeping platelets from the second flat filter smooth surface by maintaining a sufficient shear rate of the erythrocyte-depleted filtrate over the second flat filter smooth surface whilst permitting plasma to flow through the second flat filter pores thereby to concentrate platelets in the erythrocyte-depleted filtrate and to form a platelet-enriched stream. The method can also include maintaining the first flat filter in a horizontal orientation above the second flat filter, and storing or transfusing the platelet-enriched stream.

In embodiments, a system for separating platelets from whole blood can include a pump. The pump can be configured to pump blood through a channel having a first flat filter in a wall thereof. The first flat filter can have a smooth surface and pores of uniform diameter. The pores can be sized to permit the passage of platelets therethrough but to block the passage of erythrocytes. The channel can be configured such that erythrocytes may be swept from the first flat filter smooth surface by maintaining a sufficient shear rate of the blood over the first flat filter smooth surface. The system can further include a second channel and a recovery channel. The second channel can be configured to permit the recovery of an erythrocyte-depleted filtrate passing through the first flat filter and to permit the flowing of the filtrate over a second flat filter. The second flat filter can have a smooth surface and pores of uniform diameter. These pores can be sized to block the passage of platelets. The second channel can also be configured to permit the sweeping of platelets from the second flat filter smooth surface through shear developed at the second flat filter smooth surface by the movement of the erythrocyte-depleted filtrate over the second flat filter smooth surface while permitting plasma to flow through the second flat filter pores thereby to concentrate platelets in the erythrocyte-depleted filtrate and to form a platelet-enriched stream. The recovery channel can be configured to permit recovery of filtrate from the second channel and to convey the filtrate to a storage vessel.

Objects and advantages of embodiments of the present disclosure will become apparent from the following description when considered in conjunction with the

accompanying drawings.

BRIEF DESCRIPTION OF DRAWINGS

Embodiments will hereinafter be described in detail below with reference to the accompanying drawings, wherein like reference numerals represent like elements. The accompanying drawings have not necessarily been drawn to scale. Indeed, the scale of some features may be exaggerated to assist in the description. Where applicable, some features may not be illustrated to assist in the description of underlying features.

FIG. 1 is a diagram showing the principles of blood component separation, according to one or more embodiments of the disclosed subject matter. FIG. 2 shows a blood component separation device where filling of the device produces separation of components, according to one or more embodiments of the disclosed subject matter.

FIGS. 3A-3B illustrates operation of a blood component separation device incorporating a piston for producing separation of components, according to one or more embodiments of the disclosed subject matter.

FIG. 4 illustrates sweeping of the filter surface by directional fluid inlets, according to one or more embodiments of the disclosed subject matter.

FIGS. 5A-5C illustrates operation of a blood component separation device with filter inserts to produce separation of components, according to one or more embodiments of the disclosed subject matter.

FIGS. 6A-6C illustrates operation of a blood component separation device using threaded filter inserts to produce separation of components, according to one or more embodiments of the disclosed subject matter.

FIG. 7 is a simplified schematic diagram of a flow-through blood component separation system, according to one or more embodiments of the disclosed subject matter.

FIG. 8 is a simplified schematic diagram showing variations in filter arrangement of a flow-through blood component separation system, according to one or more embodiments of the disclosed subject matter.

FIG. 9 is an isometric view of microfluidic channels in a flow-through blood component separation system, according to one or more embodiments of the disclosed subject matter.

FIG. 10 is a simplified schematic diagram of an array of microfluidic channels in a flow-through blood component separation system, according to one or more embodiments of the disclosed subject matter. DETAILED DESCRIPTION

The present disclosure is directed toward the fractionation of whole blood into discrete, clinically useful components (i.e., concentrated red cells, platelet-rich plasma, and cell-free plasma) without the use of centrifugation. In particular, filters can be used to size select between cells in whole blood. Appropriate control of the filter size, filter arrangement, and/or flow conditions can allow for effective filtration without blocking or fouling of filter pores due to cell accumulation on the filter surface. A thin filter having precisely defined pores that occupy a large percentage of the filter area can be used to select between cells or other components in the whole blood (or a filtrate thereof) based on size. The filters can be very thin and can be supported on a matrix which has considerably larger pores. For example, the filters can have a thickness less than 1 μιη, although other thicknesses are also possible.

Referring to FIG. 1, blood can be provided to a device 100 and contacted with a filter 106 having pores selected for a particular component of the blood, e.g., platelets 118. The pores of the filter 106 can have a precisely defined diameter. For example, the pores of the filter 106 may have diameters between 2 and 5 μιη. Such a pore size and configuration may inhibit the passage of erythrocytes 116 and leukocytes (not shown) through the filter 106 while allowing plasma and platelets 118 to pass therethrough. The blood can be flowed in direction 122 in volume or channel 110 while in contact with the filter 106. Filter 106 and wall 102 can thus define a portion of channel 110. The blood can pass at a low shear rate and at a very low transmembrane pressure (i.e., less than 5 Torr) over the filter 106.

Plasma and platelets 106 passing through the pores of the filter 106 can then be collected in a second volume or channel 112. Filter 106 and a second filter 108 can define a portion of the second channel 112. Similar to the fluid in channel 110, the fluid and components in channel 112 can also be flowed in a direction 124, i.e., as a second low-shear flow parallel to the initial flow 122. The second flow can pass in contact with filter 108, which can have pores with a precisely defined diameter selected for another component of the blood. For example, the pores of filter 108 may have diameters between 0.2 and 1.0 μιη. Such a pore size and configuration may inhibit the passage of platelets 118 through filter 108 while allowing plasma to pass therethrough and concomitantly the concentration of platelets in the second flow stream. Thus, filter 108 can allow the extraction of cell free plasma into volume or channel 114. Filter 108 and wall 104 can define a portion of channel 114. As with the other two channels, fluid and components in channel 114 can also be flowed in a direction 126.

The design of and the operating conditions imposed on the filter 106 are carefully controlled so as to afford discrimination between various blood components, for example, erythrocytes 116 and platelets 118. Erythrocytes are flexible and are able to penetrate pores that are smaller in diameter than that of the cell. The use of a precisely defined pore size and flow conditions which minimally distort the erythrocyte can act together to effect the platelet- erythrocyte separation.

The two parallel pore structures (i.e., filters 106 and 108) can be arranged such that the pore faces of the structures are perpendicular to the direction of gravity, denoted as 'g' in the figures. For example, the first flow stream 122 of whole blood can be generated adjacent to a lower face of filter 106. The filter 106 can be effective to retain erythrocytes 116 in the first flow stream 122 while allowing the passage of platelets 118 and plasma therethrough.

The passage of platelets 118 and plasma into a fluid layer 112 adjacent to and above an upper face of the filter 106, which produces platelet-containing plasma stream 124.

The second filter 108 can be effective to retain platelets 118 while allowing the passage of plasma therethrough. The second filter 108 can be arranged above the platelet- containing plasma stream 124, such that the platelet-containing plasma stream 124 is adjacent to and below a lower face of the second filter 108. The passage of plasma from the platelet- containing plasma stream 124 through the second filter 108 into a top layer 114 above the second filter 108, at a controlled rate, can produce a substantially platelet-free plasma stream 126 above the second filter 108. Extraction of plasma into the top layer 114 thus leaves a platelet-rich stream 124 in the volume 112 between the filters 106, 108. The orientation of the filters and flows with respect to gravity can be used as a natural force to cause particles whose passage through the filters is not desired to naturally move away from each filter surface, as shown in FIG. 1.

In embodiments, a system for fractionation of whole blood may separate fresh whole blood (FWB) into three component streams: a first stream with packed erythrocytes, a second platelet-rich plasma stream, and a third, substantially acellular plasma stream. The system may operate, for example, on citrate-anticoagulated blood at a flow rate of, for example, at least 7.5 mL/min. At such exemplary flow rates, the system can process one unit of blood in less than one hour. Such a system can weigh less than, for example, 500 g.

For the first and second filters of the system, micropore filters, such as those provided by Aquamarijn BV (Zutphen, Netherlands) may be used. For example, micropore filters can be prepared by lithographic or other techniques. In embodiments, the filters may be mounted on silicon wafers about 675 μιη thick, which provide support and collecting channels for the thin filter layer.

The micropore filters can have precisely sized and shaped pores over relatively large areas. Pore sizes for the filters may range from, for example, 0.2μιη to 5μιη. The micropore filters can have a large open area fraction, for example, on the order of 23-25%. The pores can be non-branching channels that extend through a thickness of the filter. The filters may be less than, for example, 1 μιη in thickness, so as to result in a very low flow resistance. These pores may be arranged in a regular array with a substantially uniform size. The surface of the filter in contact with the fluid to be filtered, i.e., a filtration surface, can be a substantially smooth surface with the pores having respective axes extending perpendicular to the smooth surface.

Filtration requires, by definition, that at least one component be "rejected" and in practical circumstances this component must be cleared from the filter surface. By controlling the relationship among flowrate through each filter, fluid flow over each filter surface, and the effect of filtration conditions on the penetration of the highly deformable erythrocyte, the clearance of components from the filter surface may also be controlled. The cross sectional area of the filters can be selected to allow for low-pressure filtration but while permitting sufficient speed in the separation process.

As discussed above, three streams 122, 124, and 126 can be formed, each having concentrated red cells 116, plasma rich with platelets 118, and cell-free plasma, respectively. However, it is noted that flow of the blood and components parallel to the filter surface is not necessarily required to effect discrimination between blood components. As explained in further detail herein, component discrimination may also be accomplished using non-flowing volumes by providing relative movement between the blood and the filters, for example, by filling a designated volume with filters therein, by displacing the blood volume with respect to the filters, or by displacing the filters with respect to the volume.

Referring to FIG. 2, a blood component separation system 200 that does not necessarily result in flow streams of separated components parallel to the filter surfaces is shown. Platelet-erythrocyte separation can be accomplished by achieving a controlled, low- velocity flow, upward in the gravitational field, g, through a horizontal filter 204 with pores selected to allow the passage of platelets therethrough but to restrict the passage of erythrocytes. Filtration may thus be accomplished in the absence of any lateral (shear) flow. For example, if the filter has an en- face area of 150 cm , the velocity required to process 7.5 ml/min would be 0.05 cm/min. One unit of blood (450 ml) under such a filter would have a height of 3 cm. Typical erythrocyte sedimentation rates are 1-2 cm/hr, indicating that most erythrocytes would not contact the filter during most of the plasma extraction process. This process may decrease the height of the blood layer by about 1.65 cm, assuming a hematocrit of 0.45. Platelet settling rates are on the order of 3 μηι/ηώι, and they are expected both to remain suspended and to be convected upward by the settling

erythrocytes. In the absence of appreciable pressure gradients and shear flow, the filter stabilizes the interface between blood and platelet-bearing plasma and can provide an absolute barrier to erythrocyte penetration.

Separation between plasma and platelet-rich plasma can be accomplished by achieving a second controlled, low-velocity flow, also upward in the gravitational field through a second horizontal filter 206 with pores of a diameter significantly smaller than that of a platelet. This second filter 206 can be placed above a growing layer of platelet bearing plasma. Flow through the second filter can be regulated so as to generate a substantially cell- free plasma layer above the second filter, leaving a platelet concentrate below. For example, the flow rates in flow streams adjacent to the filters may be greater than 0.05 cm/min such that platelets do not clog the filters.

In FIG. 2, blood may be provided to a first chamber 212 through inlet 208 so as to progressively fill the container 202. Once a sufficient volume of blood has been introduced into chamber 212 through inlet 208, it comes into contact with filter 204, which has pores sized (for example, 3μιη in diameter) to prevent the passage of erythrocytes but allow the passage of platelets therethrough. Blood can be continually introduced into chamber 212, such that platelets and plasma in chamber 212 move into chamber 214 through filter 206. Similarly, as the volume increases, the platelets and plasma in chamber 214 are brought into contact with filter 206, which prevents the passage of platelets therethrough. The plasma can thus move into chamber 216 while platelets remain in chamber 214 and erythrocytes in chamber 212. The filling can continue until the container is filled, with each chamber 212, 214, and 216 containing a specified blood component.

Although not required, filters 204 and 206 can be designed to have the same filtration area, A (i.e., based on W₂ and the filter geometry), which in turn determines the appropriate size (e.g., height) of chambers 212, 214, and 216 in order to accommodate a specific blood sample volume. For example, when dealing with a 450mL blood sample having 50mL of anticoagulant therein, the total height of the three chambers can have a value of 500cm /A. The first chamber 212 can finish the process with approximately 250mL of blood cells in plasma. This would concentrate the blood cells by a factor of 1.8, leading to a hematocrit of 0.8 for a starting hematocrit of 0.45. The second chamber 214, housing the concentrated platelets, can have a volume of 50mL. The third chamber 216, which houses the remaining plasma, can contain the remaining volume, i.e., 200mL. The relative heights of these chambers can be determined based on the filter area, i.e., 250/A, 50/A, and 200/A, respectively. For example, the total filter surface area, A, of each filter can be on the order of 200cm , thereby leading to a total height, H₂, of the device of approximately 2.5cm.

The inlet 208 for chamber 212 can be located near the surface of the filter 204, such that incoming blood flows serve to sweep from the filter surface components that may have become stuck there. Alternatively or additionally, chambers 212 and 214 may include respective inlets 222 and 224 located near the respective surface of filters 204 and 206. The inlets 222 and 224 may be used to introduce a flow to sweep from the filter surface components that may have become stuck there. The source of the flow for inlets 222 and 224 may be provided from the contents of the respective chambers 212 and 214, for example, through outlets 210 and 218. Outlets 210, 218, and 220 may also serve to remove the contents of the individual chambers 212, 214, and 216, respectively, once separation of components is completed.

Alternatively, the uppermost contents and filters may be sequentially removed to obtain access to the contents of the individual chambers 212, 214, and 216. For example, substantially cell free plasma layer in volume 216 may be collected, after which filter 206 can be removed to provide access to volume 214. Similarly, the platelet-rich plasma in volume 214 may then be collected, after which filter 204 can be removed to provide access to volume 212. The concentrated erythrocytes in the bottom volume 212 can then be collected.

Referring now to FIGS. 3A-3B, a blood component separation device 300 employing a piston 330 for moving an initial blood volume 312 through one or more filters is shown. For example, the separation device 300 can have a cylindrical chamber with a circular area of 150cm (i.e., diameter, W₃, of about 27.6cm) and a height, H₃, of about 10cm. The chamber can contain a movable piston 330 or diaphragm.

In FIG. 3A, the chamber has just been charged with a 450ml unit of blood into the space 312 between the piston 330 and a filter 304 that discriminates between erythrocytes and platelets. The piston 330 can be moved upward at a velocity of, for example, 0.05 cm/min, thus driving 7.5 ml/min of fluid through both the platelet-permeable filter 304 and the platelet-retaining filter 306. The initial volume 314 between the filters 304, 306 may be filled with a fluid, for example, sterile saline. An initial volume 316 of fluid may also be provided above filter 306. The distance between the first and second filters 304, 306 may be chosen based on the expected volume of the platelet-rich component that is produced after the piston has traveled a predetermined distance, for example, about 1.5cm. FIG. 3B shows the position of piston 330 after processing is complete. The displacement of the volume of blood initially in chamber 312 by piston 330 has produced a volume 342 of concentrated erythrocytes, a volume 344 between filter 304 and 306 of platelets, and a volume 346 above filter 306 of substantially cell-free plasma.

In embodiments, the movement of blood into contact with the filter without a lateral (shear) flow may result in accumulation of blood components on the surface of the filter, thereby resulting in clogging or fouling, which may reduce system efficiency or result in system damage. The arrangement of components such that filtration is achieved against the direction of gravity helps to reduce the impact of cell accumulation on the filter surface, as discussed above. Alternatively or additionally, the inlet or outlet arrangement in a chamber may be defined to induce an in chamber flow to sweep the filter surface of accumulated components. For example, in FIG. 4, device 402 has a chamber 404 includes one or more inlets 408 that have been tangentially arranged. This arrangement may induce a circular flow 410 within chamber 404 which may serve to sweep the surface of filter 406 and prevent accumulation of blood components thereon. Such an arrangement may be employed when filling a chamber, as in the embodiment of FIG. 2, when moving the blood with respect to static filters, as in the embodiment of FIGS. 3A-3B, when moving the filters with respect to a static volume of blood, as in the embodiments of FIGS. 5A-6C, or in any other contemplated embodiment.

Referring now to FIGS, 5A-5C, a setup is shown whereby a static volume of blood is subject to component separation by displacing filters with respect thereto. In FIG. 5A, a container 502 having an interior volume 504 can hold a quantity of whole blood 506. In FIG. 5B, a filter 512 mounted on a filter holder 508 can be inserted into the interior volume 504 of container 502 a predetermined distance. As discussed above, the filter 512 can be inserted into the whole blood 506 at a rate that allows processing of at least 7.5 ml/min of blood. Filter 512 is designed with pores that allow platelets and plasma to pass through, but prevent erythrocytes from passing through. As a result, an erythrocyte -rich volume 514 is formed below filter 512 while a platelet-plasma volume 516 is formed above filter 512. O-rings 518 can be optionally provided between the filter holder 508 and the container 502 (or between the filter 512 and the container 502) to prevent fluid or cells from circumventing the filter.

In FIG. 5C, a filter 524 mounted on a filter holder 520 can be inserted at a

predetermined rate into the interior volume 510 of filter holder 508 a predetermined distance. The filter 524 can be inserted into this volume 510 at a rate that allows for the desired processing rate of blood, e.g., at least 7.5 ml/min. Filter 524 is designed with pores that prevent the passage of platelets through the filter. As a result, the platelets are concentrated in region 526 below filter 524, while a substantially cell-free plasma volume 528 is formed above filter 524 in the interior volume 522 of holder 520. O-rings 530 can be optionally provided between the filter holder 520 and the filter holder 508 (or between the filter 524 and the container 502) to prevent fluid or cells from circumventing the filter.

Although shown as sequential steps, the introduction of filters 512 and 524 into blood volume 506 may occur at the same time. In addition, the specific arrangement and orientation of filter holders 508 and 520 should not be understood as limiting. For example, filter holder 508 may extend downward from filter 512 so as to precisely define the size of volume 514. Filter holder 520 may also extend downward in a similar manner so as to precisely define the size of volume 526. Moreover, the use of o-rings 518 and 530 are not necessarily required. Other means for sealing the filter to container 502 (or filter holders 508/520) can also be employed. For example, the filter holders 508 and 520 may be designed with an interference fit with respect to container 502, such that fluid cannot circumvent filters 512 and 524.

Insertion of filters 512 and 524 into the blood volume 506 may be done manually. Alternatively, the insertion of filters 512 and 524 into the blood volume 506 is accomplished by actuators, which may be precisely controlled to vertically translate each filter to achieve a desired blood processing rate. In an alternative embodiment, the filter holders and/or filters may be displaced into the blood volume by threaded displacement. As shown in FIGS. 6A- 6C, a threaded rod 603 may be disposed in container 602. Disposed within the internal volume 604 of container 602 is an initial blood volume 606. In FIG. 6B, a filter 612 mounted on a filter holder 608 can be inserted into the interior volume 604 of container 602. As the filter holder 608 is rotated on rod 603, a threaded portion 618 of filter holder 608 interacts with the threads on the rod 603 to provide controlled vertical displacement together with rotation of the filter 612. This rotation may help to sweep cells from the filter surface so as to reduce accumulation and/or fouling. The rate of rotation of the filter holder 608 may be selected to provide processing of, for example, at least 7.5 ml/min of blood. Filter 612 is designed with pores that allow platelets and plasma to pass through, but prevent erythrocytes from passing through. As a result, an erythrocyte-rich volume 614 is formed below filter 612 while a platelet-plasma volume 616 is formed above filter 612.

In FIG. 6C, a filter 624 mounted on a filter holder 620 can be inserted into filter holder 608. As the filter holder 620 is rotated on rod 603, a threaded portion 630 of filter holder 620 interacts with the threads on the rod 603 to provide controlled vertical

displacement together with rotation of the filter 624. As with filter 612, this rotation may help to sweep cells from the filter surface so as to reduce accumulation and/or fouling. The rate of rotation of the filter holder 620 may be selected to provide processing of, for example, at least 7.5 ml/min of blood. Filter 624 is designed with pores that do not allow platelets to pass through. Thus, a concentrated platelet volume 626 is formed between filters 612 and 624, while a substantially cell-free plasma volume 628 is formed above filter 624.

Although shown as sequential steps, the introduction of filters 612 and 624 into blood volume 606 may occur at the same time. In addition, the specific arrangement and orientation of filter holders 608 and 620 should not be understood as limiting. For example, filter holder 608 may extend downward from filter 612 so as to precisely define the size of volume 614. Filter holder 620 may also extend downward in a similar manner so as to precisely define the size of volume 626. Moreover, o-rings may be used for sealing the filter holder or the filter to the container 602. Other sealing techniques can also be employed. For example, the filter holders 608 and 620 may be designed with an interference fit with respect to container 602, such that fluid cannot circumvent filters 612 and 624.

The specific example of threaded rod 603 is not to be understood as limiting, but merely illustrative of the concept of controlled displacement by rotating the filters into place. Alternative configurations are possible according to one or more contemplated embodiments. For example, the internal surface of container 602 may be threaded and the circumferential end of the filter or filter holder designed to interact with the threads to effect displacement of the filter into the fluid volume. In another example, the threaded component is not a portion of the container 602 at all, but is part of the actuator. In such a configuration, the actuator provides the filter with a rotational displacement together with the vertical displacement.

In embodiments, a high volume cross flow filtration system can be used to separate blood into three component streams, for example, a stream with red bloods, a platelet-rich plasma stream, and a substantially acellular plasma stream. As shown in FIG. 7, such a system 700 may include, for example, a component separation module 704, a flow regulation module 712, a control module 708, a detection module 716, and component storage module 718. The component separation module 704 may include one or more micro fluidic-based blood component separation devices, 706. The blood component separation device can include a lower filter that allows platelet and plasma to move into the middle flow channel while restricting erythrocytes to the bottom flow channel. The separation device 706 can also include an upper filter that restricts platelets to the middle flow channel but allows plasma to move into the top flow channel. The component separation module 704 may also include one or more manifolds (not shown) for distributing flows to the flow channels in the one or more blood component separation devices 706.

Flow control module 712 can be designed to move the fluid and components through the component separation module 704. For example, flow control module 712 can include one or more pumps 714, such as a peristaltic pump. Flow control module 712 can also include (but not shown) various conduits, valves, pumps, sensors, and the like for controlling fluid flows. Control module 708 can control operation of the system 700, including pumps 714 in flow control module 712. Detection module 716 can include various sensors and/or monitoring equipment for monitoring and regulating the blood component separation process. For certain measurements, components of the detection module 716 may be integrated with the component separation module 704, for example, to monitor transmembrane pressure or other flow conditions in the separation device 706.

The component separation module 704, the control module 708, the detection module 716, and the flow control module 712 may be configured as a single unit 702 with its own power supply 710. Component storage module 718 or components thereof may be provided as a separate unit, for example, by an end user. Fluid lines 728 from the component storage module 718 may be connected to unit 702 through connections 726, for example, in a wall of a housing of the unit 702. The same unit 702 may thus be able to process multiple samples of whole blood.

Actual separation of the individual blood components can be achieved in the separation module 706. Filtration of incoming anti-coagulated whole blood from bag 720 in component storage module 718 can be achieved by carefully regulated shear flow over a first nanopore filter configured to retain red blood cells and leukocytes in the first flow channel but allowing platelets to pass therethrough. The resulting filtrate in the second flow channel will include platelets and plasma. The resulting filtrate can pass immediately over a second nanopore filter designed to retain platelets but allowing plasma to pass therethrough. Plasma thus is able to pass through this filter into the top channel. Three separate fluid streams can then emerge from the separation module 704: a stream of packed red blood cells (for example, at a hematocrit of at least 0.7), a platelet-rich plasma stream, and a substantially cell-free plasma stream. The filters can have pore sizes ranging from, for example, 0.45 μιη to 5 μιη depending on the desired discrimination between various blood components.

For example, blood storage bag 720 may initially contain 450ml of citrated whole blood. As the blood repeatedly flows through lower channel of the component separation module 704, the middle flow channel will fill. The platelet/plasma mixture is pumped around a loop including the middle channel of the component separation module and the platelet storage bag 722 of the storage module 718. As the platelet/plasma mixture is pumped around this second loop, plasma will pass through the second filter to fill the top channel in the component separation device. As the plasma is pumped around this third loop, the plasma storage bag 724 of the storage module 718 will fill. For example, after a predetermined processing time (e.g., 1 hour) the three collection bags in storage module 718 will contain erythrocytes at a hematocrit in the range of 0.6 to 0.8 (blood storage bag 720), platelets with plasma with a count of at least 5.5 x 10¹⁰ platelets^L (platelet storage bag 722), and substantially accelular plasma (plasma storage bag 724).

Because of the flexibility of red blood cells, it may be desirable to operate at a very low pressure difference that is uniform over a relatively larger filter area. Even though the red blood cell may be substantially larger than a proposed pore size of the first filter, this cell is known to pass through orifices smaller than its diameter. Therefore, a low transmembrane pressure may decrease the probability of pushing a red blood cell through the first and/or second filters. Pressure differences across the lower filter may be less than 5 Torr at all points of the filter, for example, between 1 Torr and 3 Torr. Sensors in detection module 716 can be used to monitor the progress of the filtration, for example, by observing the content of the different streams. Sensors in the detection module can also be used to help adjust flow conditions (e.g., if transmembrane pressures are causing erythrocytes to pass through the first filter and show up in the middle flow channel circuit) or monitor for system failure (e.g., if one of the filters should fail).

In FIGS. 1 and 7, the filters are shown arranged in parallel and at substantially the same region in the device. However, it is not required for the filters to be disposed in this manner. For example, as shown in system 800 of FIG. 8, it is contemplated that the erythrocyte-platelet discrimination filter 808 can be disposed in one flow channel

arrangement 806a while the platelet-plasma discrimination filter 810 is disposed in a connected but different flow channel arrangement 806b. For example, the platelet-plasma discrimination filter 810 can be disposed at a different location of the flow channel as the erythrocyte-platelet discrimination filter 808. In another example, the platelet-plasma discrimination filter 810 can be disposed in a second flow channel different from a first flow channel in which the erythrocyte-platelet discrimination filter 808 is disposed. Appropriate fluid conduits can convey the output of the first fluid channel to the second fluid channel. Together, flow channel arrangements 806a and 806b may be considered to form a component separation module 804.

The flow of blood within the separation device may be through microchannels. For example, as shown in FIG. 9, three microchannels 914, 916, and 918 may be disposed in a layered fashion to form a separation module 900. Fluid communication between adjacent microchannels is provided by one or more wall filters 910, 912. Filter 910, formed in wall 906, is sized and shaped to allow platelets therethrough while restricting erythrocytes to channel 914. Filter 912, formed in wall 904, is sized and shaped to allow only plasma to pass through to channel 918. Walls 906 and 908 additionally define the bottom microchannel 914. Walls 902 and 904 define the top microchannel 918 while walls 904 and 906 define the middle microchannel 916.

Each of the microchannels can have a height (i.e., ¾τ, ¾M, ¾B) in the range of 80- ΙΟΟμιη. The filters 910, 912 can have a width (i.e., W₉) on the order of 3mm, which is determined in the direction perpendicular to the flow direction. The filters 910, 912 can be constructed to minimize their length (i.e., L9, which is in the direction of flow), as even low flows in a microfluidic channel can cause pressure over the filter surface to vary in the direction of flow. Thus, wide systems with correspondingly less lengths are desirable for a given filter area. If a particular width for a given length may be too large, multiple microfluidic channels with respective filters can be operated in parallel to achieve the desired filter shown in FIG. 10.

In such microchannels, the flow may be controlled such that cells migrate toward the center of the flow when fluid shear rates are sufficiently small. Such a migration may lower the load and shield cells from unnecessary wall and/or filter contact. Scaling to different flows, larger or smaller, can be accomplished by increasing the width of the flow channels and, when the width is inconveniently large, stacking channels on top of each other, as shown in FIG. 10. In particular, multiple component separation modules 900 are layered on top of each other, with an optional spacer layer 1002 or supporting structure in between. Although only three component separation modules 900 are shown, any number is possible to achieve a desired filtration area. Setup 1000 in FIG. 10 can also include one or more inlet manifolds 1004 and outlet manifolds 1006, which distribute to and collect from the microchannels 914, 916, and 918. The manifolds 1004 and 1006 may be specifically designed to ensure substantially similar pathlengths to each channel to reduce the likelihood of undesired pressure variations between the microchannels. Although separation of specific blood components has described herein, the types of blood components that may be separated using the disclosed techniques are not necessarily limited to those specific components. Separation of various other blood components is of course possible with appropriate selection of filter and flow characteristics. Accordingly, the number of filters is limited only by the number of components desired to be separated.

Different filter sizes and filtering characteristics as well as a different number of filters can be used to isolate different components in whole blood or other biological fluids.

Although particular configurations for inlets and outlets are shown in the figures, the embodiments of the disclosed subject matter are not limited to these configurations. Rather, other configurations and arrangements for inlets and outlets, as well as their inclusion or omission, are also possible according to one or more contemplated embodiments. In addition, sensors and structural features have not necessarily been shown in the figures. Physical embodiments of the disclosed subject matter may include sensors and structural features not presently illustrated.

In embodiments, the systems disclosed herein may be adapted as a highly portable unit. Moreover, as the systems need not require large amounts of electrical power or supporting elements, they can be adapted as a wearable unit in some embodiments. The systems disclosed herein have particular application to emergency and military environments and other applications where typical blood separation technologies may be deficient. In embodiments, the blood component separation system can be configured as a field

deployable unit. Such a unit can allow collection of blood components for use in forward echelons of care, for example, in emergency and/or military applications.

Embodiments described herein can be configured as mobile systems or devices, such that they can be moved, for example, from room to room in a hospital. Embodiments can be deployed in mobile medical units, such an ambulance or medivac helicopter, for use by emergency personnel. Alternatively, the embodiments can be configured as a portable device that emergency responders can carry to a treatment site. In yet another alternative, embodiments can be configured as substantially fixed units with appropriate fluid

conveyances and/or storage vessels for interfacing with a treatment location. In still another alternative, embodiments can be fixed at a particular location, such as a floor or wing of a medical treatment facility, to process blood at that location.

Although particular configurations have been discussed herein, other configurations can also be employed. Furthermore, the foregoing descriptions apply, in some cases, to examples generated in a laboratory, but these examples can be extended to production techniques. For example, where quantities and techniques apply to the laboratory examples, they should not be understood as limiting.

Features of the disclosed embodiments may be combined, rearranged, omitted, etc., within the scope of the invention to produce additional embodiments. Furthermore, certain features may sometimes be used to advantage without a corresponding use of other features.

It is, thus, apparent that there is provided, in accordance with the present disclosure, methods, systems, and devices for filtration to derive component fractions from whole blood. Many alternatives, modifications, and variations are enabled by the present disclosure. While specific embodiments have been shown and described in detail to illustrate the application of the principles of the invention, it will be understood that the invention may be embodied otherwise without departing from such principles. Accordingly, Applicant intends to embrace all such alternatives, modifications, equivalents, and variations that are within the spirit and scope of the present invention.

Claims

1. A method of filtration to derive component fractions from whole blood, comprising: passing a stream of whole blood across a first face of a first filter so as to produce a filtrate stream on a second opposite face of the first filter, the filtrate stream being substantially free of erythrocytes; and

passing the filtrate stream across a first face of a second filter so as to produce a substantially cell-free plasma stream on a second opposite face of the second filter.

2. The method of claim 1, wherein at least a portion of the second face of the first filter faces at least a portion of the first face of the second filter.

3. The method of claim 1, wherein the first filter has pores with diameters greater than diameters of pores of the second filter.

4. The method of claim 1, wherein the first filter is constructed such that erythrocytes are prevented from passing therethrough and platelets are allowed to pass therethrough.

5. The method of claim 1, wherein the second filter is constructed such that platelets are prevented from passing therethrough

6. The method of claim 1 , wherein the first filter has pore diameters of 2-4 μιη.

7. The method of claim 1 , wherein the second filter has pore diameters of 0.2- 1.Ομιη.

8. The method of claim 1, wherein the first and second filters extend perpendicular to a direction of gravity.

9. The method of claim 1 , wherein the whole blood stream is arranged below the filtrate stream with respect to gravity, and the filtrate stream is arranged below the substantially cell- free plasma stream.

10. The method of claim 1, wherein the first and second filters are micropore or nanopore filters.

11. The method of claim 1 , wherein the first and second filters constitute portions of microchannels.

12. A device for filtration of whole blood to derive component fractions thereof, comprising:

first through third microchannels;

a first filter arranged between the first and second microchannels, the first filter being constructed to prevent the passage of erythrocytes therethrough; and

a second filter arranged between the second and third microchannels, the second filter being constructed to prevent the passage of platelets therethrough,

wherein the first microchannel is configured to accept a flow of whole blood therein.

13. The device of claim 12, wherein the first filter forms an upper wall portion of the first microchannel and a lower wall portion of the second microchannel, and the second filter forms an upper wall portion of the second microchannel and a lower wall portion of the third microchannel.

14. The device of claim 12, wherein the first filter has pores with diameters greater than diameters of pores of the second filter.

15. The device of claim 12, wherein the first filter has pore diameters of 2-4μιη.

16. The device of claim 12, wherein the second filter has pore diameters of 0.2-1. Ομιη.

17. The device of claim 12, wherein the first and second filters extend perpendicular to a direction of gravity.

18. The device of claim 12, wherein the first microchannel is configured to accept a flow of whole blood therein.

19. The device of claim 12, wherein the first and second filters are micropore or nanopore filters.

20. The device of claim 12, further comprising a flow control module, which is

configured to control the flow in the first through third microchannels such that erythrocytes are unable to pass through the first filter.

21. The device of claim 20, wherein the flow control module controls the flow in the microchannels such that a pressure difference across the first filter is less than 5 Torr at all points along the first filter.

22. A device for filtration of whole blood to derive component fractions thereof, comprising:

one or more first filters, each having pores with diameters between 2μιη and 5μιη, the one or more first filters having a filtration area of at least 200 cm ; and

one or more second filters, each having pores with diameters between 0.2μιη and 1.Ομιη, the one or more second filters having a filtration area of at least 200 cm ,

wherein each first filter is arranged with respect to a corresponding one of the second filters such that a filtrate of each first filter is incident on a filtering surface of the

corresponding one of the second filters,

each first filter is constructed such that platelets in whole blood pass through the first filter when the whole blood is incident on a respective filtering surface of said each first filter, and

each second filter is constructed such that platelets do not pass through the second filter when the platelets are incident on the respective filtering surface of said each second filter.

23. The device of claim 22, wherein each first and second filter has an open area fraction on the order of 23-25%.

24. The device of claim 22, wherein each first and second filter has a thickness in the respective filtration area of less than 1 μιη.

25. The device of claim 22, wherein the first filters are arranged parallel to respective ones of the second filters.

26. The device of claim 22, wherein the first filters each form part of a bottom wall of a respective microchannel, and the second filters each form part of a top wall of said respective microchannel.

27. The device of claim 22, wherein each first and second filter has a width on the order of 3 mm.

28. The device of claim 22, further comprising a control module configured to control fluid flows adjacent to each first and second filter such that a transmembrane pressure at each point along the respective surface for each filter is less than 5 Torr.

29. The device of claim 28, wherein the control module is further configured to control fluid flows adjacent to each first and second filter so as to process at least 7.5ml/min of blood.

30. A method of separating platelets from whole blood, comprising:

passing blood with platelets over a first flat filter with a smooth surface and pores of uniform diameter, which pores are sized to permit the passage of platelets therethrough but to block the passage of erythrocytes;

sweeping erythrocytes from the first flat filter smooth surface by maintaining a sufficient shear rate of the blood over the first flat filter smooth surface;

recovering an erythrocyte-depleted filtrate from the first flat filter and flowing the filtrate over a second flat filter with a smooth surface and pores of uniform diameter, which pores are sized to block the passage of platelets;

sweeping platelets from the second flat filter smooth surface by maintaining a sufficient shear rate of the erythrocyte-depleted filtrate over the second flat filter smooth surface whilst permitting plasma to flow through the second flat filter pores thereby to concentrate platelets in the erythrocyte-depleted filtrate and form a platelet-enriched stream; maintaining the first flat filter in a horizontal orientation above the second flat filter; and

storing or transfusing the platelet-enriched stream.

31. The method of claim 30, wherein the first and second flat filters have uniformly-sized pores that are in a regular array and free of branches.

32. The method of claim 30, wherein the first and second flat filters have uniformly-sized pores that are in a regular array and have axes that are perpendicular to a respective smooth surface.

33. The method of claim 30, further comprising fabricating the first and second flat filters by machining straight channels with axes perpendicular to the respective smooth surfaces of the first and second flat filters to form the pores therein.

34. A system for separating platelets from whole blood, comprising:

a pump configured to pump blood through a channel having a first flat filter in a wall thereof, the first flat filter having a smooth surface and pores of uniform diameter, which pores are sized to permit the passage of platelets therethrough but to block the passage of erythrocytes;

the channel being configured such that erythrocytes may be swept from the first flat filter smooth surface by maintaining a sufficient shear rate of blood over the first flat filter smooth surface;

a second channel configured to permit the recovery of an erythrocyte-depleted filtrate passing through the first flat filter and to permit the flowing of the filtrate over a second flat filter with a smooth surface and pores of uniform diameter, which pores are sized to block the passage of platelets;

the second channel being configured to permit the sweeping of platelets from the second flat filter smooth surface through shear developed at the second flat filter smooth surface by the movement of the erythrocyte-depleted filtrate over the second flat filter smooth surface whilst permitting plasma to flow through the second flat filter pores thereby to concentrate platelets in the erythrocyte-depleted filtrate and form a platelet-enriched stream; a recovery channel configured to permit recovery of filtrate from the second channel and convey the filtrate to a storage vessel.

35. The system of claim 34, wherein the first and second flat filters have uniformly-sized pores that are in a regular array and free of branches.

36. The system of claim 34, wherein the first and second flat filters have uniformly-sized pores that are in a regular array and have axes that are perpendicular to a respective smooth surface.