Patent Description:
Within the industry, plasma is usually obtained by centrifuging whole blood separating red blood cells from plasma. The centrifuging process, however, may be slow and require large powered instrumentation. Additionally, once the centrifugation process has begun, the instrument is unavailable for use by another operator until completion of the process. <CIT> provides a method of collecting a plasma fraction, wherein the channel assembly includes a channel housing which is rotatably interconnected with a rotatable centrifuge rotor assembly.

Progress within the medical industry has been in the development of point-of-care systems to provide rapid and portable care. Because of time restraints, size of equipment, and one-operator use, centrifugation is generally impractical for use in such point-of-care diagnostic instruments. For example, <CIT> pertains to a system and method for separating plasma and/or serum from blood for bilirubin level estimation. Other separation methods including laminar-flow filtration or capillary-action based processes, however, are also expensive, complex, slow, require large volumes of blood, or may lead to unacceptable levels of hemolysis. In <CIT> a capillary device is disclosed for analyzing blood using minimal sample volumes, e.g., fingerstick application. The plasma separation membrane to separate plasma from a blood sample is provided in <CIT>.

To assist those of ordinary skill in the relevant art in making and using the subject matter hereof, reference is made to the appended drawings, which are not intended to be drawn to scale, and in which like reference numerals are intended to refer to similar elements for consistency.

The following detailed description refers to the accompanying drawings.

As used in the description herein, the terms "comprises," "comprising," "includes," "including," "has," "having," or any other variations thereof, are intended to cover a non-exclusive inclusion. For example, unless otherwise noted, a process, method, article, or apparatus that comprises a list of elements is not necessarily limited to only those elements, but may also include other elements not expressly listed or inherent to such process, method, article, or apparatus.

Further, unless expressly stated to the contrary, "or" refers to an inclusive and not to an exclusive "or". For example, a condition A or B is satisfied by one of the following: A is true (or present) and B is false (or not present), A is false (or not present) and B is true (or present), and both A and B are true (or present).

In addition, use of the "a" or "an" are employed to describe elements and components of the embodiments herein. This is done merely for convenience and to give a general sense of the inventive concept. This description should be read to include one or more, and the singular also includes the plural unless it is obvious that it is meant otherwise. Further, use of the term "plurality" is meant to convey "more than one" unless expressly stated to the contrary.

As used herein, any reference to "one embodiment," "an embodiment," "some embodiments," "one example," "for example," or "an example" means that a particular element, feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment. The appearance of the phrase "in some embodiments" or "one example" in various places in the specification is not necessarily all referring to the same embodiment, for example.

Referring now to the Figures, and in particular to <FIG>, shown therein and designated by reference numeral <NUM> is an exemplary plasma separation system <NUM> in accordance with the present disclosure. Generally, the plasma separation system <NUM> may provide separation of plasma from blood using a vacuum force, and without using centrifugation or laminar-flow filtration based filtration processes. The separation of plasma from blood may be with minimal hemolysis and within a relatively short amount of time (e.g., <NUM>-<NUM> seconds) as compared to processes used currently within the industry. In some embodiments, the plasma may be further used in one or more point-of-care assays.

The plasma separation system <NUM> includes a housing <NUM> supporting or encompassing a blood separation well <NUM> connected via a first channel <NUM> to a plasma collection vessel <NUM>. The housing <NUM> supports or encompasses a second channel <NUM> connecting the plasma collection vessel <NUM> to an outlet port <NUM> downstream of the plasma collection vessel <NUM>. The plasma separation system <NUM> may also include a negative pressure source <NUM> that can be attached to the outlet port <NUM>. In this instance, the outlet port <NUM> can be configured to allow for the negative pressure source <NUM> to be attached thereto assist in enabling the operation of the plasma separation system <NUM>.

Generally, in the plasma separation system <NUM>, blood is added to the blood separation well <NUM>. Negative pressure (e.g., vacuum pressure) is applied by the negative pressure source <NUM> to the outlet port <NUM>, which also causes a vacuum to form in the blood separation well <NUM>. Using a combination of capillary action and negative pressure, plasma is separated from the blood. The magnitude of the negative pressure may be controlled to prevent hemolysis and/or leakage of cellular material. The separated plasma is collected in the plasma collection vessel <NUM>.

In some embodiments, the plasma separation system <NUM> may be a single-use system. Alternatively, one or more components of the plasma separation system <NUM> may be disposable such that the plasma separation system <NUM> may be a multi-use system. For example, in some embodiments, after a first blood sample is separated, one or more channels <NUM> and <NUM> may be lined such that the plasma collection vessel <NUM> and components of the blood separation well <NUM> may be removed, disposed of, and replaced for use with a second blood sample.

The housing <NUM> may be formed of materials including, but not limited to, glass, plastic, and/or the like. The shape and size of the housing <NUM> may be dependent on shape and/or size of the blood separation well <NUM>, channels <NUM> and <NUM>, and/or the plasma collection vessel18. The housing <NUM> that is shown in <FIG> is a unitary integral device that is shaped to form the blood separation well <NUM>, channels <NUM> and <NUM> and the plasma collection vessel <NUM>. It should be understood that the housing <NUM> can be constructed of separate components which are connected together so that the blood separation well <NUM>, channels <NUM> and <NUM> and the plasma collection vessel <NUM> communicate with each other. Generally, the size of the housing <NUM> may be minimized and determinate on an amount of plasma that is desired (e.g., <NUM>-<NUM>µL) to be extracted from a blood sample. The housing <NUM> may include a surface <NUM>.

The blood separation well <NUM> is positioned to intersect the surface <NUM> of the housing <NUM> such that the surface <NUM> at least partially surrounds the blood separation well <NUM>. The blood separation well <NUM> includes a recess <NUM> intersecting the surface <NUM> of the housing <NUM> as illustrated in <FIG> and <FIG> and in this instance the surface <NUM> may form a rim surrounding the recess <NUM>. The recess <NUM> may include a proximal end <NUM> and a distal end <NUM> with a wall <NUM> spanning the length of the proximal end <NUM> to the distal end <NUM>. The proximal end <NUM> may include a capillary surface <NUM> in which microchannels are formed in the proximal end <NUM> as discussed in more detail below with respect to <FIG>.

The plasma separation device <NUM> is also provided with a filter <NUM>, a separation membrane <NUM>, and an adhesive member <NUM>. The filter <NUM>, the separation membrane <NUM> and the adhesive member <NUM> are stacked on the capillary surface <NUM> and disposed within the recess <NUM> such that the filter <NUM> is below the surface <NUM> of the housing <NUM> and fully disposed within the recess <NUM>. For example, in some embodiments, the adhesive member <NUM> may be positioned on the capillary surface <NUM> with the separation membrane <NUM> and the filter <NUM> positioned thereon, respectively. The filter <NUM> and separation membrane <NUM> are pressure fit (also known as a "press fit") within the recess <NUM>.

The size and shape of the recess <NUM> may be dependent on the size and shape of the filter <NUM>, the separation membrane <NUM>, and/or the adhesive member <NUM>. For example, in some embodiments, the size and shape of the recess <NUM> may be circular as the filter <NUM>, separation membrane <NUM>, and adhesive member <NUM> are circular. The shape, however, may be any shape including, but not limited to, rectangular, triangular, or any fanciful shape. The volume of the recess <NUM>, including the height and/or length of wall <NUM> may be dependent on thicknesses and/or the widths of one or more of the filter <NUM>, separation membrane <NUM>, and adhesive member <NUM>. Generally, the filter <NUM>, separation membrane <NUM> and the adhesive member <NUM> may be positioned within the recess <NUM> such that each remains between the proximal end <NUM> and the distal end <NUM> of the recess <NUM>. Further, although the wall <NUM> is depicted as a straight wall, it should be understood that in some embodiments, the wall <NUM> can be stepped.

In some embodiments, one or more additional layers may be positioned on a first side <NUM> of the filter <NUM> to hold the filter <NUM> and separation membrane <NUM> within the blood separation well <NUM>. For example, in one example, a plastic O-ring may be positioned on the first surface <NUM> of the filter <NUM> to hold the filter <NUM> and separation membrane <NUM> within the blood separation well <NUM>. In another example, a cap (e.g., plastic formed cap) may be positioned adjacent or proximal to the first side <NUM> of the filter <NUM> and span the length of the blood separation well <NUM>. In some embodiments, the cap may be vented to allow for escape of gas to the outside environment. The cap may serve to hold the filter <NUM> in close proximity to the separation membrane <NUM>. In one example, as blood wicks into the filter <NUM> during use, the filter <NUM> may expand in size such that the cap retains the filter <NUM> within the blood separation well <NUM>. To that end, the filter <NUM> may expand in the direction of the separation membrane <NUM> positioning the filter <NUM> in close proximity or even in contact with the separation membrane <NUM>.

The filter <NUM> is formed of one or more layers <NUM>. Each layer <NUM> may include the first side <NUM> and a second side <NUM> generally opposite of the first side <NUM>. Generally, blood is provided onto and contacts the first side <NUM> of the filter <NUM> and passes through the layer <NUM> of the filter <NUM> such that filtered blood emerges on the second side <NUM>. The filtered blood may include plasma and a portion of red blood cells that managed to pass through the layer <NUM>. Some of the red blood cells will be captured within the layer <NUM> of the filter <NUM>.

In other words, the filter <NUM> generally removes a portion of the red blood cells from the blood and also may reduce separation burden of the separation membrane <NUM>. For example, the filter <NUM> may remove red blood cells (e.g., up to <NUM>% of red blood cells) within the blood sample. The removal of a significant portion of red blood cells may aid the flow of the filtered blood through the separation membrane <NUM> and also reduce clogging of the separation membrane <NUM>.

Each layer <NUM> may be formed of materials including, but not limited to, glass fibers, polyester fibers, cellulose fibers, and/or the like. For example, one or more filters <NUM> may be a commercially available filter under the trade name designations of VF1, VF2 and GFB, manufactured and distributed by Whatman, having a location in Maidstone Kent. For example, in some embodiments, one or more filters <NUM> may be a <NUM> disc of Whatman VF2 glass fiber.

The second side <NUM> of the filter <NUM> is in proximity to or contact the separation membrane <NUM>. Generally, the separation membrane <NUM> may remove the remaining red blood cells from the filtered blood providing filtered plasma. The filtered plasma may be essentially cell-free in that the filtered plasma includes minimal cellular debris. For example, hemoglobin content with the filtered plasma may be comparable to plasma obtained by centrifuge techniques currently known within the industry.

In some embodiments not part of the invention, the plasma separation system <NUM> may solely comprise the separation membrane <NUM> without the use of the filter <NUM>. For example, the blood may be filtered solely through the separation membrane <NUM> providing filtered plasma using the methods as described herein.

The separation membrane <NUM> may be formed of one or more layers <NUM>. Each layer <NUM> may have a first side <NUM> and a second side <NUM>. Generally, the filtered blood contacts the first side <NUM> of the separation membrane <NUM> passing through the layer <NUM> of the separation membrane such that filtered plasma emerges on the second side <NUM>.

The separation membrane <NUM> may be formed of materials including, but not limited to, nylon, polysulfone, polycarbonate and/or the like. For example, in some embodiments, the separation membrane <NUM> may be an ion-tracked etched membrane.

In some embodiments, the separation membrane <NUM> may be an asymmetric membrane. For example, the separation membrane <NUM> may be formed having at least a first set of pores and a second set of pores, with the first set of pores and the second set of pores being different sizes. Generally, larger pores may be formed on the first side <NUM> of the separation membrane <NUM> and smaller pores may be formed on the second side <NUM> of the separation membrane <NUM> such that the filtered blood flows through the larger pores to the small pores. This may reduce blockage of red blood cells within the separation membrane <NUM>.

In some embodiments, one or both of the filter <NUM> and/or separation membrane <NUM> may be treated with one or more blocking agents and/or surfactants. Treatment with blocking agents and/or surfactant may enhance analyte recovery and/or plasma separation efficiency. Surfactants may include, but are not limited to, Tween-<NUM>, and/or the like.

By treating with one or more blocking agents, the filter <NUM> and/or separation membrane <NUM> may be made more or less hydrophilic, more or less hydrophobic, more or less susceptible to protein adsorption, more or less positively charged, more or less negatively charged and/or the like. For example, in some embodiments, the filter <NUM> and/or separation membrane <NUM> may be treated with PAMAM dendrimers, Merquat, or other polycations. Blocking agents may include, but are not limited to bovine serum albumin, Seablock, gelatin, and/or the like.

The second surface <NUM> of the separation membrane <NUM> may contact the adhesive member <NUM>. In some embodiments, the adhesive member <NUM> may prevent leakage of blood cells around the perimeter of the separation membrane <NUM>.

The adhesive member <NUM> may include a first surface <NUM> and a second surface <NUM> with the first surface <NUM> contacting the second surface <NUM> of the separation membrane <NUM> and the second surface <NUM> adhered to at least a portion of the capillary surface <NUM> of the recess <NUM>. Generally, the adhesive member <NUM> aids in holding the separation membrane <NUM> within the recess <NUM>. To that end, in some embodiments, each surface <NUM> and <NUM> of the adhesive member <NUM> may include an adhesive material. Adhesive material may include, but is not limited to, polyethylene terephthalate (PET) with silicone adhesive, and/or the like. For example, the adhesive member <NUM> may be a double-sided PET adhesive O-ring as illustrated in <FIG> and <FIG>. In some embodiments, the adhesive member <NUM> may be integral to the blood separation well <NUM> or adhered to the wall <NUM> of the recess <NUM>.

Size and shape of the adhesive member <NUM> may be dependent on the size and shape of the separation membrane <NUM> such that the separation membrane <NUM> may be positioned within the blood separation well <NUM> and leakage of filtered blood about edges of the separation membrane <NUM> may be minimized or eliminated.

The size and shape of the adhesive member <NUM> may be determined such that the separation membrane <NUM> is placed in close contact with the capillary surface <NUM> of the recess <NUM> of the blood separation well <NUM> while preventing leakage of blood within the blood separation well <NUM>. In some embodiments, the shape of the adhesive member <NUM> may include an opening <NUM>. The opening <NUM> may provide for flow of the filtered plasma to flow from the second surface <NUM> of the separation membrane <NUM> to the capillary surface <NUM> of the recess <NUM>. For example, in some embodiments, the adhesive member <NUM> may be formed as an O-ring, or any fanciful shape providing a direct opening <NUM> for flow of filtered plasma from the second surface <NUM> of the separation membrane <NUM> to the capillary surface <NUM> of the recess <NUM>. Additionally, in some embodiments, the material of the adhesive member <NUM> may be formed of a mesh-type material.

Referring to <FIG> and <FIG>, the capillary surface <NUM> of the recess <NUM> of the blood separation well <NUM> includes one or more microchannels <NUM>. Microchannels <NUM> may encourage capillary flow of the filtered plasma. Microchannels <NUM> may form any pattern capable of enhancing capillary flow of the filtered plasma through the blood separation well <NUM>. For example, in <FIG> and <FIG>, eight microchannels <NUM> are used to form a concentric pattern having a plurality of radial microchannels connecting at a central axis <NUM>. Although eight microchannels <NUM> are illustrated in <FIG> and <FIG>, it should be apparent to one skilled in the art that any number of microchannels <NUM> may be used so long as such microchannels <NUM> are positioned within the confines of the capillary surface <NUM>. Additionally, one or more tributaries may be included within the concentric pattern. It should be noted that the capillary surface <NUM>, in some embodiments not part of the invention, may not include microchannels <NUM> as capillary flow may still occur without such microchannels <NUM>.

In some embodiments, one or more venting channels <NUM> may be positioned within the recess <NUM> of the blood separation well <NUM>. For example, in <FIG> and <FIG>, four venting channels <NUM> are provided within the recess <NUM> of the blood separation well <NUM> extending from the capillary surface <NUM> to the surface <NUM>. Venting channels <NUM> may provide for venting of gas (e.g., air) within the blood separation well <NUM>. For example, gas entering the filter <NUM> and/or separation membrane <NUM> may exit the blood separation well <NUM> through the one or more venting channels <NUM>.

The filtered plasma may flow from the blood separation well <NUM> to the channel <NUM>. The channel <NUM> may include a first outlet <NUM> connected to the plasma collection vessel <NUM>. In some embodiments, the channel <NUM> may also include a second outlet <NUM>. Generally, the second outlet <NUM> may be blocked during operation of the plasma separation system <NUM> to cause the vacuum force to be directed into the blood separation well via the channel <NUM>. The second outlet <NUM> may be capable of being selectively opened to provide for removal of any additional filtered blood and/or filtered plasma from within the blood separation well <NUM> and/or channel <NUM>.

The filtered plasma flows through the channel <NUM> and through the first outlet <NUM> to the plasma collection vessel <NUM>. The plasma collection vessel <NUM> may have a proximal end <NUM> and a distal end <NUM> connected by a tapered wall <NUM>. For example, the width of the plasma collection vessel <NUM> may increase from the proximal end <NUM> to the distal end <NUM>. Although the shape of the plasma collection vessel <NUM> is shown as conical, it should be apparent that the plasma collection vessel <NUM> may be formed in other shapes (e.g., cylindrical). Generally, the plasma collection vessel <NUM> may be formed such that the amount of volume where the filtered plasma collects reduces dead volume. For example, the shape of the plasma collection vessel <NUM> may be formed such that the filtered plasma collects in an area wherein recovery of the filtered plasma by a pipette or other collection means may be maximized (e.g., collection of <NUM>-<NUM>µL of filtered plasma).

Generally, the first outlet <NUM> may be positioned below the output port <NUM> and near the proximal end <NUM> of the plasma collection vessel <NUM>. Collection of filtered plasma may be in a portion <NUM> positioned below the first outlet <NUM>, e.g., between the first outlet <NUM> and the proximal end <NUM> of the plasma collection vessel <NUM> as illustrated in <FIG>.

The plasma collection vessel <NUM> includes a seal <NUM>. The seal <NUM> covers the distal end <NUM> of the plasma collection vessel <NUM> so that vacuum force applied to the output port <NUM> of the channel <NUM> is directed through the plasma collection vessel <NUM> and into the channel <NUM>. In some embodiments, the seal <NUM> is formed of pierceable material. For example, the seal <NUM> may be formed of materials capable of being pierced by a pipet tip or similar mechanism for collection of plasma from the plasma collection vessel <NUM>. Such materials may include, but are not limited to, acetate, polyethylene, foil and/or the like.

The plasma collection vessel <NUM> is connected to the channel <NUM>. The channel <NUM> is positioned above the channel <NUM>, and in proximity to the distal end <NUM> of the plasma collection vessel <NUM>, e.g., between the channel <NUM> and the distal end <NUM>. The channel <NUM> connects the plasma collection vessel <NUM> to the negative pressure source <NUM> via the outlet port <NUM>. In some embodiments, the outlet port <NUM> of the channel <NUM> may include a fitted tube or luer connection at which the negative pressure source <NUM> is connected.

The negative pressure source <NUM> may be any source capable of providing force between approximately <NUM>,<NUM>-<NUM>,<NUM> kPa (<NUM>-<NUM> psi). For example, the negative pressure source <NUM> may include, but is not limited to, a pump, a syringe pump, a vacuum pump, a suction pump, and/or the like. Generally, the negative pressure source <NUM> may be capable of being controlled such that plasma may be collected without hemolysis and/or free of cellular material from the blood. For example, by controlling the force of the negative pressure source <NUM> within the boundaries discussed above, the risks of damage to the plasma may be reduced such that the blood may not hemolyze and/or cells may not deform (i.e., pass through the separation membrane <NUM>).

Control of the negative pressure source <NUM> may provide for a fixed flow rate and volume. For example, in some embodiments, one or more pressure sensors or monitors may be used to control the negative pressure source <NUM>. In another example, displacement speed of a syringe may control the rate and volume of the negative pressure source <NUM>.

<FIG> illustrates another exemplary embodiment of a plasma separation system 10a. Similar to the plasma separator system <NUM> of <FIG>, the plasma separation system 10a includes the housing <NUM> supporting or encompassing the blood separation well <NUM>. The blood separation well <NUM> is connected via a first channel 16a to a plasma collection vessel 18a such that filtered plasma enters the plasma collection vessel 18a from a proximal end <NUM> of the plasma collection vessel 18a. A second channel <NUM> connects the plasma collection vessel 18a to an outlet port <NUM> downstream of the plasma collection vessel 18a. The outlet port <NUM> may allow for a negative pressure source <NUM> (e.g., vacuum source) to be attached to the plasma separator system 10a.

The channel 16a may be implemented in a variety of manners such that the channel 16a connects the plasma collection vessel 18a to the proximal end <NUM> of the plasma collection vessel 18a. For example, the channel 16a may include a variety of linear segments that are interconnected as shown in <FIG>. In the example depicted in <FIG>, the channel 16a includes a first portion <NUM> and a second portion <NUM> that may be in parallel alignment connected by a third portion <NUM> extending between the first portion <NUM> and the second portion <NUM>. In the example shown, the third portion <NUM> extends normally to the first portion <NUM> and the second portion <NUM> and vertically within the housing <NUM>. Although the channel 16a includes corners <NUM> and <NUM> formed by intersection of the first portion <NUM> and the second portion <NUM> with the third portion <NUM>, it should be noted, the corners <NUM> may include rounded edges. Additionally, the third portion <NUM> may be positioned at an angle relative to the first portion <NUM> and the second portion <NUM> such that the third portion <NUM> provides a sloped connection between the first portion <NUM> and the second portion <NUM>. A fourth portion <NUM> of the channel 16a may provide an inlet <NUM> into the proximal end <NUM> of the plasma collection vessel 18a.

Generally, in the plasma separation system 10a, blood is added to the blood separation well <NUM>. Vacuum pressure may be applied via the outlet port <NUM>. Using a combination of capillary action and vacuum pressure, plasma may be separated from the blood. The plasma may enter and collect at the proximal end <NUM> of the plasma collection vessel 18a. The vacuum pressure may be controlled to prevent hemolysis and/or leakage of cellular material.

<FIG> illustrates another exemplary embodiment of a plasma separation system 10b which is similar in construction to the plasma separation systems <NUM> and 10a shown in <FIG> and <FIG> with the exception that that plasma separation system 10b includes a plasma collection vessel 18b in the form of a serpentine channel <NUM>, rather than a well. Since the volume of the serpentine may be controlled by the length of the serpentine channel <NUM>, the system 10b can be used to meter the plasma. The serpentine channel <NUM> can be readily integrated with other standard microfluidic features such as valves and reaction wells for performing quantitative and qualitative assays. The number of curves, the configuration and/or the length of the serpentine channel <NUM> may be determined based on the assay of interest.

<FIG> and <FIG> illustrate an exemplary method for operating the plasma separator system <NUM> of <FIG>. In particular, <FIG> illustrates a flow chart <NUM> for operating the plasma separation system <NUM>.

To separate plasma from red and white blood cells in a blood sample, in a step <NUM>, the blood sample <NUM> (e.g., <NUM>µl) is added to the blood separation well <NUM> as illustrated in <FIG>. In some embodiments, a pipet, or other similar device, may be used to add the blood sample to the blood separation well <NUM>.

In a step <NUM>, the blood sample wicks into the filter <NUM> and the separation membrane <NUM>. In some embodiments, the blood sample wicks into the filter <NUM> and the separation membrane <NUM> for a pre-determined time period. For example, in one non-limiting example, the blood sample may be allowed to wick into the filter <NUM> and the separation membrane <NUM> for approximately <NUM> - <NUM> seconds.

In some embodiments, the blood sample may wick into the filter <NUM> and the separation membrane <NUM> by capillary action. In one non-limiting example, the presence of the one or more venting channels <NUM> (shown in <FIG>) may promote wicking of the blood sample by providing escape of gas from edges of the filter <NUM> and/or the separation membrane <NUM>. Further, the presence of microchannels <NUM> on the capillary surface <NUM> of the recess <NUM> may also promote capillary flow from the second side <NUM> of the separation membrane <NUM> into the channel <NUM> (shown in detail in <FIG>).

In a step <NUM>, the negative pressure source <NUM> (e.g., vacuum source) is actuated to apply the vacuum force to the blood separation well <NUM> via the outlet port <NUM>. In a step <NUM>, the vacuum force assists the blood sample to proceed through the filter <NUM> providing filtered blood. In a step <NUM>, the blood sample proceeds through the separation membrane <NUM> providing filtered plasma <NUM> as illustrated in <FIG>. It should be noted that prior to application of vacuum source to the blood separation well <NUM>, a portion of the blood sample may proceed through the filter <NUM> and the separation membrane <NUM>.

In a step <NUM>, the filtered plasma <NUM> may flow through the channel <NUM> and collect in the plasma collection vessel <NUM> as illustrated in <FIG>. The vacuum force may be maintained at a substantially constant level until all needed filtered plasma is collected in the plasma collection vessel <NUM> or may increase over time to a set-point.

In a step <NUM>, the vacuum force is caused to cease, such as by deactuating the negative pressure source <NUM> (e.g., vacuum source). In a step <NUM>, the filtered plasma can be removed from the plasma collection vessel <NUM> such as by piercing the seal <NUM> of the plasma collection vessel <NUM>. For example, a pipet tip may pierce the seal <NUM> and filtered plasma may be removed from the plasma collection vessel <NUM>. One or more assay may then be performed using the filtered plasma. Quantitative and/or qualitative assays may be performed using the filtered plasma. For example, as the pipet may be capable of collecting a determinate amount of filtered plasma, quantitative assays may be performed.

In one example, a blood sample of <NUM>µL and <NUM>% hematocrit (HCT) containing D-dimer at a concentration of <NUM> ng/ml may be added to the blood separation well <NUM> as illustrated in <FIG>. The filter <NUM> of the blood separation well <NUM> may be formed of VF2, and the separation membrane <NUM> may be formed of a polysulfone asymmetric membrane, for example. Using the process detailed in <FIG> and <FIG>, the blood sample may be allowed to wick into the filter <NUM> and separation membrane <NUM> for approximately <NUM>-<NUM> seconds. The negative pressure source <NUM> may be applied (e.g., between <NUM>,<NUM>-<NUM>,<NUM> kPa (<NUM>-<NUM> psi) such that filtered plasma (e.g., approximately <NUM>-<NUM>µL) may be collected. The D-dimer concentration in the filtered plasma may then be measured (e.g., using a Siemens Stratus CS D-dimer immunoassay). In one example, the D-dimer concentration recovery for the filtered plasma, as compared to centrifugation was <NUM>%.

In another example, a blood sample of <NUM>µL and <NUM>% HCT containing Tnl at a concentration of <NUM> pg/ml may be added to the blood separation well <NUM>. Using the process detailed in <FIG> and <FIG>, the blood sample may be allowed to wick into the filter <NUM> and separation membrane <NUM> for approximately <NUM>-<NUM> seconds. The negative pressure source <NUM> is applied (e.g., between <NUM>,<NUM>-<NUM>,<NUM> kPa (<NUM>-<NUM> psi) such that filter plasma (e.g., approximately <NUM>µL) may be collected. The Tnl concentration in the filtered plasma may then be measured (e.g., using Siemens Dimension EXL Tnl immunoassay). In one example, the Tnl concentration recovery for the filtered plasma, as compared to centrifugation was <NUM>%.

<FIG> illustrates another exemplary embodiment of a plasma separation system 10c. The plasma separation system 10c is similar to the plasma separation systems <NUM>-10b illustrated in <FIG>, <FIG> respectively; however, the plasma separation system 10c applies positive force via a positive pressure source <NUM> upstream of the plasma collection vessel <NUM>. In particular, the positive pressure source <NUM> is connected to a cap <NUM> positioned over the filter <NUM> within the blood separation well <NUM>.

In some embodiments, the cap <NUM> is form fit over the filter <NUM> within the blood separation well <NUM>. The cap <NUM> may include an outlet <NUM>. The outlet <NUM> may connect to the positive pressure source <NUM>. The positive pressure source <NUM> may provide between <NUM>,<NUM>-<NUM>,<NUM> kPa (<NUM>-<NUM> psi) of positive pressure to the blood separation well <NUM> during use. The positive pressure may force the blood sample through the filter <NUM>, separation membrane <NUM>, and channel <NUM> into the plasma collection vessel <NUM>.

In some embodiments, the positive pressure source <NUM> may be a syringe loaded with air. By forcing air through the outlet <NUM>, positive pressure may be applied to the blood sample forcing the blood sample through the filter <NUM>, separation membrane <NUM>, and/or channel <NUM> to the plasma collection vessel <NUM>. In this embodiment, the outlet <NUM> downstream of the plasma collection vessel <NUM> may be used as a vent and opened as needed.

Claim 1:
An apparatus, comprising:
a housing (<NUM>) supporting or encompassing a blood separation well (<NUM>) for collection of a blood sample (<NUM>), the blood separation well (<NUM>) having a recess (<NUM>) intersecting a surface (<NUM>) of the housing (<NUM>), wherein the surface of the recess is defined further as a capillary surface (<NUM>) having at least one microchannel;
a separation membrane (<NUM>) positioned on the surface of the recess (<NUM>) for filtration of the blood sample to provide filtered plasma (<NUM>);
a plasma collection vessel (<NUM>) configured to collect filtered plasma; a first channel (<NUM>) connecting the blood separation well (<NUM>) to the plasma collection vessel (<NUM>) proximate to the surface of the recess (<NUM>); and,
a second channel (<NUM>) connecting the plasma collection vessel (<NUM>) to a first outlet port (<NUM>), and a filter (<NUM>) positioned within the recess (<NUM>, the second side (<NUM>) of the filter (<NUM>) being in proximity to or in contact with the separation membrane (<NUM>), wherein the filter (<NUM>) is formed of one or more layers (<NUM>),
a seal (<NUM>) is positioned on a distal end (<NUM>) of the plasma collection vessel (<NUM>), wherein the seal (<NUM>) is formed of a pierceable material;
the second channel (<NUM>) is positioned above the first channel (<NUM>) and in proximity to the distal end (<NUM>) of the plasma collection vessel (<NUM>) such that, when a negative pressure force is applied to the first outlet port (<NUM>) of the second channel (<NUM>), filtered plasma (<NUM>) is drawn into the collection vessel (<NUM>), wherein the first channel (<NUM>), the plasma collection vessel (<NUM>), and the second channel (<NUM>) are configured to convey the negative pressure force applied to the first outlet port (<NUM>) to the surface of the recess (<NUM>),
characterized in that
an adhesive member is disposed within the recess and positioned on the capillary surface, with the separation membrane and the filter positioned thereon, respectively, wherein the filter (<NUM>), the separation membrane (<NUM>) and the adhesive member (<NUM>) are stacked on the capillary surface (<NUM>) and wherein the filter (<NUM>) and the separation membrane (<NUM>) are pressure fit within the recess (<NUM>).