Patent Description:
The separation of certain cells or cell population(s) from complex biological liquids such as blood is a routine procedure in many fields of medical research. Many different processes are available and are commonly used in the research community for isolating target cells.

Centrifuging anti-coagulated blood leads to the separation of red blood cells and white blood cells. The lighter white blood cells sediment slowly and form an opaque layer on top of the red blood cells. This layer is commonly referred to as the "buffy coat". Several washing steps are normally necessary to remove the red blood cells. This process was for many years the standard procedure for isolating lymphocytes from blood.

The introduction of a separation medium with the specific density of the white blood cells, i.e., a density separation medium, makes it possible to separate white blood cells from red blood cells by centrifugation of anti-coagulated whole blood through the density separation medium. The heavier red blood cells sediment through the density separation medium, while the white blood cells remain on top of the density separation medium. The white blood cell layer can then be carefully removed with a pipette. However, this results in additional stress to living cells. This remains the standard procedure in isolating white blood cells from blood specimens today.

<CIT> discloses an apparatus for filtering blood. The apparatus includes a filter vessel with a filter disposed at the open lower end, whereby the open lower end is inserted into a container having a closed lower end and a capped upper end. The capped neck of the filter vessel is formed with a capillary channel for pressure equalization. Among other things, this reference does not teach a cap for suspending filter vessel inside the container, whereby the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the filter vessel.

<CIT> discloses an isolation system for white blood cells from blood. The system includes a capped separation tube having open access that can be placed inside a centrifuge tube with a cap and closed bottom. During centrifugation, the small cell pellets sediment at the bottom of the centrifuge tube and the white blood cells remain inside the separation tube. Among other things, this reference does not teach that the separation tube is submersed in separation medium or that the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the separation tube.

<CIT> discloses a centrifuge tube assembly to extract blood components. An inner tube has a lower open end and an open upper end in the form of a threaded male luer that is closed by a cap. The inner tube extends through the cap of an outer containment tube. During centrifugation, light density components move towards the cap and the heaviest components collect at the bottom of the inner tube. Among other things, this reference does not teach that the inner tube is submersed in separation medium or that the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the inner tube.

<CIT> discloses a fluid collection device for separating blood into its phases. An outer container has an open top closed by a cap. A phase partitioning device containing a sealant is disposed within the outer tube. During centrifugation, the cellular phases separate and the sealant is dispensed from the phase partitioning device to form a semi rigid partition between the phases. Among other things, this reference does not teach an open-bottom inner containment unit suspended within the outer tube by attachment to the cap, whereby the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the inner containment unit.

<CIT> discloses a centrifuge vessel that includes a collector having an open end and a cap. The vessel contains a fraction-density-altering solution and a blood sample. During centrifugation, the blood sample separates into its respective portion of target material based on density and the target material such as circulating tumor cells and blood cells are moved from the vessel to the collector. Among other things, this reference does not teach a cap used to fix the collector inside the vessel or that the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the collector.

<CIT> discloses a filter assembly for the separation of blood. The assembly includes a centrifuge tube that includes a filter tube having a cap to seal the upper end and a filter at an opposite open end. Among other things, this reference does not teach that a lower end of the filter tube is submersed in separation medium or that the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the filter tube.

<CIT> discloses a centrifugally-driven assembly for separating blood product. The assembly includes a tubular receptacle and a closure for supporting a port having a second cap. Among other things, this reference does not teach that the second cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the system, namely the ports or pipes extending therefrom.

<CIT> discloses a rigid tube having a closure comprising a tubular seal plug that includes a septum and a central passage. The seal plug separates the blood samples into higher and lower density components. Among other things, this reference does not teach an open-bottom inner containment unit suspended within the outer tube by attachment to the tube closure, whereby the tube closure is designed with a means to regulate, or to facilitate the regulation of, pressure within the inner containment unit.

<CIT> discloses a device for separating blood components. The device includes a separator having an open bottom end and a bellow for closing the top end. The separator is disposed inside an outer container. Among other things, this reference does not teach an open-bottom inner containment unit suspended within the outer tube by attachment to the cap, whereby the cap is designed with a means to regulate, or to facilitate the regulation of, pressure within the inner containment unit.

<CIT> discloses a system for the separation of blood samples. The system includes an outer centrifuge tube containing a wash solution and an inner tube which is insertable into the outer tube. The system separates blood samples, based on the densities of the various components. The inner tube includes an air vent to allow free communication between a lower inner area and the atmosphere. Among other things, this reference does not teach a cap designed with a means to regulate, or to facilitate the regulation of, pressure within the inner tube.

<CIT> discloses a system for separating bolgoical material, used to separate a mixed solution, comprising a bottomed centrifuge tube, a separation tube having an open bottom, and a cap, wherein the centrifuge tube and the separation tube are configured to sealingly and releasably couple to the cap, wherein the cap comprises means for facilitating or regulating air- or gas-low between an area outside of the cap and an interior of the separation tube, when the separation tube and the centrifuge tube are coupled to the cap.

<CIT> teaches a system with a centrifugation tube, a separation medium disposable with the centrifuge tube and a separation member, wherein a volume of the separation medium is such that the open bottom of the separation member is submersed in the separation medium.

<CIT> teaches a system wherein the means for facilitating or regulating air- or gas-flow comprise one or more channels that pass through the top of the cap and into the cavity, wherein, when the cap is coupled to a container, the one or more channels provide open communication between the top of the cap and an interior of the container, wherein the plug is configured to releasably seal the one or more channels.

<CIT> teaches a system comprising a cap and centrifuge tube, wherein the means for regulating a flow of matter is selected from the group consisting of a syringe, a pump and an air compressor.

<CIT> discloses a system for separating biological material, the system comprising a centrifuge tube, a separation tube having an open bottom, a cap, wherein the centrifuge tube and the separation tube are configured to sealingly and releasably couple to the cap, wherein, when coupled to the cap, a predetermined length of the separation tube is positioned within the centrifuge tube, wherein an area between an exterior surface of the separation tube and an interior surface of the centrifuge tube is unobstructed along the pre-detremined length, wherein a top of the cap includes an aperture that opens into the cavity of the cap and wherein the cap comprises means for facilitating or regulating air- or gas-flow between an area outside of the cap and an interior of the separation tube.

Based on the foregoing, there is a need in the art for an apparatus and system that provides a means for isolating target material from non-target material, combined with a means for regulating air pressure within the system to allow for controlled release and analysis of the isolated target material, thus minimizing the number of processes/steps for isolation and collection and, hence, mitigating stress to the target material.

A system for separating biological material includes, at least, a separation apparatus, i.e., a centrifuge tube, a separation tube having an open bottom, and a cap, and a separation medium disposable within the separation apparatus. The centrifuge tube and the separation tube are configured to sealingly and releasably couple to the cap. The cap is configured to facilitate and/or regulate air- or gas-flow between an area outside of the cap and an interior of the separation tube. A volume of the separation medium is such that, when the separation tube and the centrifuge tube are coupled to the cap, the open bottom of the separation tube is submersed in the separation medium.

In an embodiment, an aperture extends through the top of the cap and opens into an interior portion, i.e., a cavity, of the cap, such that, when the cap is coupled to the separation tube, the cap provides open communication between an area outside of the cap and an interior of the separation tube. In a further embodiment, the cap includes one or more channels that extend through the top of the cap, such that, when the cap is coupled to the separation tube, the channels provide open communication between an area adjacent to the top of the cap and an interior of the separation tube.

In an embodiment in which an aperture and channels extend through the top of the cap, the cap includes a plug configured to releasably seal the aperture and the channels. The lower portion of the plug is insertable into the aperture and is configured to seal the aperture. In an embodiment which is not claimed, the upper portion of the plug includes a flange. In an embodiment which is not claimed, the flange is configured to transition between a closed position and an open position. In the closed position, the flange is configured to seal the channels, and in the open position, the flange is configured to unseal, i.e., open, the channels to provide open communication between the interior of the separation tube and an area outside of the cap. In an embodiment which is not claimed, the upper portion of the plug is a mechanical actuator configured to transition the flange between the open position and the closed position.

In an embodiment, the plug is releasably retained within the cap by an interference fit between the lower portion of the plug and an interior surface of the cap and/or the interior wall of the separation tube. In a further embodiment, a groove extends around the lower portion of the plug. The groove is configured to matingly engage a retaining lip protruding from the upper portion of the cavity, whereby the retaining lip seats into the groove to facilitate a snap fit between the plug and the cap. In addition to the interference fit, this serves to further assist in releasably coupling the plug to the cap.

The system also includes a hollow needle coupled to a means for regulating a flow of air, gas, or other matter. The needle is insertable through the cap or plug, and is used to facilitate the introduction of air, gas, or other matter into the separation tube. Example means for regulating a flow of air, gas, or other matter include, but are not limited to, a syringe, a pump, and an air compressor.

A method for separating and collecting a target material from a fluid sample using the separation system includes: depositing a density separation medium into the separation apparatus, such that a lower portion of the separation tube is submersed in the density separation medium; depositing the fluid sample on top of the density separation medium inside of the separation tube; sealing the cap, if necessary, e.g., if the cap requires a plug; centrifuging the fluid sample; uncoupling the centrifuge tube from the cap once centrifugation is complete, wherein the target material is retained within the separation tube; introducing air or gas into the separation tube to facilitate efflux of an amount of the target material from the separation tube's open bottom; and halting air- or gas-flow into the separation tube to halt efflux of target material from the separation tube' s open bottom. The process of introducing air or gas into the separation tube and halting air- or gas-flow into the separation tube can be repeated as necessary to collect a target material. The target material may be analyzed during effluxion and/or post-effluxion.

The foregoing, and other features and advantages of the invention, will be apparent from the following, more particular description of the preferred embodiments of the invention, the accompanying drawings, and the claims.

For a more complete understanding of the present invention, the objects and advantages thereof, reference is now made to the ensuing descriptions taken in connection with the accompanying drawings briefly described as follows.

Preferred embodiments of the present invention and their advantages may be understood by referring to <FIG>, wherein like reference numerals refer to like elements.

With reference to <FIG>, separation apparatus <NUM> includes centrifuge tube <NUM>, separation tube <NUM>, and cap <NUM> (with or without plug <NUM>), each of which are further individually illustrated in <FIG>.

With reference to <FIG>, centrifuge tube <NUM> has an open top that opens into cavity <NUM>. The bottom of centrifuge tube <NUM> is closed. Connector <NUM> is disposed at an upper portion of centrifuge tube <NUM>.

With reference to <FIG>, separation tube <NUM> has an open top and an open bottom, each of which open into hollow interior <NUM>. Connector <NUM> is disposed at an upper portion of separation tube <NUM>. With additional reference to <FIG>, the figures depict connectors <NUM>, <NUM> as threading that extends around an outer circumference of an upper portion of centrifuge tube <NUM> and separation tube <NUM>, respectively. However, one skilled in the art would understand and appreciate that alternative connection means could be employed to serve the same function.

With reference to <FIG>, cap <NUM> has an open bottom that opens into cavity <NUM>. Cavity <NUM> includes connectors <NUM>, <NUM> configured to matingly engage connectors <NUM>, <NUM>, respectively, to releasably couple centrifuge tube <NUM> and separation tube <NUM> to cap <NUM>. Aperture <NUM> extends through the top surface of cap <NUM> and opens into cavity <NUM>. Cavity neck <NUM> is disposed adjacent to, and is set inward from, connector <NUM>. In an embodiment, retaining lip <NUM> protrudes from a lower portion of cavity neck <NUM>. In an assembled state, wherein separation tube <NUM> is coupled to an un-plugged cap <NUM>, cap <NUM> provides open communication between an area outside of separation apparatus <NUM> and hollow interior <NUM>.

In an embodiment, one or more channels <NUM> extend through top surface of cap <NUM>. In an assembled state, wherein separation tube <NUM> is coupled to cap <NUM> (plugged or un-plugged), channels <NUM> provide open communication between the top of cap <NUM> and cavity <NUM>. In other words, channels <NUM> serve as a bypass for influent air to travel to cavity <NUM> when cavity <NUM> is otherwise sealed. For example, in an embodiment, as shown in <FIG>, ID and 7A, when separation tube <NUM> is coupled to cap <NUM>, the interior wall of separation tube <NUM> is separated from a rear surface of cavity neck <NUM> by open space <NUM>. Channels <NUM> pass through cap wall and open into open space <NUM>, allowing air to travel into and through open space <NUM> without passing through aperture <NUM>.

With reference to <FIG>, plug <NUM> includes flange <NUM> that extends from an outer circumference of plug <NUM>. The bottom surface of flange <NUM> is configured to complement the contour the top surface of cap <NUM>. For example, in a preferred embodiment, the bottom surface of flange <NUM> is disposed orthogonal to flange's central axis, allowing it to seat onto and create an air-tight seal with the top surface of cap <NUM>. Pull tab <NUM> extends from an outer edge of flange <NUM> and facilitates removal of plug <NUM> from cap <NUM>.

Lower portion of plug <NUM>, i.e., the portion of plug <NUM> below flange <NUM>, includes cavity <NUM> and is configured to be received into cavity <NUM>. Upon insertion, lower portion of plug <NUM> is removably retained within cavity <NUM> via an interference, i.e., friction, fit between lower portion of plug <NUM> and the wall surface of cavity neck <NUM> and/or the inner wall of separation tube <NUM> to create a sealed, i.e., air-tight, engagement between plug <NUM> and cavity neck <NUM> and/or separation tube <NUM>, respectively.

With reference to <FIG>, in an embodiment, groove <NUM> extends around an outer circumference of plug's lower portion to facilitate a snap fit engagement with cap's retaining lip <NUM> as plug <NUM> is inserted into cap <NUM>.

With further reference to <FIG>, in an embodiment, central portion <NUM> atop plug <NUM> serves as a mechanical actuator to transition flange <NUM> between an open position and a closed position. The circumference of central portion <NUM> is smaller than the circumference of cavity <NUM>, allowing downward displacement of central portion <NUM>. Depression of central portion <NUM> toward cavity <NUM> forces flange <NUM> to invert upward to the open position. Conversely, release of central portion <NUM> allows flange <NUM> to relax and return to its native configuration, i.e., the closed position.

With reference to <FIG>, in an embodiment, the circumference of central portion <NUM> is larger than the circumference of cavity <NUM>, thus preventing downward displacement of central portion <NUM>. By preventing downward displacement of central portion <NUM>, flange <NUM> is held in the closed position, i.e., it is unable to transition to the open position. This ensures that channels <NUM> remain sealed to prevent inadvertent spills or leakage during transport, etc..

In an embodiment, hollow needle <NUM> is used as a means to introduce air or gas into the upper portion of separation tube <NUM> through plug <NUM>. Needle <NUM> is connected to regulator <NUM>. Regulator <NUM> may be a syringe, a pump, or any other device/machine, e.g., an air compressor, configured to regulate a flow of air, gas, or other matter. This allows air or gas to be introduced into the upper portion of separation tube <NUM> in a regulated, i.e., controlled and calculable, manner.

Plug <NUM> is constructed of a flexible and/or compressible material with resilient qualities, e.g., plastic, rubber, or silicone. This facilitates the interference fit between lower portion of plug <NUM> and a wall surface of cavity neck <NUM> and/or the interior wall of separation tube <NUM> by allowing the lower portion of plug <NUM> to undergo a pre-determined degree of deformation, e.g., compression, during engagement/disengagement with cavity neck <NUM>. Further, it facilitates the process of transitioning flange <NUM> between its open and closed positions. Additionally, it facilitates penetration of needle <NUM> through top of plug <NUM> and into cavity <NUM> for introducing air or gas into separation tube <NUM>.

In an embodiment, a valve or port (not shown) is integrated into plug <NUM>. The valve or port provides an alternative means, whereby needle <NUM>, coupled to regulator <NUM>, may be punctured through valve or port, to inject matter, e. g , air, gas, or biological material, into separation tube <NUM>.

In an embodiment, cap <NUM> is configured without aperture <NUM>, such that the top of cap <NUM> is closed, i. , sealed, and, therefore, cap <NUM> does not require plug <NUM> to ensure an air tight seal. In such an embodiment, the interior of separation tube <NUM> is accessed through cap <NUM> using one or more valve means. For example, needle <NUM>, coupled to regulator <NUM>, may be punctured through cap <NUM>, or other valve or port (not shown) integrated into cap <NUM>, to inject matter, e. g , air, gas, or biological material, into separation tube <NUM>. At least a portion of cap <NUM> is preferably constructed of a flexible and/or compressible material with resilient qualities, e.g., plastic, rubber, or silicone, to facilitate penetration of needle <NUM> through cap <NUM> and into cavity <NUM> for introducing matter into separation tube <NUM>.

Referring again to <FIG>, assembly of separation apparatus <NUM> begins with coupling separation tube <NUM> to cap <NUM> via engaging connector <NUM> with connector <NUM>. Once fully engaged, upper edge of separation tube <NUM> abuts cap ledge <NUM> to provide a sealed, i.e., air tight, engagement between cap <NUM> and separation tube <NUM>. Next, separation tube <NUM> is received into centrifuge tube <NUM>, and centrifuge tube <NUM> is coupled to cap <NUM> via engaging connector <NUM> with connector <NUM>. Once fully engaged, upper edge of centrifuge tube <NUM> abuts cap ledge <NUM> to provide a sealed, i.e., air-tight, engagement between cap <NUM> and centrifuge tube <NUM>. Next, lower portion of plug <NUM> is removably inserted into cavity <NUM>. Once fully inserted, lower portion of plug <NUM> seats into cavity <NUM>, and bottom edge of flange <NUM> abuts a top surface of cap <NUM> to provide a sealed, i.e., air-tight, engagement between cap <NUM> and plug <NUM>. The sealed engagement between cap <NUM> and plug <NUM>, and cap <NUM> and separation tube <NUM>, coupled with a means for introducing air or gas into separation tube <NUM> in a controlled manner, e.g., channels <NUM> and flange <NUM>, or needle <NUM> and regulator <NUM>, provides a means for controlled release of liquid from the open bottom of separation tube <NUM>.

With further reference to <FIG>, density separation medium <NUM> is used within separation apparatus <NUM> to isolate target biological material. Density of density separation medium <NUM> depends on densities of the material to be separated. For example, to separate target white blood cells from red blood cells, the specific density of density separation medium <NUM> should be higher than the target white blood cell population. This will allow the higher density red blood cells to sediment through the open bottom of separation tube <NUM> to the bottom of the centrifuge tube <NUM>. Meanwhile, the lower density white blood cells will remain on top of density separation medium <NUM> inside of separation tube <NUM>.

<FIG> outlines a method of using the separation system, according to an embodiment of the present invention. At step <NUM>, a pre-determined volume of density separation medium <NUM> is deposited into separation apparatus <NUM>. This can be accomplished, for example, by depositing density separation medium <NUM> into centrifuge tube <NUM>, followed by inserting separation tube <NUM>, coupled to cap <NUM>, into centrifuge tube <NUM> and coupling centrifuge tube <NUM> to cap <NUM>, whereby density separation medium <NUM> passes into separation tube <NUM> through open bottom of separation tube <NUM>. As another example, separation tube <NUM>, coupled to cap <NUM>, can be inserted into centrifuge tube <NUM>, followed by coupling centrifuge tube <NUM> to cap <NUM> and depositing density separation medium <NUM> into separation tube <NUM> through cavity <NUM>, whereby density separation medium <NUM> flows into centrifuge tube <NUM> through the open bottom of separation tube <NUM>. The amount of density separation medium <NUM> deposited into separation apparatus <NUM> is such that, a lower portion of separation tube <NUM> is submersed in, and filled with, density separation medium <NUM>.

At step <NUM>, a sample containing biological material, is deposited into separation tube <NUM> on top of density separation medium <NUM>. This could be accomplished, for example, by puncturing needle <NUM> through cap <NUM> and dispensing the sample through needle <NUM> into separation tube <NUM> or, alternatively, by depositing the sample into separation tube <NUM> through cavity <NUM> via aperture <NUM>. At step <NUM>, in an embodiment in which aperture <NUM> extends through a top surface of cap <NUM>, plug <NUM> is engaged with cap <NUM> to ensure a complete air-tight seal of separation apparatus <NUM> in preparation for centrifugation. At step <NUM>, separation apparatus <NUM> containing density separation medium <NUM> and the sample is placed in a centrifuge and the sample is centrifuged.

With reference to <FIG>, following centrifugation, at step <NUM>, centrifuge tube <NUM> is removed from cap <NUM>, and separation tube <NUM> remains connected to cap <NUM>. The sealed, i.e., air-tight engagement between cap <NUM> and plug <NUM>, and cap <NUM> and separation tube <NUM> prevents liquid from flowing out of the lower end of separation tube <NUM>. By sealing the top of separation tube <NUM>, air is prevented from entering and exerting a downward force on liquid inside separation tube <NUM>, leaving only the upward force of atmospheric pressure being exerted on the liquid at the open bottom. As such, the pressure inside separation tube <NUM> is less than the external atmospheric pressure. Further, the upward force of atmospheric pressure is greater than the force of gravity pulling down on the liquid. Thus, the liquid is held inside separation tube <NUM>.

At step <NUM>, air or gas is introduced into the upper portion of separation tube <NUM>. This increases the pressure inside separation tube <NUM>, such that it is equal to or greater than the external atmospheric pressure. Air or gas can be introduced into separation tube <NUM> in a number of ways. For example, in an embodiment, pressure is applied to plug <NUM> to transition flange <NUM> to its open position, allowing an in-flow of air through channels <NUM> and into separation tube <NUM>. The degree of inversion is dependent on the amount of pressure applied to plug <NUM>. As such, the volume of influent air can be controlled by adjusting the amount of pressure exerted on plug <NUM>. As air enters the top of separation tube <NUM>, the upward and downward forces of air pressure on the liquid equalizes, leaving gravity as the dominant force causing the liquid to drop out of the bottom of separation tube <NUM>. As air is introduced into separation tube <NUM>, a corresponding volume of liquid is expelled through the lower end of separation tube <NUM>. Therefore, the release of liquid from separation tube <NUM> can be controlled by adjusting the amount of pressure exerted on cap <NUM>.

In another embodiment, needle <NUM> is punctured into and through plug <NUM> and air or gas is introduced into the upper portion of separation tube <NUM> through needle <NUM>. Needle <NUM> is connected to regulator <NUM>, such that the air or gas can be introduced into the upper portion of separation tube <NUM> in a regulated manner. The regulated influx of air or gas defines the release speed of the fluid from the open bottom of separation tube <NUM>. As air or gas is introduced into separation tube <NUM>, a corresponding volume of liquid is expelled through the lower end of separation tube <NUM>. Therefore, the release of liquid from separation tube <NUM> can be controlled by adjusting the amount of air or gas injected into separation tube <NUM>.

At step <NUM>, air- or gas-flow into separation tube <NUM> is halted. This is accomplished, for example, by releasing pressure from plug <NUM>, allowing flange <NUM> to revert to its native configuration, i.e., the closed position, thereby sealing channels <NUM> and preventing additional air from flowing into separation tube <NUM>. Similarly, air or gas-flow through needle <NUM> may be shut off. By halting air- or gas-flow into separation tube <NUM>, the pressure inside separation tube <NUM> drops below the external atmospheric pressure, preventing release of any remaining fluid inside from separation tube <NUM> until air- or gas-flow resumes.

Optionally, at step <NUM>, the efflux from separation tube <NUM> is analyzed, based on characteristics including, but not limited to, light absorption, emission, refraction, reflection and diffraction.

While the foregoing method is described and shown in a numerical, step-wise order, it should be understood that the steps are not limited to any specific order. Additionally, some steps may overlap in time, i.e., they may be carried out simultaneously, with other steps.

Claim 1:
Apparatus (<NUM>) for separating biological material, the apparatus comprising:
a. a centrifuge tube (<NUM>);
b. a separation tube (<NUM>) having an open bottom;
c. a cap (<NUM>), wherein the centrifuge tube (<NUM>) and the separation tube (<NUM>) are configured to sealingly and releasably couple to the cap (<NUM>), wherein, when coupled to the cap (<NUM>), a predetermined length of the separation tube (<NUM>) is positioned within the centrifuge tube (<NUM>), wherein an area between an exterior surface of the separation tube (<NUM>) and an interior surface of the centrifuge tube (<NUM>) is unobstructed along the pre-determined length, wherein a top of the cap (<NUM>) includes an aperture (<NUM>) that opens into a cavity (<NUM>) of the cap (<NUM>) and wherein the cap (<NUM>) comprises means for facilitating or regulating air- or gas- flow between an area outside of the cap (<NUM>) and an interior of the separation tube (<NUM>), said means for facilitating or regulation air- or gas-flow comprise one or more channels (<NUM>) that pass through the top of the cap (<NUM>) and into the cavity (<NUM>), wherein, when the cap (<NUM>) is coupled to the separation tube (<NUM>), the one or more channels (<NUM>) provide open communication between the top of the cap (<NUM>) and an interior of the separation tube (<NUM>), and further comprises a plug (<NUM>) that is configured to releasably seal the aperture (<NUM>) and one or more channels (<NUM>),
characterized by that
the lower portion of the plug (<NUM>) is insertable into the aperture (<NUM>) and is configured to seal the aperture (<NUM>), and that the apparatus (<NUM>) is comprising a hollow needle (<NUM>) coupled to a means for regulating a flow of air or gas, wherein the needle (<NUM>) is insertable into the cap (<NUM>) or plug and is used to facilitate the introduction of air or gas into the separation tube (<NUM>).