EA042126B1 - METHOD AND DEVICE FOR SAMPLE VOLUME REDUCTION - Google Patents

Description

Technical field

The present invention relates to a method and apparatus for sample volume reduction.

State of the art

It is known that biological samples must be processed in different ways in order to obtain the isolation of certain types of microparticles (usually cells).

Examples in this regard are the devices and methods described in patent applications PCT/IB2010/000615, PCT/IB2010/000580 (regarding the DEP Array TM system).

Typically, at the end of the above treatments, samples are obtained in which the microparticles are introduced into the liquid component. In connection with the above, it should be noted that the liquid component is usually a buffer that cannot be used in subsequent phases of the analysis, and that the sample volume is usually too large. For example, samples obtained after using the DEP Array TM system have volumes of approximately 13 μl, while subsequent phases (such as WGA, Whole Genome Amplification - amplification of the whole genome) require volumes of several microliters, in particular less than 5 μl, more moreover, less than 1.5 μl.

It is therefore necessary that samples be processed by centrifugation at high speed and that the operator manually collect excess fluid with great care and attention using a pipette (and tilting the control tube containing the sample). This process entails many problems. These include the following:

the success of operations depends to a large extent on the ability of the operator, who must be properly trained and periodically practiced;

there is a risk, which can be high if the operator does not work correctly, of removing the microparticle along with excess fluid;

the success rate of the procedure is not reliable and always reproducible, and also depends on the type of buffer used;

operations are relatively slow (it takes approximately 3 hours to isolate 96 samples);

the procedure requires special care, such as the use of specially provided pipettes and dual filter tips without contamination to reduce the risk of contamination of the sample during processing by the operator;

there is a relatively high risk of damage to the microparticle/microparticles due to centrifugation, which, as said, is carried out at relatively high speeds (hence giving a relatively high load on the microparticle/microparticles); and this procedure is not recommended for in vitro diagnostic (IVD) applications.

Similar problems arise when it is necessary to prepare biological samples prior to the above treatments in order to obtain isolation of certain types of microparticles (usually cells). In these cases, the initial and final volumes are larger (typically about 200 and 12 cl, respectively), but the disadvantages of known methods (high-speed centrifugation and subsequent manual collection by the operator) are those described above, with the addition of the fact that often for to obtain the desired volume, some work has to be done repeatedly.

In addition, almost identical problems occur in other cases where the sample volume is small (such as cell staining, cell washing, buffer change, cell fixation, cell permeabilization, and a combination of these). These problems are exacerbated even if the number of microparticles (cells) is small.

More generally, methods have not yet been proposed that are satisfactory, sufficiently accurate and/or reproducible to reduce the volume of a sample with small dimensions.

It is an object of the present invention to provide a method and apparatus for sample reduction that overcomes, at least in part, the disadvantages of the prior art and, if possible, is both simple and inexpensive to manufacture.

The essence of the invention

According to the present invention, there is provided a method and apparatus for reducing sample volume as claimed in the following independent claims and preferably in any of the claims that depend directly or indirectly on the independent claims.

Unless explicitly stated otherwise, in this text the following terms have the meanings given below.

By equivalent section diameter we mean the diameter of a circle having the same area as the section.

By microfluidic system we mean a system containing at least one

- 1 042126 microfluidic channel and/or at least one microfluidic chamber. Advantageously, but not necessarily, the microfluidic system comprises at least one pump (more specifically, a plurality of pumps), at least one valve (more specifically, a plurality of valves), and, if necessary, at least one gasket (more specifically, a plurality of gaskets) .

In particular, by microfluidic channel we mean a channel having a cross section with an equivalent diameter of less than 0.5 mm.

In particular, the microfluidic chamber has a height of less than 0.5 mm. More specifically, the microfluidic chamber has a width and length greater than the height (more specifically, at least five times the height).

In the present text, by microparticle we mean the largest cell size, which is less than 500 μm (preferably less than 150 μm). According to some non-limiting examples, the microparticle is selected from cells, cellular debris (in particular, cellular fragments, such as DNA and/or RNA), cellular aggregates (such as, for example, small aggregations of cells derived from stem cells, such as neurospheres or mammospheres), bacteria, lipo-beads, microballoons (in polystyrene and/or magnets), nanoballoons (for example, nanoballoons up to 100 nm in size), complexes formed from microballoons (in particular, magnetic ones; in particular, with the largest size less than 500 μm), bound with cells, circulating tumor cells associated with ferrofluid, exosomes, colloidal suspension (eg ferrofluid), liposomes, nuclei, spores, as well as combinations of them. Preferably, but not necessarily, the microparticles are cells.

In accordance with some non-limiting embodiments, the largest size of microparticles (preferably cells and/or cell debris) is less than 60 μm.

The sizes of the microparticles can be measured in a standard manner using graduated scale microscopes or conventional microscopes used with glass slides (on which microparticles are deposited) having a graduated scale.

In the present text, by the dimensions of a microparticle, we mean the length, width, and thickness of the microparticle.

Brief description of the figures

The invention will now be described with reference to the accompanying drawings, which illustrate several non-limiting embodiments of the invention, in which FIG. 1 and 2 are schematic and perspective views of an apparatus in accordance with the present invention in successive operating phases;

fig. 3 is a side section through a portion of the device of FIG. 1;

fig. 4 is a front view of the device of FIG. 1;

fig. 5 is a schematic front view, with some details shown transparently, of a further embodiment of an apparatus in accordance with the present invention;

fig. 6 is a perspective view of a detail of the device of FIG. 5;

fig. 7 is a cross section of the detail of FIG. 6;

fig. 8 is a perspective view of the detail of FIG. 6 with some components removed for clarity;

fig. 9 is a schematic side section of a detail from a further embodiment of an apparatus in accordance with the present invention;

fig. 10 is a schematic detail view of a further embodiment of an apparatus in accordance with the present invention;

fig. 11 is a schematic detail view of a further embodiment of an apparatus in accordance with the present invention;

fig. 12 is a schematic side section of a device component of one of the preceding figures;

fig. 13-16 illustrate schematically and in section a further embodiment of the component of FIG. 12 in successive working phases;

fig. 17 and 18 illustrate schematically and in section a further embodiment of the component of FIG. 12 in successive working phases;

fig. 19 illustrates a detail of the component of FIG. 17 on an enlarged scale;

fig. 20-22 illustrate sections from alternative embodiments of containers used in accordance with the present invention;

fig. 23, 24, 26 and 27 schematically illustrate some of the forces present during the implementation of the present invention, the x-axis shows the distance from the rotation axis, the y-axis shows the forces;

fig. 25 schematically illustrates some of the forces present during the implementation of the present invention, the x-axis shows the distance relative to the end of the container, the y-axis shows the forces;

fig. 28 and 29 schematically illustrate some of the forces present during implementation.

- 2 042126 of the present invention;

fig. 30 is a graphical representation of the experimental results obtained using the prior art method; the x-axis shows the identification of the operator who performed the tests; the y-axis shows the volumes (in μl) of the received samples;

fig. 31 is a graphical representation of the experimental results obtained using the prior art method; the x-axis shows the volumes (in μl) of the received samples; the y-axis indicates the repeatability (number of times) with which the indicated volume was obtained;

fig. 32 is a graphical representation of the experimental results obtained by testing the method of the present invention with a structure similar to that shown in FIG. 27; the Y-axis shows the received volume (in rl); the x-axis shows the angular velocity used for the experiment (rpm); and in FIG. 33 is a block diagram relating to possible uses of the method according to the present invention.

Detailed description

In FIG. 1, the number 1 denotes the entire sample volume reduction device 2 (FIGS. 9, 10 and 23-29) containing at least one (at least partially) liquid component 3.

The device 1 (in particular, Fig. 1-11) contains a processing unit 4, equipped with at least one socket 5 for supporting (at least partially), at least one container 6 (Fig. 7, 12 and 20-22 ) having an interior, a closed end 7, an end 8 provided with an opening 8' which establishes contact between the interior and exterior, and (at least) one side wall 9 (which extends between ends 7 and 8). In particular, the interior space is delimited (at least partially) by the side wall 9 and the closed end 7.

In particular, the container 6 can be inserted into and removed from the processing unit 4 (more precisely, can be inserted into the socket 5 and removed from it). More specifically, the device 1 contains a container 6.

The processing unit 4 contains a collecting device 10 (in particular, Fig. 3, 4, 6, 7 and 12-19), which is adapted to collect the first part of the specified (at least partially) liquid component 3, and is located outside the socket 5 and close (in the zone) of the indicated nest 5.

The processing unit 4 is adapted to move the socket 5 in such a way as to subject it to an acceleration (having at least one component) oriented by the collecting device 10 towards the socket 5, in particular in such a way that the first part of said (at least partially ) of the liquid component 3 flows out of the container 6 (passing through the opening 8') and reaches the collecting device 10, while the second part (at least partially) of the liquid component 3 (in particular, substantially with a certain volume) remains in the container 6, in particularly at the closed end 7.

It should be noted that acceleration is a vector quantity (vector), therefore it has a module (intensity - the absolute value of the vector) and direction. In particular, it should be noted that acceleration (as opposed to deceleration) is understood to be positive (hence with a positive modulus) and therefore entails an increase in speed (in its own direction).

More specifically, the acceleration used determines at least one first force of inertia on the first part of the sample 2 and at least one second force of inertia on the second part of the sample 2. The first and second forces of inertia are oriented from the first closed end 7 towards the second end 8 in transversely (in particular perpendicularly) to said opening 8'. The processing unit 4 is adapted to adjust the acceleration so that the first inertial force is greater than the first holding force acting between the first sample part and the container 6, and the second inertial force is less than the second holding force acting between the second sample part and the container 6 .

In accordance with some non-limiting embodiments (see, in particular, Fig. 9 and 25), the processing unit 4 includes a moving device 11, which is adapted to force the container 6 to make a substantially linear movement, accelerated in a given direction, so that the seat 5 facing forward with respect to the given direction and collecting device 10 facing backward with respect to the given direction (or rather, the closed end 7 faces forward and the end 8 faces back). In other words, the collecting device 10 is positioned below the socket 5 in the designated predetermined direction.

In accordance with some non-limiting embodiments (see, in particular, Fig. 1-11, 23, 24, 26 and 27), the processing unit 4 includes a moving device 11, which is adapted to rotate the socket 5 about the axis A of rotation in such a way that centrifugal force moves said first portion of said (at least partially) liquid component 3 (from container 6) into collecting device 10.

In some cases (socket 5 is shaped so that) the axis of rotation A passes through the container 6 between the closed end 7 and the end 8 (in particular, Fig. 7, 11, 24 and 27).

Alternatively (socket 5 is shaped so that) the closed end 7 is located between

- 3 042126 axis of rotation A and end 8 (in particular, Fig. 10, 23 and 26).

In some non-limiting cases, the collection device 10 includes a collection zone in which (at least a fraction) of the first part (at least partially) of the liquid component 3 is collected (for any additional subsequent applications). In these cases, the collection device 10 may be, for example, a control tube.

Preferably, but not necessarily (see, in particular, Fig. 12-19) collecting device 10 contains a holding system 12 for holding (at least a fraction) of the first part (at least partially) of the liquid component 3.

Thus, the risk of leakage of a portion of the first part (at least partially) of the liquid component 3 back into the container 6 is eliminated.

In particular, the used collecting device 10 (more specifically the holding system 12) faces the opening 8'.

In accordance with some non-limiting embodiments, the containment system 12 comprises an element selected from the group consisting of an absorbent material 13 (FIG. 12), a capillary trap (FIGS. 17-19), a liquid trap (FIGS. 13-16) (and combinations of them).

The capillary trap (FIGS. 17-19) comprises a plurality of grooves 14 having a width of less than 2.0 mm, in particular less than 0.9 mm (more specifically, greater than 0.5 mm). In this case, the liquid enters the grooves 14 due to the force (in particular centrifugal) to which it is subjected while the container 6 is accelerating, and remains in said grooves 14 due to capillarity (because the surface tension force is greater than the force of gravity). In particular, each groove 14 has a depth of at least 2 mm (typically up to 7 mm).

The liquid trap comprises a collecting chamber 15 (FIGS. 13-16) provided with an inlet 16, at least one movable wall 17 (in this case two movable walls 17) moving between a closed position (FIGS. 13 and 16) in which it prevents (liquid from and inlet from collection chamber 14) fluid from passing through inlet 16, and the opening position (FIGS. 14 and 15) in which fluid can pass through the inlet. The wall 17 is adapted to move to the opening position when the specified acceleration used is applied. In particular, when no acceleration is applied to wall 17 (more generally, when no force is applied to wall 17), wall 17 is in the closed position. More specifically, when acceleration is no longer applied to seat 5, wall 17 returns to the closed position. In particular, the collecting device 10 used faces the opening 8'.

More specifically, the acceleration used brings the movable wall 17 into the opening position. Even more precisely, the acceleration used brings the movable wall 17 into the opening position, acting directly and/or indirectly on the movable wall. In particular, in some cases, acceleration acts on the at least partially liquid component 3, which pushes the movable wall 17.

According to some non-limiting examples, the wall 17 is movable in the sense that it is (resilient) and/or deformable (elastically) hinged.

In accordance with some non-limiting embodiments, the absorbent material 13 is absorbent paper (FIG. 12).

Preferably, but not necessarily, the processing unit 4 comprises a plurality of sockets 5 (in particular, Figs. 8-10), each of which is adapted to receive a respective container 6. In particular, in these cases, the processing unit 4 comprises a moving device 11, which is adapted to force all sockets 5 make the same movement.

In accordance with some non-limiting embodiments (see, for example, Fig. 8), the seats 5 are arranged according to (at least) one row, essentially parallel to the axis A of rotation.

In some cases, the nests 5 are arranged in accordance with a plurality of rows substantially parallel to the axis of rotation A. In these cases, Fig. 10 shows the layer of the moving device 11 which is repeated several times. In other words, each slot 5 (from FIG. 10) is a slot belonging to a row of slots 5 (each of which is located together with other slots 5 of other rows around axis A).

One or more flexible well plates can be mounted directly on the transfer device 11 (more precisely on the rotor of the transfer device 11), in particular (curved) around axis A.

It should be noted that, in accordance with some preferred but non-limiting embodiments, the sample contains at least one microparticle 18 (in particular, a plurality of microparticles 18).

With specific reference to FIG. 11, in accordance with some preferred but non-limiting embodiments, the processing unit 4 comprises a plurality of additional peripheral pockets 19 which are disposed around the axis of rotation A and adapted to receive additional containers 6 containing (at least partially) the liquid component 3 and to be rotated by a transfer device. 11 about the axis A of rotation so that the centrifugal force is applied to the (at least partially) liquid component 3 contained in the additional containers 6 towards the closed end 7 of the other containers 6.

Thus, the moving device 11 in one movement (rotation around axis A) can simultaneously reduce the volume of the sample 2 contained in the container 6 located in the slot 5, and prepare the samples 2 contained in other containers 6 located in the peripheral slots 19 (by moving microparticles 18 in closed ends 7).

In particular, additional peripheral seats 19 are arranged around the seat 5, which, in particular, is located (in a central position) on the axis A (more precisely, so that the axis A passes through the container 6 located in the seat 5).

Preferably, but not necessarily, the processing unit 4 comprises a moving device 20 (particularly with a cam drive mechanism shown only partially and schematically) which is adapted to move the peripheral seat 8 (in practice becoming seat 5) from its peripheral position to a position around which other peripheral sockets 8 are located, and vice versa. In FIG. 11 this movement is shown by the arrow T.

In accordance with some non-limiting embodiments, the moving device 20 is adapted to move a row (substantially parallel to axis A) of peripheral sockets 19. In these cases, what is shown in FIG. 11 is a layer of construction which provides for a plurality of rows of peripheral sockets 19 (obtained, for example, in the same support 21).

It should be noted that the seat 5 (in particular, the support 21) and the collecting device 10 together constitute a cartridge 22, which can be removed from and inserted into the processing unit 4 (in particular, inserted into and removed from the transfer device 11).

Preferably, but not necessarily, the processing unit 4 (in particular the transfer device 11) contains a sensor (of a type known per se and not shown) to detect the presence (and/or correct position) of the cartridge 22 (in the transfer device 11).

Preferably, but not necessarily, the cartridge 22 comprises a writable memory (of a type known per se and not shown, e.g. not shown) writable memory. In accordance with some non-limiting embodiments, information may be written to writable memory regarding cartridge 22, such as an identification of cartridge 22, parameters for using the cartridge, and/or the number of times cartridge 22 has been used. In particular, the used reader and/or writer writes to the above memory when the corresponding cartridge 22 is used for the first time, and whenever said cartridge 22 is installed in the moving device 11, it determines that it is not the first use and emits an error signal. This eliminates the repeated use of cartridge 22 and contamination of samples 2.

Preferably, but not necessarily, the processing unit 4 comprises a support 21 on which there are a plurality of seats 5. In these cases, the cartridge 22 comprises a support 21 and a collecting device 10.

With specific reference to FIG. 8, according to some non-limiting embodiments, the support 21 has a plurality of seats 5 arranged in a row.

In particular (FIGS. 6 and 7), the collecting device 10 is located above the nests 5, more specifically to cover the openings 8' of the containers 6. More specifically, the collecting device 10 is located above the support 21.

In accordance with some non-limiting embodiments, which are not shown, the processing unit 4 includes a magnet (permanent magnet and/or electromagnet) located in the socket 5.

In some cases, the magnet is located at one end of the socket 5 opposite the collecting device 10. In particular, the magnet is located at the closed end 7. In some cases, the magnet is adapted to reduce the risk of leakage of the microparticle(s) 18 (which contains/which contain at least one magnetic functionalization ) from the container 6, while the container 6 in use is accelerated by the processing unit 4 (in particular by the transfer device 11).

Additionally or alternatively, the processing unit 4 comprises an (additional) magnet (permanent magnet and/or electromagnet) located in a socket 5 located in the collecting device 10, in particular at one end of the socket 5 located in the collecting device 10. More specifically, the magnet is located at the end 8, more specifically at the hole 8'. In these cases, the magnet improves the removal of unwanted (magnetic) components from sample 2.

In accordance with some non-limiting embodiments, the processing unit 4 also includes a supply device F (for example, shown in Fig. 11), which is adapted to supply the container 6 with a substance (in particular, a liquid), such as, for example, a reagent and/or a buffer solution . Feeder F is particularly useful when additional treatments (beyond simple volume reduction) are desired in Sample 2 (eg, staining and/or permeabilization and/or washing and/or fixation of the microparticle(s) 18).

With reference to FIG. 5, in particular, the moving device 11 comprises a motor 23 and an actuator 24 (linear or rotating). In the embodiments of FIG. 1-8, the actuator 24 is a rotor and has a housing H for the cartridge 22.

Preferably, but not necessarily, the movement device 11 also includes a brake 25 which is adapted to block movement of the seat 5 (in particular, the actuator 24).

In particular, the processing unit 4 also includes a control unit 26 (FIG. 5) which is adapted to control the motor 23 (and optionally the brake 25). More specifically, the control unit 26 is adapted to control the operation of the motor (and optionally the brake 25) based on the resulting final volume of sample 2 (and/or the characteristics of the control tubes and/or liquid).

Preferably, but not necessarily, the control unit 26 is connected to the above sensor to detect the presence (and/or correct position) of the cartridge 22 and is adapted to be processed by the motor 23 (and optionally the brake 25) based on the data detected by the sensor. More precisely, if the sensor does not detect the presence of the cartridge 22 or it detects an incorrect location of the cartridge 22, then the motor 23 is not driven (by the control unit 26). Preferably, but not necessarily, the control unit 26 also controls the operation of the power supply F.

Preferably, but not necessarily, the control unit 26 is connected to the aforementioned reader and/or writer and adapted to be processed by the motor 23 (and optionally the brake 25) based on the data detected by the reader and/or writer. More specifically, if the reader and/or writer detects that the cartridge 22 is not being used for the first time, then the control unit 26 does not drive the motor 23.

According to some non-limiting embodiments, the processing unit 4 comprises an operator interface (HMI, Human Machine Interface) 27 provided with, for example, a touch screen and/or mechanical buttons.

Preferably, but not necessarily, the processing unit 4 also includes a cover 28 that moves between an open position and a closed position.

When the cover 28 is in the open position, the container 6 (more specifically the cartridge 22) can be inserted into and removed from the transfer device 11. In other words, when the cover 28 is in the open position, the aforementioned body H is accessible from the outside.

When the lid 28 is in the closed position, it is not possible to insert and/or remove the container 6 (in particular the cartridge 22). In other words, housing H is not accessible from the outside.

In accordance with some non-limiting embodiments (eg, shown in Figs. 1-4), the movable block includes a body H that slides between an outward position (Fig. 1) and an inward position (Figs. 2 and 3).

In these cases, the container 6 to be used (more precisely, the cartridge 22) is inserted into the housing H located in the outward position. At this moment, the housing H moves (with the container 6, more precisely the cartridge 22) to the inner position. In the internal position, the body used rotates around axis A.

The embodiment of FIG. 1-4 differs from the embodiment of FIG. 5-8 because motor 3 and brake 25 are on the same side as actuator 24 (which is also a rotor in this case; see in particular FIG. 3).

According to some non-limiting embodiments (FIG. 20), container 6 is a conventional type control tube (eg, PCR tubes).

Preferably, but not necessarily (Fig. 21), the container 6 has a part located at the closed end 7 with a reduced internal section (relative to a conventional control tube, in particular, the specified section has a radius of less than 0.8 mm; more precisely, the radius is approximately 0.5 mm) and has a substantially cylindrical shape. This type of geometry has various advantages, including the possibility of obtaining a smaller final volume of sample 2; and a further increase in volume reduction reproducibility.

Preferably, but not necessarily (FIG. 22), the container 6 has an internal constriction 29 near the closed end 7. In particular, the internal volume of the portion of the container 6 located between the constriction 29 and the closed end 7 is less than the initial sample volume 2 and greater than the final volume obtained from sample 2.

This type of geometry reduces the risk of microparticle 18 leaking out of container 6.

According to an aspect of the present invention, there is provided a method for reducing the volume of a sample 2 containing at least one (at least partially) liquid component 3 and having a volume of up to 10 ml (in particular up to 2 ml).

The method involves the use of at least one container 6 as defined above.

The method comprises an acceleration step during which the processing unit 4 moves the container 6 containing the sample 2 so as to subject the container to an acceleration (having at least one component) oriented from the end 8 towards the closed end 7 and transversely (at in particular perpendicular) to the opening 8' in such a way that the first part (at least partially) of the liquid component 3 flows out of the container 6 passing through said opening

- 6 042126

8' and the second part (at least partially) of the liquid component 3 (in particular with a substantially defined volume) remains in the container 6, in particular in the closed end 7.

Preferably, but not necessarily, the method is carried out using the device 1 described above.

Preferably, but not necessarily, sample 2 contains at least one microparticle 18 (in particular, a plurality of microparticles 18). During the acceleration step, the processing unit 4 subjects the container 6 to acceleration in such a way that the microparticle 18 remains in the container 6, in particular at the closed end 7.

Preferably, but not necessarily, the method comprises a pre-treatment step that precedes the acceleration step, during which the container 6 containing the sample 2 is subjected to additional acceleration (having at least one component) oriented from the closed end 7 towards the end 8 (and transverse, in particular perpendicular to said opening 8') so that the microparticle 18 is located at the closed end 7, in particular in contact with the inner surface 30 of the container 6. In particular, the pre-treatment step includes centrifuging the container 6 containing sample 2.

It has been experimentally observed that in this way the risk of leakage of the microparticle 18 from the container 6 during the acceleration step is further reduced.

In this regard, it should be noted that an adhesive force F _a is generated between the microparticle 18 and the inner surface 30, which opposes the inertial force F _ic that is applied to the microparticle 18 during the acceleration step.

The above is illustrated in FIG. 28 and 29, which show an attempt to explain what has been observed experimentally. In FIG. 28 shows container 6 and sample 2 at the start of the acceleration step. In FIG. 29 shows container 6 and sample 2 at the end of the acceleration step.

In these figures, the inertial force applied to sample 2 is based on the amount of sample 2 and is denoted as F _c , and the holding force that maintains the surface of the sample without damage (and is related to surface tensions in action) is denoted as F _Y .

In particular, according to some embodiments, during the acceleration phase, as a consequence of the acceleration applied to the container 6, the first part of the sample 2 is affected by at least one first inertial force, and the second part of the sample 2 by at least one second inertial force. The first and second inertial forces are oriented from the closed end 7 towards the end 8 (transversely, in particular perpendicularly) towards the opening 8'. During the acceleration step, the first inertial force is greater than the first holding force acting between the first sample part and container 6, and the second inertial force is less than the second holding force acting between the second sample part and container 6. In particular, the first and second holding forces are determined (by the main way) by the surface tension of the sample 2 and the container 6 (more precisely, the side wall 9 of the container 6).

In a more detailed description below and in FIG. 23-29 the holding force is given as F _Y .

Preferably, but not necessarily, the processing unit 4, during the acceleration step, simultaneously subjects a plurality of containers 6 (each containing a respective sample 2) to the above acceleration. In particular, the processing unit 4 contains a plurality of sockets 5, each of which houses a corresponding container 6.

In accordance with some non-limiting embodiments, the processing unit 4 includes a moving device 11 which, during the acceleration step, imparts to the container 6 a substantially linear movement in a given direction such that the closed end 7 faces forward with respect to a given direction and end 8 faces backward with respect to a given direction. directions (figs. 9 and 25).

According to alternative embodiments, the processing unit 4 comprises a moving device 11 which, during the acceleration step, rotates the container 6 about the axis of rotation A in such a way that the centrifugal force moves the first part (at least partially) of the liquid component 3 from the container 6 passing through hole 8'.

In particular, during the acceleration phase, the container 6 is oriented essentially radially about the axis A in such a way that the end 8 faces outward.

Preferably, but not necessarily, the axis A of rotation passes through the container 6 between the closed end 7 and the end 8. Thus, during the acceleration step, the container essentially cannot be emptied beyond the limit determined by the position of the axis A (in other words, below the axis A). In these cases, by choosing the position in which the axis A passes through the container 6, it is possible to control the volume of that part (at least partially) of the liquid component 3 that remains in the container 6.

According to alternative non-limiting embodiments, during the acceleration step, the closed end 7 is positioned between the axis of rotation A and the end 8. Thus (as will be explained in more detail below), the volume of the second part of the liquid component 3 is based on the angular velocity of the container 6 during the acceleration step .

In accordance with certain non-limiting embodiments, microparticles 18 are selected from the group consisting of cells, cellular debris (in particular, cellular fragments, such as DNA and/or RNA), cellular aggregates (such as, for example, small clusters of cells, originating from stem cells such as neurospheres or mammospheres), bacteria, lipospheres, microbeads (made of polystyrene and/or magnetic), nanobeads (for example, nanobeads up to 100 nm in size), complexes formed from microbeads (in particular magnetic; in particular, with the largest size less than 500 μm) associated with cells, circulating tumor cells associated with ferrofluid, exosomes, colloidal suspensions (for example, ferrofluid), liposomes, nuclei, spores, as well as combinations of them.

In particular, the microparticles 18 are selected from the group consisting of stem cells, erythroblasts, trophoblasts, neuronal cells, epithelial cells, tumor cells, leukocytes (WBCs), stromal cells, spermatozoa, circulating tumor cells (CTCs), embryonic cells, microballoons (in in particular, with a maximum dimension less than 500 μm), a colloidal suspension (e.g. ferrofluid), complexes formed from microspheres associated with cells (e.g. stem cells, erythroblasts, trophoblasts, neuronal cells, epithelial cells, tumor cells, leukocytes (WBC) , stromal cells, spermatozoa, circulating tumor cells (CTCs), fetal cells), erythrocytes, ferrofluid-associated circulating tumor cells, and combinations thereof.

Preferably, but not necessarily, the method comprises an adjustment step which precedes the acceleration step and during which the estimated acceleration (and/or estimated angular velocity) is determined (by control unit 6) based on a substantially determined volume to be obtained (and in particular , the geometry of the container, and the constant of interaction between the material constituting said side wall and said liquid component). During the acceleration step, the effect of acceleration to which the processing unit 4 subjects the container 6 is the design acceleration (and/or the angular velocity with which the processing unit 4 exposes the container 6 is the design angular velocity).

It should be noted that, in reality, angular velocity and acceleration are equivalent (for circular motions), since they are related by the relation a _c =ω ² χd, in which a _c is the centrifugal acceleration, ω is the angular velocity, and d is the distance from the axis of rotation.

According to some non-limiting embodiments, the method comprises an insertion step during which sample 2 is inserted into container 6.

In particular, during the insertion step, the sample 2 is inserted into the container 6 using an instrument selected from the group consisting of microfluidic devices (for example, containing an inkjet system, in particular, obtained by inkjet technology), pipetting instruments, flow cytometers, micromanipulators, optical tweezers. In some non-limiting embodiments, microfluidic devices are of the types described in patent applications WO 2010/106434 and WO 2012/085884.

Preferably, but not necessarily, sample 2 is selected to contain at least one microparticle 18 using imaging, immunofluorescence, impedance, size, geometry, morphological features, and a combination of these.

In accordance with specific non-limiting embodiments, a microfluidic device is used that selects a sample containing at least one microparticle 18 (of a given type) using images, immunofluorescence, impedance, dimensions, geometry, morphological features, and a combination of these.

In some cases, it is also possible to provide for a processing step which is subsequent to the insertion step and preceding the centrifugation step, during which the sample is treated, in particular, with an additional substance.

In accordance with some non-limiting embodiments, the additional agent is a reagent (eg, for coloring and/or permeating the microparticles 18) that is inserted into the container. Alternatively or additionally, the reagent is a microparticle fixation reagent 18.

It is also possible to provide for a washing step during which a washing liquid (washing solution - washing buffer) is introduced into the container 6 and then removed during the acceleration step.

In particular, the processing step comprises a first addition sub-step during which a reagent (particularly a staining reagent and/or a reagent for permeating the microparticles 18) is introduced into container 6 containing sample 2; and a second adding sub-step during which the washing liquid is introduced into the container 6 containing the sample 2. In particular, which follows the first adding sub-step.

In accordance with some non-limiting embodiments, the processing step includes a holding sub-step that is subsequent to the first adding sub-step and preceding the second adding sub-step, during which the reagent is kept in the container 6 along with the sample 2 (in particular, at a controlled temperature between minimum and maximum temperatures).

According to some non-limiting embodiments, the soak sub-step has a duration of 10 seconds to 24 hours.

Fig. 33 illustrates a flowchart of procedures that may be followed. In these cases, the method comprises a pre-processing step SA (as described above) and an acceleration step SB following the pre-processing step. The method also includes a processing step which is subsequent to the acceleration step SB and comprises an addition sub-step SC and usually an addition sub-step SD, during which the additional substance is stored in the container 6 in contact with the sample 2; a washing step SE which is subsequent to the processing step (more precisely, subsequent to the aging sub-step SD) and during which the washing liquid is introduced into the container 6.

Here (after the washing step), according to the first variant, the pre-treatment step SA is again scheduled (and, therefore, the steps SB, SC, SD again in succession). In accordance with the second option, the acceleration stage (SB) is scheduled again (and, therefore, the stages SC, SD again in succession). The procedure can be performed several times (and usually ends with the SB step).

Preferably, but not necessarily, during the acceleration step, the temperature of the containers 6 is maintained within a predetermined interval. In particular, the processing unit 4 comprises a temperature maintenance system adapted to maintain the temperature of the containers 6 (more specifically the nests 2) within the desired range. According to some non-limiting embodiments, the temperature maintenance system includes a temperature sensor and a heating and/or cooling device that operates based on data detected by the temperature sensor.

The adjustment step can be implemented using previously obtained experimental data (which, for example, show that with a given container 6 - a given shape and a given material - with a given component 3 and applying a given acceleration - and / or angular velocity - a given final volume is obtained ). An example of a curve obtained from the obtained experimental data that relates the angular velocity to the volume of the second part of the liquid component 3 (which remains in the container after the acceleration step) is shown in FIG. 32.

In particular (therefore), the adjustment step is in some cases preceded by a calibration step during which different final volume values obtained for different accelerations (and/or angular velocities) are measured. In particular, during the calibration step, a calibration curve (or calibration function) is created, which is then used during the tuning step to obtain a calculated acceleration (and/or a calculated angular velocity) based on a substantially defined volume to be obtained.

Alternatively or additionally, during the adjustment step, the first function that relates the inertial force to the acceleration intersects with the second force function due to surface stresses and evaluates (also graphically) for what amount of acceleration (and/or angular velocity) these two forces are equivalent for desired volume of the second part (at least partially) of the liquid component 3. In other words, in the control step, after determining the volume of the second part (at least partially) of the liquid component 3 to be left in the container 6, it is estimated for what amount acceleration (and/or angular velocity) the inertial force is equivalent to the force due to surface stresses based on a first function that relates the inertial force to the acceleration (and/or angular velocity) and a second function that relates the inertial force to the distance from the closed end 7 or from the A axis.

During the acceleration step, the processing unit subjects the container 6 to an acceleration with a specified amount.

If the closed end 7 is located between axis A and end 8, and the cross section of the container 6 is constant (as shown in Fig. 23), then the first function is

F _c (d) amord (1) the second function is

Ρ _γ α2πΚΔγ (2) where F _c - inertial force (more precisely, centrifugal force), m - sample mass, ω - angular velocity, d - distance from axis A,

F _γ - holding force (due to surface stresses),

R is the inner radius of container 6, and

Δγ is a parameter depending on the type of sample 2 (more precisely, on the type of liquid component 3) and on the material from which the container 6 is made.

Parameters Δγ can be found in tabular form in reference books or can be determined experimentally.

- 9 042126

In this case, the volume of the second part (at least partially) of the liquid component 3 (namely, the part that remains in the container after the acceleration step) is based (while maintaining the inner perimeter of the container, mass and fixed surface stress) on the angular velocity (and, therefore, can be modulated by changing the angular velocity). More precisely, as the angular velocity increases, the volume decreases.

In this case, the volume of the second part (at least partially) of the liquid component 3 (namely, the part that remains in the container after the acceleration step) is based (while maintaining the inner perimeter of the container, the mass and a fixed surface tension) on the angular velocity (and, therefore, can be modulated by changing the angular velocity). More precisely, as the angular velocity increases, the volume decreases.

If the axis A passes through the container 6 and is located between the closed end 7 and the end 8, and the cross section of the container 6 is constant (as shown in Fig. 24), then the first and second functions are the functions (1) and (2) described above. In contrast to the previous situation, in this case it is not possible (by increasing the angular velocity) to force the entire sample 2 to flow out, since the part located between the axis A and the closed end 7 remains (in any case) inside the container 6.

If the container 6 has a variable diameter and moves in a linear manner (as shown in Fig. 25), then the second function is the function (2) (note, however, that in this case the inner perimeter of the container 6, and hence the inner radius R, changes according to as it moves along the longitudinal extension of the container 6) and the first function is

F _c amaap d ² (3) in which Fc is the inertial force, m is the mass of the sample 2, a is the acceleration, d is the distance relative to the closed end 7, β is the proportionality factor depending on the specific gravity of the sample 2 (more precisely, the liquid component 3) and container geometry 6 (and therefore can be predetermined).

If the closed end 7 is located between axis A and end 8, and the section of the container 6 is variable (as shown in Fig. 26), then the second function is the function (2) (note, however, that in this case the inner perimeter of the container 6, and therefore, the inner radius R, changes as it moves along the longitudinal extension of the container 6) and the first function is

F _c (d)o0d ² ro ² (4) where F _c is the inertial force, d is the distance from axis A, β is the proportionality factor, which depends on the specific gravity of the sample 2 (more precisely, the liquid component 3) and the geometry of the container 6 (and therefore can be predetermined).

In particular, function (4) is obtained from

F _c (d)amco ² dapKR ² co ² d (5) where ρ is the specific gravity of the sample.

If the closed end 7 is located between the axis A and the end 8, and the cross section of the container 6 is variable (as shown in Fig. 27), then the second function is the function (2) (note, however, that in this case the inner perimeter of the container 6, and therefore, the inner radius changes as it moves along the longitudinal expansion of the container 6) and the first function is the function (4). Unlike the previous situation, in this case it is not possible (by increasing the angular velocity) to force the entire sample 2 to flow out, since the part located between the axis A and the closed end 7 remains in the container 6.

In view of the above, the first function is the function (1), or (3), or (4) and the second function is the function (2). This is especially important when the closed end 7 is located between axis A and end 8.

It should be noted that the functions (1)-(5) and the illustrations of FIG. 23-27 were formulated (to substantiate what was observed and to automate some embodiments) after the present invention experimentally demonstrated unexpectedly that it can obtain sample 2 volume reductions in an extremely accurate and reproducible manner.

In accordance with some non-limiting embodiments, sample 2 is prepared from a pre-sample containing microparticle(s) 18 and additional microparticles. In particular, sample 2 is obtained by selectively isolating the microparticle/microparticles 18 relative to additional microparticles. This is done with a separating unit containing a system selected from the group consisting of dielectrophoresis, optical tweezers, magnetophoresis, acoustic phoresis, traveling waves, heat flow, local fluid movements generated by electrothermal flow, local fluid movements generated by electrohydrodynamic forces, and a combination of them.

In some non-limiting cases, the separator block contains a system selected from

- 10 042126 group consisting of dielectrophoresis, optical tweezers, magnetophoresis, acoustophoresis and a combination of them.

In particular, the separating block comprises a system capable of exerting force directly on the microparticle(s) 18 (particularly without applying force to the fluid that transmits the motion to the microparticle(s) 18).

In accordance with specific embodiments, the separation unit contains a dielectrophoresis unit (or system) as described, for example, in at least one of patent applications WO-A-0069565, WO-A-2007010367, WO-A-2007049120. More specifically, the separating block functions as described in patent applications WO 2010/106434 and WO 2012/085884).

Preferably, but not necessarily, the separation unit is part of one of the above microfluidic devices, in particular as described in patent applications with publication numbers WO 2010/106428 and WO 2010/106426. This microfluidic system is used to obtain sample 2 from the preliminary sample.

According to some non-limiting embodiments, during the acceleration step, a magnetic force is applied to the microparticle 18, in particular having at least one magnetic component directed towards the closed end 7 (and/or side wall 9).

Alternatively (or additionally) during the acceleration phase, an (additional) magnetic force is applied towards the opening 8'. In particular, said magnetic force acts on the ferrofluid colloidal slurry (so as to facilitate the outflow of the colloidal slurry from the container 6).

The method and apparatus according to the present invention can also preferably be used for sample preparation for genetic analysis, sample preparation for cell sorting, cell staining and cell washing.

Unless expressly stated otherwise, the contents of references (articles, books, patent applications, etc.) cited in this text are referenced in their entirety in this document. In particular, these references are included herein for reference.

Further characteristics of the present invention will become apparent from the following description of purely illustrative, non-limiting examples.

Example 1

This example describes the tests performed in the traditional way of reducing the volume of samples 6, including the liquid component 3 and at least one microparticle 18. These samples were pre-processed using a centrifuge to ensure that the microparticle 18 is located in the closed end 7 of the container 6 (control tube, as shown in Fig. 20).

In particular, a total of 260 tests were performed by three different operators (A, B and C) who, starting with 6 samples with an initial volume of approximately 113 μl, should have obtained a volume of 1 rl.

Each operator collected the excess liquid manually using a pipette (and tilting the control tube containing the sample).

Operators A and B performed 90 decrements each. Operator C performed 80 decrements.

It took 10 hours of work to complete the operations (adding the working time of each operator). All the results obtained are given in table. 1 and in FIG. 31 (the x-axis shows the sample volume obtained after reduction, and the y-axis shows the number of times the reduced sample represented this volume), in which the darker color shows the times the microparticle was lost, and the broken line defines the area in which ( for which finite volumes) the indicated losses occurred.

Table 1

Manual sample reduction: 260 tests Meaning 1.73 rl Maximum spacing 4.3 rl Standard deviation 1.03 rl Performance indicator 94.7%

By the maximum interval, we mean the difference between the maximum volume and the minimum volume obtained after reduction operations. The average value is the average of the sample volumes obtained after reduction. The results obtained show that the conventional procedure is not reliable (the failure rate relative to the cases in which the microparticle 18 was lost is not negligible) and the average volume significantly exceeds the target volume (1 rl).

The results obtained, broken down by operators, are shown in the following tables. 2-4.

- 11 042126

table 2

Operator A Meaning 2.65 rl Maximum spacing 3.35μΐ Standard deviation 0.79 rl Performance indicator 98.9%

Table 3

Operator B Meaning 1.16 rl Maximum spacing 2.60 rl Standard deviation 0.51 rl Performance indicator 94.7%

Table 4

Operator C Meaning 1.28 rl Maximum spacing 4.06 rl Standard deviation 0.96 rl Performance indicator 89.9%

These results show that the accuracy of the procedure is highly dependent on the operator and his/her ability to work manually.

It should be noted that each operator performed reductions in three separate work sessions. The results obtained by each operator (indicated on the x-axis) for each section are shown graphically in FIG. 30 (Y-axis shows sample volume obtained after reduction).

As you can see, even a single operator tends to get different results at different times.

Example 2

This example describes a comparison between tests performed by the traditional volume reduction method and the (automatic) method according to the present invention. The tests were carried out with an initial volume of approximately 113 rl, and the target volume was 1 rl.

The traditional method was implemented as described in example 1.

To carry out the method in accordance with the present invention, the device 1 (made to operate at 4200 rpm) shown in Figures 5-8 and the container of FIG. 20 (more precisely, a strip of control PCR tubes with a volume of 0.2 ml each). The distance between the axis A (which passed through the container) and the closed end 7 was 2.22 mm.

The results obtained are presented in table. 5.

Table 5

Manually Automatically Number of reduced samples 360 180 Meaning 1.66 rl 1.22 rl Maximum spacing 4.34 rl 1.05 rl IQR(interquartile range, interquartile range) 1.31 rl 0.30 rl Standard deviation 0.95 rl 0.25 rl Performance indicator 92.8% 99.4% 1 particle lost every 10 samples 1 particle lost every 180 samples

Based on the above data, the present invention represents a significant and unexpected improvement in every recorded aspect.

In table. 6 below compares the same number of tests performed by the traditional method and the method according to the present invention.

- 12 042126

Table 6

Manually Automatically Number of reduced samples 96 96 elapsed time 220 minutes 15 minutes Meaning 1.66 rl 1.21 rl Standard deviation 0.95 rl 0.25 rl Performance indicator 92% 98%

It can be immediately noticed that the time required to obtain samples with reduced volume is dramatically reduced when implementing the method of the present invention.

Claims (25)

1. A device for reducing the volume of a sample (2) containing at least one, at least partially liquid component (3) and having a volume of up to 10 ml, and the device (1) contains a processing unit (4) equipped with at least at least one socket (5) for at least partial support of at least one container (6) having an internal space, a first closed end (7), a second end (8) made with a hole (8') providing communication between the inner space and the outer part, and at least one side wall (9), while the processing unit (4) contains a collecting device (10), which is configured to collect the first part of the specified at least partially liquid component (3 ) and is located outside the seat (5) and near the seat (5), and the processing unit (4) is configured to move the seat (5) in such a way as to expose the seat (5) to an acceleration effect having at least one component that is orientedfrom the collecting device (10) towards the socket (5) in such a way that the first part of the specified, at least partially liquid component (3) flows out of the container (6) and reaches the collecting device (10), and the second part of the specified, at least partially liquid component (3) with essentially a certain volume remained in the container (6), in particular in the area of the specified first closed end (7), while the processing unit (4) contains a moving device (11), which is made with the possibility of rotation of the nest (5) around the axis (A) of rotation in such a way that the centrifugal force forces the specified first part of the at least partially liquid component (3) to flow from the container (6) into the collecting device (10), and the nest ( 5) is made in such a form that the axis (A) of rotation passes through the container (6) between the first closed end (7) and the second end (8). 2. The device according to claim 1, in which the processing unit (4), in particular the transfer device (11), is made in such a form that one or more flexible well plates can be installed directly on the transfer device (11) so that pass (bend) around the axis (A) of rotation. 3. The device according to any one of claims 1 and 2, in which the collecting device (10) contains a holding system (12) (in particular, a reservoir) for retaining at least a fraction of said first part of said at least partially liquid component (3). 4. The device according to claim 3, in which the containment system (12) contains an element selected from the group consisting of an absorbent material (13), a capillary trap, a liquid trap, and a combination of them; a capillary trap containing a plurality of grooves (14) having a width of less than 2.0 mm, in particular less than 0.9 mm; a liquid trap comprising a collecting chamber (15) provided with an inlet (16) and at least one movable wall (17) capable of moving between a closed position in which it prevents liquid from flowing through the inlet (16) and a position opening in which fluid can flow through the inlet (16), wherein the wall (17) is adapted to move to the opening position when the said acceleration used is applied, and as soon as the acceleration is no longer applied to the seat (5), the wall (17) returns to closing position. 5. The device according to any one of claims 1 to 4, in which the processing unit (4) contains a plurality of sockets (5), each of which is adapted to accommodate the corresponding container (6), and the processing unit (4) contains a moving device (11) , which is adapted to force all sockets (5) to make the same movement. 6. The device according to claim 5, in which the moving device (11) is adapted to rotate the seats (5) around the axis (A) of rotation in such a way that the centrifugal force moves the said first parts of the specified at least partially liquid component (3) in collecting device (10), wherein the nests (5) are located in at least one row, which is essentially parallel to the axis (A) of rotation. 7. The device according to any one of claims 1 to 6, in which the processing unit (4) contains a moving device - 13 042126 stem (11), which is adapted to rotate the socket (5) around the axis (A) of rotation in such a way that the centrifugal force moves the specified first part of the specified at least partially liquid component (3) to the collecting device (10), moreover, the processing unit (4) contains a plurality of additional peripheral sockets (19), which are located around the axis (A) of rotation and are adapted to accommodate additional containers (6), each of which contains a corresponding sample (2), and must be rotated by a moving device (11 ) around the axis (A) of rotation in such a way that centrifugal force is applied to said at least partially liquid component (3) contained in additional containers (6) towards the first closed end (7) of additional containers (6) . 8. Apparatus according to any one of claims 1 to 7, wherein the processing unit (4) comprises a temperature maintenance system which is adapted to keep the temperature of the containers (6), in particular the nests (5), within a desired range, in particular between a predetermined minimum temperature and a predetermined maximum temperature, wherein, in particular, the temperature storage system comprises a temperature sensor and a heating and/or cooling device which is adapted to operate on the basis of data detected by the temperature sensor. 9. The device according to any one of claims 1 to 8, in which the processing unit (4) contains a first magnet, which is located in the region of the end face of the socket (5) opposite the collecting device (10), and, in particular, the first magnet is located in the region of the closed end (7). 10. The device according to any one of claims 1 to 9, in which the processing unit (4) contains a second magnet, which is located in the area of the socket (5), in particular in the area of the end face of the socket (5) in the area of the collecting device (10), and , in particular, the magnet is located in the area of the second end (8), more specifically the hole (8'). 11. Apparatus according to any one of claims 1 to 10, wherein during the acceleration step, due to the acceleration applied to the container (6), at least one first inertial force and at least one second inertial force acts on the first part of the sample (2). force acts on the second part of the sample (2), and the processing unit (4) contains a moving device (11), which is adapted to rotate during the acceleration stage the container (6) around the rotation axis (A) passing through the container (6) between the first closed end (7) and second end (8) such that centrifugal force forces said first portion of said at least partially liquid component (3) to flow out of container (6) into collecting device (10); or the first inertial force, which is greater than the first holding force acting between the first sample part and the container (6), and the second inertial force, which is less than the second holding force acting between the second sample part and the container (6), are oriented from of the first closed end (7) towards the second end (8) transversely (in particular perpendicularly) to said opening (8'). 12. A method for reducing the volume of a sample (2) containing at least one at least partially liquid component (3) and having a volume of up to 10 ml, implemented by means of a device according to any one of claims 1 to 11, the method comprising using at least one container (6) having an inner space, a first closed end (7), a second end (8) provided with an opening (8') providing communication between the inner space and the outer part, and at least one side wall (9), characterized in that it includes an acceleration step during which the processing unit (4) moves the container (6) containing the sample (2), exposing the container (6) to an acceleration that has at least one component that oriented from the second end (8) towards the first closed end (7) and is transverse (in particular, perpendicular) to said opening (8'), so that the first part of said at least partially liquid component (3) flows out leaves the container (6), flowing through the specified opening (8'), and the second part of the specified at least partially liquid component (3) with a substantially defined volume remains in the container (6), in particular in the specified first closed end (7), while the processing unit (4) contains a moving device (11), which during the acceleration stage rotates the container (6) around the rotation axis (A) in such a way that the centrifugal force forces the said first part of the said at least the partially liquid component (3) to flow out of the container (6) passing through said opening (8'), with the rotation axis (A) passing through the container (6) between the first closed end (7) and the second end (8). 13. The method according to claim 12, characterized in that the sample (2) contains at least one microparticle (18); during the acceleration step, the processing unit (4) exposes the container (6) to acceleration in such a way that the microparticle (18) remains in the container (6), in particular in the region of said first closed end (3). - 14 042126 14. The method according to claim 13, characterized in that it comprises a pretreatment step which precedes the acceleration step and during which the container (6) containing the sample (2) is subjected to additional acceleration having at least one component that is oriented from the first closed end (7) towards the second end (8) and is transverse (in particular, perpendicular) to said hole (8'), so that the microparticle (18) is located in the region of the first closed end (3). 15. The method according to any one of claims 12 to 14, characterized in that the processing unit (4) simultaneously exposes a plurality of containers (6) to said acceleration during the acceleration step; in particular, the processing unit (4) contains a plurality of sockets (5), each of which houses a corresponding container (6). 16. Method according to any one of claims 12 to 15, characterized in that during the acceleration step the first closed end (7) is located between the rotation axis (A) and the second end (8). 17. The method according to any one of claims 12-16, characterized in that the microparticles (18) are selected from the group consisting of cells, cell debris (in particular cell fragments, e.g. DNA and/or RNA), cell aggregates (such as , for example, small aggregations of cells derived from stem cells such as neurospheres or mammospheres), bacteria, lipo-beads, microbeads (polystyrene and/or magnetic), nanobeads (e.g. nanobeads up to 100 nm in size), aggregations formed microbeads (in particular magnetic beads, in particular with a maximum dimension less than 500 μm) associated with cells, circulating tumor cells associated with ferrofluid, exosomes, colloidal suspension (e.g. ferrofluid), liposomes, nuclei, spores and combinations thereof ; in particular, the microparticles (18) are selected from the group consisting of stem cells, erythroblasts, trophoblasts, neuronal cells, epithelial cells, tumor cells, leukocytes (WBCs), stromal cells, spermatozoa, circulating tumor cells (CTCs), fetal cells, microbeads (in particular, with the largest dimension less than 500 µm), colloidal suspension (for example, ferrofluid), accumulations formed by microspheres associated with cells (for example, stem cells, erythroblasts, trophoblasts, neuronal cells, epithelial cells, tumor cells, leukocytes (WBC ), stromal cells, spermatozoa, circulating tumor cells (CTCs), fetal cells), erythrocytes, ferrofluid-associated circulating tumor cells, and combinations thereof. 18. The method according to any one of claims 12 to 17, characterized in that it includes an insertion step during which the sample (2) is inserted into the container (6); and a processing step following the insertion step and up to the acceleration step, during which the sample (2) is treated, in particular with an additional agent. 19. The method according to claim 18, characterized in that the processing step comprises a first adding sub-step during which the reagent (in particular the microparticle staining reagent (18) and/or the microparticle permeation reagent (18) and/or the microparticle fixation reagent (18)) is inserted into the container (6) containing the sample (2); and a second adding sub-step which follows the first adding sub-step and during which the rinsing liquid is inserted into the container (6) containing the sample (2), and in particular the processing step comprises a holding sub-step which follows the first adding sub-step and before the second an addition sub-step and during which the reagent remains with the sample (2) (more specifically, at a controlled temperature ranging between a minimum temperature and a maximum temperature). 20. A method according to any one of claims 12 to 19, characterized in that during the acceleration step, the collecting device (10), which collects the first part of said at least partially liquid component (3), is located in the region of the opening (8' ) outside the container (6). 21. The method according to claim 20, characterized in that the collecting device (10) comprises a holding system (12) for retaining at least a fraction of the first part of said at least partially liquid component (3), moreover, in particular , the containment system (12) contains an element selected from the group consisting of an absorbent material (13), a capillary trap, a liquid trap, and a combination of them; a capillary trap comprising a plurality of grooves (14) having a width which is less than 2.0 mm, in particular less than 0.9 mm; a liquid trap comprising a collection chamber (15) which is provided with an inlet (16) and at least one movable wall (17) that is movable between a closed position in which it prevents liquid from flowing through the inlet (16) and a position an opening in which fluid can flow through the inlet (16); during the acceleration step, said acceleration (in particular directly and/or indirectly by means of said at least partly liquid component (3)) moves the movable wall (17) to the opening position; as soon as acceleration is no longer applied to the container (6), in particular to the holding system (12), the movable wall (17) returns to the closed position. 22. A method according to any one of claims 12 to 21, characterized in that during the acceleration step, the temperature of the container (6) is maintained within a predetermined temperature range. 23. The method according to any one of claims 12 to 22, characterized in that it comprises an insertion step during which the sample (2) is inserted into the container (6) using an instrument selected from the group consisting of microfluidic devices, pipetting instruments, flow cytometers, micromanipulators, optical tweezers, and, in particular, the sample (2) is selected so that it contains at least one microparticle (18) using images, immunofluorescence, impedance, dimensions, geometry, morphological features, as well as their combination . 24. The method according to any one of claims 12-23, characterized in that the sample (2) contains at least one microparticle (18), in particular provided with at least one magnetic component; during the acceleration step, the magnetic force acts on the microparticle (18) towards the side wall (9) and/or the closed end (7). 25. A method according to any one of claims 12 to 24, characterized in that during the acceleration step a magnetic force is applied towards the opening (8'), in particular in such a way as to remove undesired material (e.g. ferrofluid) from the container ( 6).