Electrophoresis cell, free-flow electrophoresis device containing same and uses thereof

Abstract

The invention relates to a free-flow electrophoresis cell for purifying and / or separating an initial solution containing a product to be purified and / or separated, comprising - at least one plate X between two adjacent plates Y: X being an electrophoresis plate made of inert material, Y being a sealed plate made of sapphire or alumina Al2O3 containing 99% α-Al2O3, - an electrophoresis chamber comprising s sub-units Ui, s representing an integer from 2 to 5 and i being an integer varying from 1 to s, - a cooling circuit, characterized in that the electrophoresis chamber comprises: - intermediate channels connecting the two successive sub-units.

Description

[0001] DESCRIPTION

[0002] Electrophoresis cell, free-flow electrophoresis device containing it and its uses

[0003] The present invention relates to a new electrophoresis cell, a new free-flow electrophoresis device with a new configuration and its uses.

[0004] High-performance liquid chromatography (HPLC) and zone electrophoresis using a support are analytical and / or preparative techniques for separating molecules, particularly biomolecules, present in a mixture. However, the need for a stationary phase in HPLC used in industrial processes for separation and / or purification leads to significant costs due to the recycling of various solvents and the regular replacement of solid phases. Furthermore, zone electrophoresis has limitations for industrial-scale preparative use and is primarily employed in analytical techniques.

[0005] There is a need to develop a new electrophoresis cell enabling effective and optimized purification and / or separation of products contained in an initial solution by free-flow electrophoresis.

[0006] There is therefore a need to develop an adaptable, robust device without the need for demanding maintenance, operating in continuous flow, designed for easy use and modular implementation, in order to allow purification and / or separation of the expected products in volumes and quantities on an industrial scale.

[0007] One of the aims of the invention is to provide an electrophoresis cell.

[0008] Another objective of the invention is to provide a device for the purification and / or separation of molecules by free-flow electrophoresis that can operate in continuous flow and on an industrial scale.

[0009] Another objective of the invention is a process for the purification and / or separation of molecules, in particular biomolecules, adaptable to an industrial scale.

[0010] A first object of the present invention relates to a free-flow electrophoresis cell for purifying and / or separating an initial solution containing a product to be purified and / or separated, comprising at least one plate X between two adjacent plates Y:

[0011] X being an electrophoresis plate (1) made of inert material,

[0012] Y being a sealed plate (2) of sapphire or alumina Al2O3 with 99% a-ALCh, an electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5 and / being an integer ranging from 1 to s, in which each subunit comprises o a hollowed-out part in a plate X closed between two plates Y,

[0013] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, c, di) and 2 upper and lower faces (ei, f),

[0014] ■ said hollowed-out part being of length Loi and width Lai,

[0015] ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0016] ■ the lateral faces (ai, bi) being parallel to each other, the face (a) being delimited by two edges (AT, A2 ) of dimension Lai and the face (bi) being delimited by two edges (B1 i, B2>) of dimension Lai,

[0017] ■ the lateral faces (c, di) being parallel to each other, the face (c) being delimited by two edges (CT, C2) of dimension Law and the face (di) being delimited by two edges (D1 i, D2>) of dimension Law, o ni successive inputs Ei(1), Ei(2) to Ei(ni-1), Ei(n), ni being an integer from 4 to 9, distributed on the face (a) between A1 i and A2i and aligned along a direction parallel to A1 i and A2i, o mi successive outputs from Si(1), Si(2) to Si(mi-1), Si(mi), mi being an integer from 4 to 12, distributed on the face (bi) between B1 i and B2i and aligned along a direction parallel to B1 i and B2i, such that Si(1) faces Ei(1) and Si(mi) faces Ei(n) along a direction parallel to C1 i and D1 i, a cooling circuit, characterized in that the electrophoresis chamber comprises:

[0018] (s-1) intermediate channels Vk, k being an integer ranging from 1 to (s-1), each channel Vk connecting the two successive subunits Uk and Uk+i by one of the outputs SVk(Uk) chosen from Sk(2) to Sk(mk-1) of the subunit Uk to one of the inputs EVk(Uk+i) of the subunit Uk+i chosen from Ek+i (2) to Ek+i (nk+i-1), said input EVk(Uk+i) and said output SVk(Uk) being respectively the input and output connected by the intermediate channel Vk.

[0019] The inventors have demonstrated a new electrophoresis cell configuration that optimizes the purification and / or separation of an initial solution containing a product to be purified and / or separated, which may consist of a mixture of compounds of interest. In this cell, separation and / or purification are effectively implemented, as the cell allows for the recovery, at its various outlets, of a solution of the purified and / or separated product or solutions of the purified and / or separated components of said product.

[0020] The configuration of the cell of the invention, comprising an electrophoresis chamber made up of subunits connected by intermediate channels, allows for a succession of separations and / or purifications by free-flow electrophoresis. This is achieved by selecting the portion of the flow as the solution to be purified and / or separated, with the possibility of modulating and renewing the free-flow electrophoresis separation conditions independently in each subunit. Furthermore, the cell is adapted to recover, at the outlet of one of the subunits, one of the compounds of interest from the purified and / or separated product and to continue the separation and / or purification of the other compounds of said product in the other subunits.

[0021] This new electrophoresis cell can be integrated into an industrial-sized electrophoresis cell device and used in a continuous flow free-flow electrophoresis purification and / or separation process.

[0022] The electrophoresis cell according to the invention is characterized by the presence of an intermediate channel or intermediate channels linking two subunits of the electrophoresis chamber.

[0023] An electrophoresis chamber of s subunits has (s-1) intermediate channels Vk, k being an integer ranging from 1 to (s-1), each intermediate channel linking the two successive units Uk and Uk+i.

[0024] Two subunits Uk and Uk+i are said to be successive when they are connected to each other by at least one intermediate channel Vk, and when, in the circulation of fluids in the electrophoresis chamber, a part of the flux passes from the subunit Uk to the subunit Uk+i through the channel Vk.

[0025] Each intermediate channel extends from an output of a first subunit, distinct from the two outputs located respectively at the two ends of the alignment of outputs, to an input of the next subunit, distinct from the two inputs located respectively at the two ends of the alignment of inputs.

[0026] The Vk channel extends from an output of the unit Uk, hereafter named SVk(Uk), which is different from the two outputs Sk(1) and Sk(mk) located at the ends of the alignment of outputs, to one of the inputs of the unit Uk+i, hereafter named SEk(Uk+i), which is different from the two inputs Ek+i(1) to Ek+i(nk+i) located respectively at the ends of the alignment of inputs.

[0027] The intermediate channel Vk has the function of directing part of the flow circulating in the subunit Uk, this flow containing the product to be separated and / or purified or one of the compounds to be purified and / or separated said product or a mixture of compounds of said product to be purified and / or separated, towards the subunit Uk+i, this in the absence of an electric field.

[0028] The intermediate channel allows for the selection of a portion of the outgoing flow from a first subunit, excluding the two flows corresponding to the liquid cathode and liquid anode respectively. This portion of the flow is then introduced as a new solution to be purified and / or separated into a subsequent second subunit. This enables another separation and / or purification step by free-flow electrophoresis under adaptable conditions that may be identical or different from those of the first subunit. These adaptable conditions include, in particular, the dimensions of the subunit, the electric field, the fluid flow rate, and the buffer solutions.

[0029] Thus, for s equal to 2, the electrophoresis chamber comprises two subunits Ui and U2 and an intermediate channel V1 linking the two subunits U1 and U2.

[0030] For s equal to 3, the electrophoresis chamber comprises three subunits Ui, U2 and U3 and two intermediate channels V1 and V2, V1 linking U1 and U2, V2 linking U2 and U3.

[0031] For s equal to 4, the electrophoresis chamber comprises four subunits Ui, U2, U3 and LU and three intermediate channels Vi, V2 and V3, V1 linking U1 and U2, V2 linking U2 and U3, V3 linking U3 and LU.

[0032] For s equal to 5, the electrophoresis chamber comprises five subunits Ui, U2, U3, U4 and U5 and four intermediate channels Vi, V2, V3 and V4, V1 linking U1 and U2, V2 linking U2 and U3, V3 linking U3 and U4, V4 linking U4 and U5.

[0033] The intermediate channel V1 extends from an output SVi(Ui) of subunit U1, chosen from the (mi-2) outputs from Si(2) to Si(mi-1), to one of the inputs EVI(U2) of subunit U2 chosen from the (n2-2) inputs from E2(2) to E2(n2-1), where r is the number of outputs of U1 and n2 is the number of inputs of U2. The intermediate channel V2 extends from an output SV2(U2) of subunit U2, chosen from the (ni2-2) outputs from E2(2) to S2(ri2-1), to one of the inputs EV2(U3) of subunit U3 chosen from the (ns-2) inputs from Es(2) to Es(n3-1), where m2 is the number of outputs of U2 and ns is the number of inputs of U3. The intermediate channel V3 extends from an output SVs LL) of the subunit U3, chosen from the (ms-2) outputs from Ss(2) to S3(rri3-1), to one of the inputs EVs LL) of the subunit U4 chosen from the (n4-2) inputs from E4(2) to E4(n4-1), m3 being the number of outputs of U3 and being the number of inputs of LU.The intermediate channel V4 extends from an output SV4(U4) of the subunit U4, chosen from the (ni4-2) outputs of S4(2) to S4(rri4-1), to one of the inputs EV4(U5) of the subunit U5 chosen from the (ns-2) inputs of Es(2) to E5(n5-1), ru being the number of outputs of U4 and ns being the number of inputs of U5.

[0034] The term "free-flow electrophoresis" refers to electrophoresis that does not employ a stationary phase, i.e., without the use of a solid phase to support the migration of species during electrophoresis.

[0035] The "electrophoresis cell" is a component unit of a device. The electrophoresis cell comprises an electrophoresis chamber, including the walls of plates X and Y that form and close the electrophoresis chamber, and a fluidic circuit connecting the inlets and outlets of said chamber. This internal fluidic circuit includes supply, recovery, and discharge channels connecting the inlets and outlets of said electrophoresis chamber to circuits outside the cell.

[0036] The term "plate" refers to a rigid element, generally in the form of a rectangular parallelepiped, in which at least two faces are parallel to each other and these parallel faces constitute the majority of the element's total surface area; that is, the area of ​​these two parallel faces represents more than half of the total surface area. These two faces are called the "surfaces" or "lower and upper surfaces" of the plate.

[0037] The distance between these two surfaces of the plate defines the thickness of the plate.

[0038] The term "electrophoresis plate" refers to the plate, designated X, containing at least one subunit or a hollowed-out portion of the electrophoresis chamber. Plate X is made of a material inert to electrophoresis.

[0039] By "sealed plate", named Y, we mean a plate whose purpose is to ensure the fluidic sealing of the electrophoresis cell.

[0040] The cell includes at least one plate X located between two plates Y, which ensure the sealing of the electrophoresis chamber of plate X.

[0041] The Y plate is made of inert material, electrically insulating and thermally conductive in order to ensure the chemical and electrical inertness of the device necessary for the free-flow electrophoresis process and to ensure thermal conductivity between the cooling circuit and the subunits of the electrophoresis chamber to allow control of the temperature of the electrophoresis cell.

[0042] The Y plate is made of sapphire or alumina (Al2O3) with 99% a-ALCh.

[0043] In the preferred mode of the invention, the Y plates are made of sapphire or of alumina Al2O3 with 99% of a-Al2O3, preferably sapphire.

[0044] The term "sapphire" refers to a material composed of corundum, i.e., alumina (Al2O3) comprising 99% by weight of the (1-Al2O3) phase.

[0045] Advantageously, the Y plate exhibits a Mohs hardness of 9 (Coridon), a thermal conductivity of 30 W / m / K to 50 W / m / K. The Y plate exhibits high mechanical resistance, particularly compared to a glass plate.

[0046] As a non-limiting example, the Y-shaped sapphire plate is supplied by Saint-Gobain (Luxium Solutions).

[0047] The mechanical resistance of the sapphire plates makes it possible to ensure the sealing of the electrophoresis cell by mechanical clamping of the plates, advantageously, without resorting to the use of a chemical glue to ensure the seal.

[0048] The sapphire plate also has the advantage of being transparent.

[0049] The introduction of a cooling circuit with the use of sapphire or alumina plates with 99% a-AhCh as a means of separation between the cooling circuit and the subunits of the electrophoresis chamber induces excellent temperature control and homogeneity such that it allows: a thickness hi of the subunits constituting the electrophoresis cell that can vary from micrometer to millimeter and thus the increase in the processing capacity of the solutions to be purified or separated, purification and / or separation of thermosensitive molecules, the introduction of a temperature gradient in the subunits.

[0050] The nature of the materials of the Y plates, which are sapphire or alumina (Al2O3) with 99% a-AhCh, is an important characteristic of the electrophoresis cell of the invention for regulating and controlling the temperature, which is a critical parameter in the free-flow electrophoresis process, particularly for maintaining a laminar flow system, in addition to the aspect of protein denaturation.

[0051] The electrophoresis cell comprises at least one sequence of YXY plates, with plate X located between two adjacent Y plates. "Adjacent plates" refers to plates directly joined at their surfaces.

[0052] The electrophoresis cell contains a single YXY sequence when all subunits are located on the same plate. It can contain up to s different YXY plate sequences when all subunits are distributed across different X plates, allowing the subunit height (hi) to be adjusted to suit the operating conditions for separation and / or purification. The number of YXY sequences is less than s when at least two subunits are located in the same plate.

[0053] The "electrophoresis chamber" is the part of the electrophoresis cell in which the circulation and separation of the product to be purified and / or separated take place.

[0054] In the present invention, the electrophoresis chamber is made up of multiple subunits connected to each other by intermediate channels.

[0055] By "a subunit of the electrophoresis chamber" we mean a part of an X-ray electrophoresis plate comprising:

[0056] - a hollowed-out section in the electrophoresis plate X, delimited by lateral walls and the walls of the surfaces of the plates Y adjacent to the aforementioned plate X, closing off the hollowed-out section, and

[0057] - the entrances and exits located in the side walls of said hollowed-out part.

[0058] The subunits of the electrophoresis chamber form the space in which free-flow electrophoresis takes place.

[0059] The number of subunits "s" is at least two, it can vary from 2 to 5, namely 2, 3, 4 and 5, but it can also be beyond 5, between 6 and 10, for example 6, 7, 8, 9 or 10 when the separation conditions, in particular flow rate and yield of the product to be purified and / or separated, are appropriate.

[0060] Each subunit U in which free-flow electrophoresis is carried out comprises a hollowed-out portion of a plate X, closed by the lateral walls in the plate X and by the upper and lower walls of the surfaces of the two adjacent plates Y, said hollowed-out portion being in the general shape of a rectangular parallelepiped.

[0061] The term "in the shape of a rectangular parallelepiped" means that this hollowed part generally fits within a rectangular parallelepiped of length Loi, width Lai and height hi corresponding to the thickness of the plate X. The general shape of the hollowed part is not taken into account in defining the means that may be present in the electrophoresis chamber, such as the piping means present at the inlets and outlets and the protrusions.

[0062] The rectangular parallelepiped comprises four lateral faces (ai, bi, c, di) and two faces (ei, f), respectively lower and upper.

[0063] The lateral faces (ai, bi) are parallel to each other and of the same dimensions.

[0064] The face (a) is delimited by two edges (A1 i, A2 ) of dimension Lai, spaced hi apart.

[0065] The face (bi) is delimited by two edges (B1 i, B2>) of dimension Lai, spaced hi apart.

[0066] The lateral faces (c, di) are parallel to each other and of the same dimensions.

[0067] The face (c) is delimited by two edges (C1 i, C2 ) of dimension Law, spaced hi apart.

[0068] The face (di) is delimited by two edges (D1 i, D2>) of dimension Law, spaced hi apart.

[0069] The upper (e) and lower (f) faces are parallel to each other and of the same dimensions. They form the surfaces framing the hollowed-out part, carved into plate X.

[0070] The face (e) is delimited by the edges (A1 i, B1 i, C1 i, D1) forming a rectangle of width Lai and length Loi.

[0071] The face (f) is delimited by the edges (A2i, B2i, C2i, D2>) forming a rectangle of width Lai and length Loi.

[0072] The height hi corresponds to the thickness of the plate X, and thus to the height of the subunit U and its hollowed-out part.

[0073] Two subunits of the chamber can be of different heights when their hollowed-out part are respectively in two different YXY plate sequences.

[0074] The height hi is 25 pm to 20 mm.

[0075] The 25 µm to 20 mm range includes the following ranges: 25 to 50 µm; 50 to 75 µm; 75 to 100 µm; 100 to 200 µm; 200 to 300 µm; 300 to 400 µm; 400 to 500 µm; 500 to 600 µm; 600 to 700 µm; 700 to 800 µm; 800 to 900 µm; 900 µm to 1.0 mm; 1.0 to 2.0 mm; 2.0 to 3.0 mm; 3.0 to 4.0 mm; 4.0 to 5.0 mm; 5.0 to 6.0 mm; 6.0 to 7.0 mm; from 7.0 to 8.0 mm; from 8.0 to 9.0 mm; from 9.0 to 10.0 mm; from 10.0 to 11.0 mm; from 11.0 to 12.0 mm; from 12.0 to 13.0 mm; from 13.0 to 14.0 mm; from 14.0 to 15.0 mm; from 15.0 to 16.0 mm; from 16.0 to 17.0 mm; from 17.0 to 18.0 mm; from 18.0 to 19.0 mm; from 19.0 to 20.0 mm.

[0076] Advantageously the height hi is from 25 to 200 pm.

[0077] Advantageously the height hi is from 1.0 to 5.0 mm.

[0078] The dimensions of the subunits are independent of each other; that is, the width Lai, the length Loi, and the height hi of each subunit U can be identical or different. Similarly, the number of inputs ni and the number of outputs mi of the subunits Ui can be identical or different.

[0079] Thus, for the "s" subunits, these dimension parameters and numbers of inputs and outputs (Lai, Loi, hi, ni, mi) of each subunit Ui, i being an integer ranging from 1 to s, can be adapted and configured in the cell to optimize the purification or separation of a product in an initial solution.

[0080] The term "inlet" refers to a passage allowing the flow of a liquid fluid from the outside to the inside of a system containing a closed hollow section configured to contain said fluid, for example, the subunits of an electrophoresis chamber. The term "outlet" refers to a passage allowing the flow of a liquid fluid from the inside to the outside of a system containing a closed hollow section configured to contain said fluid, for example, the U subunits of an electrophoresis chamber.

[0081] On face (ai), the ni inputs into the hollowed-out portion of a U subunit of the electrophoresis chamber are distributed successively, respectively referenced Ei(1) to Ei(n), i.e., Ei(1), Ei(2) to E(ni-1), Ei(n). The number of ni inputs varies from 4 to 9, i.e., 4, 5, 6, 7, 8, and 9, preferably 5 or 6.

[0082] The entries Ei(1) to Ei(n) are located between the edges A1 i and A2i and aligned along a direction parallel to A1 i and A2i, advantageously at a substantially equal distance from each other.

[0083] By "aligned" we mean that the inlets, and respectively the outlets, are spatially close to the same straight line. The inlets can therefore be positioned slightly in front of or behind the face (ai) and / or slightly above or below each other. The positioning of the inlets is configured to introduce the different fluxes into the U subunit of the electrophoresis chamber in such a way as to allow their circulation within said U subunit parallel to the edge C1 i.

[0084] On face (b), the outputs mi are distributed successively, respectively referenced Si(1) to Si(rrii), i.e. Si(1), Si(2) to Si(mi-1), Si(nrii). The number of outputs mi varies from 4 to 12, i.e. 4, 5, 6, 7, 8, 9, 10, 11 and 12, preferably 5 or 7.

[0085] The entries Si(1) to Si(rrii) are located between the edges B1 i and B2i and aligned along a direction parallel to B1 i and B2i, advantageously at a substantially equal distance from each other.

[0086] The entries Ei(1) and Ei (n) are located near the two ends of the edges A1 i and A2i, that is to say as close as possible to the faces (c) or (d).

[0087] The outputs Si(1) and Si (m) are located near the two ends of the edges B1 i and B2i, that is to say as close as possible to the faces (c) or (d).

[0088] Thus, for each subunit U, the inlet Ei(1) is configured for the introduction of a liquid cathode CAL(i). The outlet Si(1) is configured for the evacuation of said liquid cathode after circulation in the subunit U of the electrophoresis chamber.

[0089] The inlet Ei(n) is configured for the introduction of a liquid anode ANL(i). The outlet Si(rrii) is configured for the evacuation of said liquid anode after circulation in the subunit U of the electrophoresis chamber.

[0090] The inlets and outlets of a subunit U of the electrophoresis chamber are configured so that Ei(1) faces Si(1) and Ei(n) faces Si(rrii), thereby inducing the electric field CE(i) in the subunit U of the electrophoresis chamber between the liquid anode ANL(i) and the liquid cathode CAL(i) during fluid circulation, thus enabling the implementation of an electrophoresis step. The liquid anode and liquid cathode are electrolyte solutions, i.e., solutions containing ions.

[0091] By "liquid cathode" we mean an electrolyte solution configured to act as a cathode during electrophoresis.

[0092] The term "liquid anode" refers to an electrolyte solution configured to act as an anode during electrophoresis.

[0093] In operation during fluid circulation, the presence of the liquid cathode CAL(i) and the liquid anode ANL(i) allows an electric field to be generated in the U subunit of the electrophoresis chamber when the electrolytic solutions are charged.

[0094] The liquid cathode and liquid anode are electrolytic solutions, capable of generating an electric field, which can be increased or decreased by the action of the generator which charges the liquid anode and liquid cathode and / or with an increase in the concentration of ions.

[0095] Thus the electric fields generated CE(i) in the U subunits can be identical or different from one subunit to another depending on the liquid anode and the liquid cathode in circulation.

[0096] Function of the inputs and outputs of the Ui subunit

[0097] In the first subunit Ui, one of the inlets Ei (2) to Ei (ni-1) is configured for the introduction of the initial solution to be purified and / or separated into the first subunit Ui of the electrophoresis chamber.

[0098] At least one of the inputs Ei (2) to Ei (ni-1), distinct from the aforementioned input of the initial solution to be purified and / or separated, is configured for the introduction of at least one buffer solution into the subunit Ui in order to allow free-flow electrophoresis from the initial solution in the presence of at least one buffer solution under an electric field CE(1) generated by the circulation of the liquid cathode and anode.

[0099] This subunit Ui of ni entries therefore contains (ni-3) remaining entries dedicated to the introduction of one or more buffer solutions of identical or different compositions, in addition to the entries dedicated to the liquid electrodes Ei (1 ) and Ei(ni) and that dedicated to the introduction of the initial solution to be purified and / or separated.

[0100] The outlets Si (1 ) and Si(mi) are dedicated to the respective circulation of the two liquid electrodes.

[0101] At least one of the outputs Si(2) to Ei(mi-1), named SVi(Ui) is dedicated to the intermediate channel Vi linking the subunit Ui to the subunit U2.

[0102] At least one of the remaining outlets is dedicated either to the recovery of a stream of one of the compounds present in the product to be purified and / or separated, or to the evacuation of a buffer solution that has circulated in the Ui subunit.

[0103] Function of the inputs and outputs of subunit U

[0104] In each subunit U different from U1 and U s , at least one of the inputs Ei(2) to Ei(ni-1) is connected at least to the intermediate channel V-1 which allows to recover a part of the flux, from the subunit UM, containing the purified and / or separated product or a mixture of the compounds constituting said product and to introduce said part into the subunit Ui, without presence of electric field, as a solution to be purified and / or separated in order to be able to carry out a polishing step, namely a purification and / or separation step, by electrophoresis in the subunit Ui.

[0105] At least one other of the inputs Ei(2) to Ei(ni-1), distinct from the aforementioned input connected to the VM channel, is configured for the introduction of at least one buffer solution into the subunit Ui in order to permit free-flow electrophoresis from said solution to be purified and / or separated in the presence of at least one buffer solution under an electric field CE(i) generated by the circulation of the liquid cathode CAL(i) and the liquid anode (ANL(i).

[0106] This subunit Ui of ni entries therefore contains (ni-3) remaining entries dedicated to the introduction of one or more buffer solutions of identical or different compositions, in addition to the entries dedicated to the liquid electrodes Ei(1) and Ei(n) and that dedicated to the VM channel.

[0107] The outlets Si(1) and Si(rrii) are dedicated to the respective circulation of the two liquid electrodes.

[0108] At least one of the outputs Si(2) to Ei(mi-1), named S (Ui) is dedicated to the intermediate channel linking the subunit Ui to the subunit +i.

[0109] At least one of the remaining outlets is dedicated either to the recovery of a stream of one of the compounds present in the product to be purified and / or separated, or to the evacuation of a buffer solution that has circulated in the Ui subunit.

[0110] Function of the inputs and outputs of subunit U s

[0111] In subunit U s , at least one of the E entries s (2) to E s (n s -1) is connected at least to the intermediate channel V s -i which allows recovery of a portion of the stream containing the purified and / or separated product or a mixture of the compounds constituting said product, said portion originating from subunit U s -i which has been purified and / or separated and (s-2) times polished and introduced into the subunit U s, without the presence of an electric field, as a solution to be purified and / or separated in order to be able to carry out a polishing step (purification and / or separation step) by electrophoresis.

[0112] At least one other of the E entries s (2) to E s (n s -1), distinct from the aforementioned input connected to channel V s -i, is configured to introduce at least one buffer solution into the U subunit s in order to allow free-flow electrophoresis from said solution to be purified and / or separated in the presence of at least one buffer solution under an electric field CE(s) generated by the circulation of the cathode CAL(s) and the anode (ANL(s) liquids.

[0113] This subunit U s of n s entries therefore contain (n s-3) remaining inlets dedicated to the introduction of one or more buffer solutions of identical or different compositions, in addition to the inlets dedicated to the liquid electrodes E s (1) and E s (n s ) and that dedicated to the intermediate channel V s -i.

[0114] In the last subunit U s of the electrophoresis cell, at least one of the S outputs s (2) to S s (m s - 1) is configured for the recovery of the purified and / or separated solution after the electrophoresis process carried out in the electrophoresis chamber. The S outputs s (1) and S s (m s ) are dedicated to the respective circulation of the two liquid electrodes.

[0115] In the electrophoresis cell of the invention, as described above, the inputs and outputs of each subunit have distinct functions:

[0116] - The inlets and outlets at the ends are configured to allow the circulation of liquid electrodes to generate an electric field across the entire subunit,

[0117] - one entry is dedicated to the introduction of the solution to be purified and / or separated, or said to be polished,

[0118] - the other input(s) are dedicated to the introduction of one or more buffer solutions,

[0119] - in the subunits Ui to U s -i, at least one of the outlets, separate from those of the electrolytes, is dedicated to recovering a portion of the flow containing the solution to be purified and / or separated in the next subunit,

[0120] - in all subunits at least one of the outlets, separate from that of the electrolytes, is dedicated either to the recovery of a compound from the product to be purified and / or separated or to the evacuation of the buffer solution after circulation,

[0121] - in subunit U sat least one of the outlets, separate from that of the electrolytes, is dedicated to recovering the purified and / or separated product or one of the compounds of said product and another outlet is dedicated to evacuating the buffer solution.

[0122] The cell according to the invention may include supply channels for the liquid electrodes (anode and cathode), one or more buffer solutions, a supply channel for the product to be purified and / or separated into Ui cells, and at least one outlet recovery channel in U s .

[0123] According to a particular embodiment, the present invention relates to a cell as defined above, comprising: supply channels configured to connect the s inputs Ei(1), i varying from 1 to s, of each subunit of the electrophoresis chamber to at least one external micro / millifluidic circuit for supplying at least one liquid cathode; supply channels configured to connect the s inputs Ei(n) of each subunit of the electrophoresis chamber to at least one external micro / millifluidic circuit for supplying at least one liquid anode; a supply channel configured to connect at least one of the inputs Ei(2) to Ei(ni-1) of the subunit Ui to an external micro / millifluidic circuit for supplying an initial solution containing a product to be purified and / or separated.supply channels configured to connect at least one input of each subunit of the electrophoresis chamber to at least one external micro / milli fluidic circuit supplying at least one buffer solution, at least one recovery channel configured to connect one of the outputs of subunit U, s chosen from S s (2) to S s (m s -1) from the electrophoresis chamber to an external micro / milli fluidic recovery circuit.

[0124] The inlets and outlets of the electrophoresis chamber are those connected to the outside of the cell, meaning those not linked to intermediate channels connecting the subunits. These inlets and outlets are connected via supply, recovery, or discharge channels to external supply, recovery, or discharge circuits configured for micro- or millifluidic fluid flow.

[0125] The term "supply channel" refers to a channel connected to an inlet, adapted to supply a subunit of the chamber with flow.

[0126] The term "recovery channel" refers to a channel connected to an output, adapted to recover at least one stream of interest, namely one containing the product or one of its compounds.

[0127] The term "evacuation channel" refers to a channel connected to an outlet, adapted to evacuate a flow that has circulated in the chamber.

[0128] The term “microfluidic circuit” refers to a set of channels with a cross-sectional area on the order of a micrometer.

[0129] A "millifluidic circuit" is defined as a set of channels with cross-sectional dimensions on the order of a millimeter. These supply, recovery, or discharge channels are advantageously partially or totally etched into the X-ray plates.

[0130] Supply and disposal channels for liquid electrolytes

[0131] The electrophoresis cell containing s subunits comprises s inputs Ei(1) and s outputs Si(1), i ranging from 1 to s.

[0132] Each input Ei(1) is linked to a supply channel for a liquid cathode CAL(i). These s supply channels are connected to at least one external supply circuit for at least one solution constituting the liquid cathode. Other external supply circuits for a liquid cathode, up to a number s, can be linked to these inputs when different electric fields CE(i) are required.

[0133] Each outlet Si(1) is linked to a liquid cathode evacuation channel CAL(i).

[0134] The electrophoresis cell containing s subunits comprises s inputs Ei(n) and s outputs Si(rrii), i ranging from 1 to s.

[0135] Each input Ei(n) is linked to a supply channel of a liquid anode ANL(i). These s supply channels are connected to at least one external supply circuit of at least one solution constituting the liquid anode. Other external supply circuits for a liquid anode, up to a number s, can be linked to these inputs when different electric fields CE(i) are required.

[0136] Each outlet Si(rrii) is linked to a liquid anode evacuation channel ANL(i).

[0137] Supply channels for buffer solution(s) for electrophoresis

[0138] In the subunit Ui, at least one of the inputs Ei(2) to Ei(ni-1) is connected via a supply channel to an external micro / millifluidic circuit supplying an initial solution containing a product to be purified and / or separated. The remaining (ni-3) inputs, with at least one remaining input ni greater than 4, are connected via supply channels to one or more buffer solution(s).

[0139] In the other subunits U2 to U s , the (ni-3) inputs of the U subunit which are distinct from the two inputs located respectively at the ends and distinct from the input dedicated to the intermediate channel, are respectively linked by supply channels to one or more buffer solution(s).

[0140] Product or product compound recovery channels or buffer solution evacuation channels in the last subunit U s , at least one of the S outputs s (2) to S s (ms -1) is linked to a recovery channel connected to an external micro / millifluidic circuit for recovering the purified and / or separated product, and at least one of these outlets is linked to a discharge channel for a buffer solution that has circulated in U s .

[0141] In the other subunits U1 to U s -i, for (rrii-3) outputs of the subunit U which are distinct from the two outputs located respectively at the ends and distinct from the output dedicated to the intermediate channel connecting the subunits, at least one of these outputs is: either linked to a recovery channel, advantageously to recover a purified and / or separated compound constituting said product, or linked to a channel for the removal of a buffer solution which has circulated in U.

[0142] These supply, recovery and evacuation channels fulfill the distinct function of each input and output of each subunit described above.

[0143] The cell advantageously contains supply and discharge channels (distinct from those of the liquid electrodes) which are present for at least one buffer solution so as to effectively allow the separation and / or purification by free-flow electrophoresis of the product to be purified and / or separated contained in a solution.

[0144] According to a particular embodiment, the present invention relates to a cell as defined above, said cell being configured to: in each subunit Ui, in the presence of an electric field generated CE(i) between a liquid cathode CAL(i) and a liquid anode ANL(i) parallel to A1 i and perpendicular to C1 i, and during operation, a) in subunit Ui: circulate the liquid cathode CAL(1) from inlet Ei(1) to outlet Si(1), circulate the liquid anode ANL(1) from inlet Ei(ni) to outlet Si(mi), circulate the initial solution containing the product to be separated and / or purified and at least one buffer solution in subunit Ui, between the liquid cathode CAL(1) and the liquid anode ANL(1), from inlets Ei(2) to Ei(ni-1) to outlets Si(2) to Si(mi-1), to obtain the product purified and / or separated from the Ui subunit, b) in each intermediate channel Vk, k varying from 1 to (s-1),to recover at the SVk(Uk) outlet the purified and / or separated product from the Uk subunit; to circulate the aforementioned product from the SVk(Uk) output of the Uk subunit through channel Vk, from SVk(Uk) to the EVk(Uk+i) input of the Uk+i subunit; c) in the Uk+i subunit: to circulate the liquid cathode CAL(k+1) from the Ek+i (1) input to the Sk+i (1) output; to circulate the liquid anode ANL(k+1) from the Ek+i(nk+i) input to the Sk+i(rrik+i) output; to circulate the purified and / or separated product from the SVk(Uk) output of the Uk subunit and at least one buffer solution in the Uk+i subunit, between the liquid cathode CAL(k+1) and the liquid anode ANL(k+1), from the Ek+i(2) to Ek+i(nk+i-1) inputs to the outputs Sk+i(2) to Sk+i(mk+i- 1), d) retrieve at one of the outputs S, s (2) to S s (m s -1) the purified and / or separated product.

[0145] The Ui subunit is configured to perform a first step of separation of the product to be separated and / or purified in an initial solution by free-flow electrophoresis under an electric field CE(1) generated by the circulation of the liquid cathode and liquid anode and in the presence of at least one buffer solution.

[0146] The following successive subunits are configured to refine the separation, i.e., "polish", by successive purification and / or separation steps, called "polishing" steps, carried out by free-flow electrophoresis under an electric field CE(i) in the presence of at least one other buffer solution, on a portion of the outgoing flow from the previous subunit through an intermediate channel connecting the two subunits.

[0147] The subunits are also advantageously configured to be able to retrieve output from one of the subunits Ui different from the last subunit U s, an outflow containing one of the product compounds to be separated and / or purified and to continue the separation and / or purification in successive subunits of other outflows containing the product.

[0148] Additional intermediate channel l / l /

[0149] According to a particular embodiment, the present invention relates to a cell as defined above, further comprising: at least one complementary intermediate channel W, said complementary intermediate channel W connecting two successive subunits U q and U q +i, q being an integer from 1 to (s-1), said channel W extending: from one of the outputs of unit U q chosen from S q (2) to S q (m q -1), different from the output linked to channel Vq, to ​​the input linked to channel V q of subunit U q +i, the aforementioned channel V input q being connected to at most (m q-3) chosen exit(s) from S q (2) to S q (m q -1).

[0150] In this embodiment, the electrophoresis chamber comprises, between two successive subunits, in addition to the intermediate channel V connecting them, another complementary intermediate channel W extending from one of the outputs of the first subunit, distinct from the three outputs of the cathode, the anode, and the aforementioned intermediate channel V, to the same input as that of the aforementioned intermediate channel V of the second subunit. The complementary intermediate channel W can be located between any combination of two successive subunits of the electrophoresis chamber, q being any integer from 1 to (s-1).

[0151] This complementary intermediate channel W allows for the expansion of the collected portion of the flow in the first subunit, intended to be introduced as a solution to be purified and / or separated in the subsequent second subunit. This is particularly advantageous when the flow containing the desired product or compound extends between two adjacent outlets of the first subunit.

[0152] Advantageously, in a particular embodiment, the output corresponding to channel V may not be adjacent to that of channel W. This configuration allows recovery in an output of the first subunit located between that of channel V and that of channel W, a stream containing the purified product or a compound separated from this product and to continue in the second following subunit the purification and / or separation with the streams collected by the intermediate channels V and W containing a product not sufficiently purified or a mixture of compounds of the product.

[0153] The subunits are designed to purify and / or separate a product circulating in the flow between the two electrodes in the presence of at least one buffer solution, and the intermediate channels are designed to collect a portion of this flow circulating between the liquid electrodes. Therefore, the inlet related to channel V q of a subunit U q +i can only be connected at most to (m q -3) U exits q , m q being the number of outputs of the subunit U q , because the two outlets dedicated to electrolytes are excluded, as well as an outlet dedicated either to the recovery of a purified and / or separated product or compound, or to the evacuation of the buffer solution that has circulated in unit U q Indeed, there would be no benefit in introducing, through this inlet, the electrolyte solutions and the entire outgoing flux of U into the second subunit. q .

[0154] According to a particular embodiment, the present invention relates to a cell as defined above, further comprising:

[0155] R is an intermediate channel (or channels)^) complementary channel(s), from Wi to WR, where R is an integer from 1 to Q: as r varies from 1 to R, the complementary intermediate channel W r connects the U subunit qr to the successive subunit U qr +i, qr being an integer from 1 to (s-1), said complementary intermediate channel W r extends from: one of the outputs of unit U qr chosen from S qr (2) to S q r(m qr -1) different from the output linked to channel V qr , at the entrance to the EV q r(U q r+i) linked to channel V qr of subunit U qr +i, the aforementioned EV entry q r(U q r+i) being connected to at most (m qr -3) chosen exit(s) from S qr (2) to S qr (m q -1).

[0156] In this embodiment, the number Q represents the maximum number of complementary intermediate channels W that can be introduced. It corresponds to the total sum of the number of chamber outlets, subtracting the number of outlets of the last subunit that are not connected to intermediate channels and subtracting, for the (s-1) spacings between two successive subunits, the four outlets dedicated respectively to the two liquid electrodes, the intermediate channel V and the recovery of a product or the evacuation of a buffer solution.

[0157] A chamber can contain R complementary intermediate channels W, with R being an integer from 1 to Q.

[0158] These complementary intermediate channels are called W r , r varying from 1 to R.

[0159] The complementary intermediate channel W r is associated with two successive subunits U qr and U qr+i, qr being any integer from 1 to (s-1), namely that there is no link between the index r of the channel and the index of the subunits qr.

[0160] This is subject, as previously stated, to the condition that the input linked to channel V qr of a subunit U qr +i can only be connected at most to (m qr -3) U exits qr , m qr being the number of outputs of the subunit U qr According to a particular embodiment, the present invention relates to a cell as defined above, comprising R complementary intermediate channel(s) Wi to WR, R being an integer from 1 to (s-1), and in which said channels Wi to W each connect two different successive subunits.

[0161] In this embodiment, the W channels of the electrophoresis chamber each connect two different successive subunits, so the chamber includes at most one W channel per two successive units.

[0162] Cell of subunits of the same height

[0163] According to a particular embodiment, the present invention relates to a cell as defined above, comprising a plate X between two adjacent plates Y, said plate X comprising the hollowed parts of the subunits U of said electrophoresis chamber, the height of the hollowed part of each subunit being of identical value.

[0164] In this embodiment, since all the hollowed parts of the subunits are located on the same plate and have the same height h, the electrophoresis cell comprises only one plate X of height h.

[0165] Cell of subunits of different heights

[0166] According to a particular embodiment, the present invention relates to a cell as defined above, comprising at least two plates X, each plate X being between two adjacent plates Y, each plate X comprising at least a hollowed-out portion of a subunit U of said electrophoresis chamber.

[0167] This particular embodiment corresponds to a cell configuration in which at least two subunits of the electrophoresis chamber are not on the same plate and may have different heights (hi). The cell thus comprises at least two different YXY sequences, with X plates that may be of different heights.

[0168] According to a particular embodiment, the present invention relates to a cell as defined above, comprising a plate X between two adjacent plates Y, said plate X comprising the hollowed parts of the subunits U of said electrophoresis chamber, the height of the hollowed part of each subunit being of identical value, or comprising at least two plates X, each plate X being between two adjacent plates Y, each plate X comprising at least one hollowed part of a subunit U of said electrophoresis chamber.

[0169] Z. cooling plates

[0170] According to a particular embodiment, the present invention relates to a cell as defined above, in which said a cooling circuit comprises at least one cooling plate Z comprising a heat transfer system, said plate Z being adjacent to one of the two plates Y adjacent to said plate X.

[0171] According to a particular embodiment, the present invention relates to a cell as defined above, in which said a cooling circuit comprises at least two plates Z comprising a heat transfer system, said cell comprising the following sequence of plates ZYXYZ, in particular the sequence YZYXYZY.

[0172] According to a particular embodiment, the present invention relates to a cell as defined above, in which said heat transfer system is a network of pipes configured to allow the circulation of one or more heat transfer fluids parallel to the edges Ci of the subunits Ui.

[0173] In this embodiment, the heat transfer fluid flows in the same direction as the electrophoresis flow within the chamber, facilitating temperature control, which is important for controlling electrophoretic mobility. Advantageously, the piping network can be adapted to generate a temperature gradient.

[0174] According to a particular embodiment, the present invention relates to a cell as defined above, in which said heat transfer system is a network of pipes configured to allow the circulation of one or more heat transfer fluids perpendicular to the edges Ci of the subunits Ui.

[0175] In this embodiment, the heat transfer fluid is perpendicular to the direction of the electrophoresis flow in the chamber. In this configuration, the inlets and outlets are perpendicular to the inlet and outlet alignments of the subunits, simplifying the fluid circuit layout of the cell by distributing the inlets and outlets.

[0176] According to a particular embodiment, the present invention relates to a cell as defined above, in which said heat transfer system is a network of pipes configured to allow the circulation of one or more heat transfer fluids parallel to the edges Ci or perpendicular to the edges Ci of the subunits Ui, in particular said network of pipes comprising inlets and outlets of the heat transfer system, engraved in the plate Z.

[0177] According to a particular embodiment, the present invention relates to a cell as defined above, comprising at least one cooling plate Z comprising a heat transfer system, said plate Z being adjacent to one of the two plates Y adjacent to said plate X, and / or comprising at least two plates Z comprising a heat transfer system, said cell comprising the following sequence of plates ZYXYZ, in particular the sequence YZYXYZY, in particular wherein said heat transfer system is a network of pipes configured to permit the circulation of one or more heat transfer fluids parallel to the edges Ci or perpendicular to the edges Ci of the subunits Ui, in particular said network of pipes comprising inlets and outlets of the heat transfer system, engraved in the plate Z.

[0178] Channeling methods

[0179] According to a particular embodiment, the present invention relates to a cell as defined above, in which the subunits Ui of the electrophoresis chamber comprise means for channeling the flows opening onto the inlets Ei(1) to Ei(n) and / or onto the outlets Si(1) to Si(rrii) of the subunits Ui of the electrophoresis chamber, said channeling means being preferably etched into the electrophoresis plate X, said means being in particular triangular in shape, located between two adjacent inlets or two adjacent outlets of a subunit.

[0180] These channeling means are configured to allow the orientation of flows in the subunit of the electrophoresis chamber at the level of each of the inlets and outlets, in order to better distribute the flows in the case of the inlets over the entire width of the hollowed part and to better concentrate the flows in the case of the outlets.

[0181] Advantageously, these channeling means are an integral part of the electrophoresis chamber subunit, that is, fused with the walls of the electrophoresis chamber subunit and made of the same material as the electrophoresis plate. These channeling means are located along faces (ai) and (b) of the rectangular parallelepiped in which the hollowed-out portion of subunit Ui is inscribed.

[0182] The means of channeling are, for example, triangular or beveled point-shaped elements located between two inlets or two outlets.

[0183] Dimensions of subunits

[0184] According to a particular embodiment, the present invention relates to a cell as defined above, in which the width Lai of each of the s subunits Ui is from 1.0 to 8.0 cm, and / or the length Loi of each of the s subunits U of the electrophoresis chamber is from 2.0 to 20.0 cm.

[0185] The range of "1.0 to 8.0 cm" includes the ranges: from 1.0 to 2.0 cm; from 2.0 to 3.0 cm; from 3.0 to 4.0 cm; from 4.0 to 5.0 cm; from 5.0 to 6.0 cm; from 6.0 to 7.0 cm; from 7.0 to 8.0 cm.

[0186] The "2.0 to 20.0 cm" range includes the following ranges: 2.0 to 3.0 cm; 3.0 to 4.0 cm; 4.0 to 5.0 cm; 5.0 to 6.0 cm; 6.0 to 7.0 cm; 7.0 to 8.0 cm; 8.0 to 9.0 cm; 9.0 to 10.0 cm; 10.0 to 11.0 cm; 11.0 to 12.0 cm; 12.0 to 13.0 cm; 13.0 to 14.0 cm; 14.0 to 15.0 cm; 15.0 to 16.0 cm; 16.0 to 17.0 cm; 17.0 to 18.0 cm; from 18.0 to 19.0 cm; from 19.0 to 20.0 cm

[0187] X-Plate Material

[0188] According to a particular embodiment, the present invention relates to a cell as defined above, in which the X-ray electrophoresis plate is made of a material selected from polytetrafluoroethylene (PTFE), perfluoroalkoxy (PFA) and fluoroethylene propylene (FEP), in particular Teflon™, Teflon™-PFA and Teflon™-FEP plates.

[0189] According to a particular embodiment, the present invention relates to a cell as defined above, wherein the U subunits of the electrophoresis chamber comprise means for channeling the fluxes exiting at the inlets Ei(1) to Ei(n) and / or at the outlets Si(1) to Si(r1i) of the U subunits of the electrophoresis chamber, said channeling means preferably being etched into the electrophoresis plate X, said means being in particular triangular in shape, located between two adjacent inlets or two adjacent outlets of a subunit, and / or wherein the width Lai of each of the s U subunits is from 1.0 to 8.0 cm, and / or the length Loi of each of the s U subunits of the electrophoresis chamber is from 2.0 to 20.0 cm, and / or wherein the electrophoresis plate X is made of a material selected from polytetrafluoroethylene (PTFE), perfluoroalkoxy (PFA) and fluoroethylene propylene (FEP), in particular Teflon™ sheets,Teflon™-PFA and Teflon™-FEP.

[0190] Two-subunit electrophoresis cell

[0191] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is 2, the electrophoresis chamber comprising two subunits U1 and U2, the subunit U1 comprising a hollowed-out portion in a plate X closed by two plates Y

[0192] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, ci, di) and 2 upper and lower faces (ei, fi),

[0193] ■ said hollowed-out part being of length L01 and width Lai,

[0194] ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0195] ■ the lateral faces (ai , bi) being parallel to each other, the face (ai) being delimited by two edges (A1 i, A2i) of dimension Lai and the face (bi) being delimited by two edges (B11 , B2i) of dimension Lai ,

[0196] ■ the lateral faces (ci, di) being parallel to each other, the face (ci) being delimited by two edges (C11, C2i) of dimension L01 and the face (di) being delimited by two edges (D11, D2i) of dimension L01, o ni successive entries Ei(1), Ei(2) to Ei(ni-1), Ei(ni), ni being an integer from 4 to 9, distributed on the face (ai) between A11 and A2i and aligned along a direction parallel to A11 and A2i, or successive exits from Si (1), Si(2) to Si(mi-1), Si(mi), r being an integer from 4 to 12, distributed on the face (bi) between B1i and B2i and aligned along a direction parallel to B1i and B2i, such that Si(1) faces Ei(1) and Si(mi) faces Ei(ni) along a direction parallel to C1 i and D1 i, the subunit U2 comprises o a hollowed-out part in a plate X, possibly in the aforementioned plate X of the aforementioned subunit Ui,

[0197] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (a2, b2, C2, d2) and 2 upper and lower faces (e2, f2),

[0198] ■ said hollowed-out part being of length L02 and width La2,

[0199] ■ of height i2 corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0200] ■ the lateral faces ai, b2) being parallel to each other, the face (32) being delimited by two edges (AI2, A22) of dimension La2 and the face (b2) being delimited by two edges (BI2, B22) of dimension La2,

[0201] ■ the lateral faces (C2, d2) being parallel to each other, face (C2) being delimited by two edges (CI2, C22) of dimension L02 and face (d2) being delimited by two edges (DI2, D22) of dimension L02, o n2 successive inputs E2(0), E2(2) to E2(n2-1), E(n2), n2 being an integer from 4 to 9, distributed on face (32) between AI2 and A22 and aligned along a direction parallel to AI2 and A22, o m2 successive outputs from S2(1), S2(2) to 82(012-1), 82(012), m2 being an integer from 4 to 12, distributed on face (b2) between BI2 and B22 and aligned along a direction parallel to BI2 and B22, so that 82(1) faces E2O) and 82(012) face £2(02) in a direction parallel to CI2 and DI2, characterized in that it comprises an intermediate channel V1, connecting the two subunits U1 and U2 by one of the outputs SVi(Ui) chosen from Si(2) to Si(mi-1) of the subunit U1 to one of the inputs EVI(U2) of the subunit U2 chosen from E2(2) to E2(n2-1).

[0202] The cell according to the invention may comprise two subunits U1 and U2, supply and discharge channels, and a U-shaped outlet recovery channel. s .

[0203] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is 2, comprising: a supply channel configured to connect the inlet Ei(1) of the electrophoresis chamber subunit U1 to an external micro / millifluidic circuit for supplying a first liquid cathode CAL(1), a supply channel configured to connect the inlet Ei(ni) of the electrophoresis chamber subunit U1 to an external micro / millifluidic circuit for supplying a first liquid anode ANL(1), a supply channel configured to connect at least one of the inlets Ei(2) to E1 (ni-1) of the U1 subunit to an external micro / millifluidic circuit for supplying an initial solution containing a product to be purified and / or separated,at least one supply channel configured to connect at least one remaining input of unit Ui to at least one external micro / milli-fluidic circuit supplying at least one buffer solution, a supply channel configured to connect the E2(n2) input of the electrophoresis chamber subunit U2 to an external micro / milli-fluidic circuit supplying a second liquid cathode CAL(2), a supply channel configured to connect the E2(n2) input of the electrophoresis chamber subunit U2 to an external micro / milli-fluidic circuit supplying a second liquid anode ANL(2), at least one supply channel configured to connect at least one remaining input of unit U2 to at least one external micro / milli-fluidic circuit supplying at least one buffer solution,at least one recovery channel configured to connect one of the outputs of subunit U2 selected from 82(2) to 82(012-1) of the electrophoresis chamber to an external micro / millifluidic recovery circuit, said cell being configured to, in the presence of: o an electric field generated CE(1) in subunit Ui between liquid cathode CAL(1) and liquid anode ANL(1) parallel to A1 i and perpendicular to C1 i, o an electric field generated CE(2) in subunit U2 between liquid cathode CAL(2) and liquid anode ANL(2) parallel to AI2 and perpendicular to CI2, o and in operation, a) in subunit U1: o circulate liquid cathode CAL(1) from inlet E1 (1 ) to outlet Si (1 ) , o circulate liquid anode ANL(1) from inlet Ei(ni) at the outlet Si(mi), to circulate the initial solution containing the product to be separated and / or purified and at least one buffer solution in the subunit Ui,between the liquid cathode CAL(1) and the liquid anode ANL(1), from the inlets Ei(2) to Ei(ni-1) to the outlets Si(2) to Si(mi-1), b) in the intermediate channel Vi, to recover at the outlet SVi(Ui) the purified and / or separated product contained in said initial solution; a) Circulate the purified and / or separated product from the SVi(Ui) outlet of subunit Ui in channel V1, from SVi(Ui) to the EVI(U2) inlet of subunit U2; c) in subunit U2: circulate the liquid cathode CAL(2) from the E2(0) inlet to the S2(1) outlet; circulate the liquid anode ANL(2) from the E2(n2) inlet to the 82(012) outlet; circulate the purified and / or separated product from the SVi(Ui) outlet of subunit U1 and at least one buffer solution in subunit U2, between the liquid cathode CAL(2) and the liquid anode ANL(2), from inlets E2(2) to E2(n2-1) to outlets 82(2) to 82(012-1); d) collect at one of the outlets 82(2) to 82(012-1) the purified and / or separated product.

[0204] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is 2, the number of outputs r of the unit U1 is greater than or equal to 5, further comprising an intermediate channel W1 connecting the subunit U1 to the subunit U2, extending: o from one of the outputs SWi(Ui) of the unit U1 chosen from Si(2) to Si(mi-1), different from the output SVi(Ui) of the channel V1, o to the input EVI(U2) of the subunit U2.

[0205] In addition to a portion of the flow from subunit U1 collected by channel V1 and directed to subunit U2 as a solution to be purified and / or separated, channel W1 allows another portion of the flow collected in subunit U1 to be directed to complete the solution to be purified and / or separated in subunit U2.

[0206] Ratio of subunit dimensions and number of inputs and outputs.

[0207] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 2, in which the number of outputs r and m2 of the subunits U1 and U2 is greater respectively than the number of inputs and m of the subunits U1 and U2.

[0208] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is 2 and in which the subunits U1 and U2 have width and length dimensions such that the width La2 of subunit U2 is equal to the width Lai of subunit U1 and the length L02 of subunit U2 is equal to the length L01 of subunit Ui.

[0209] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 2 and in which the width La2 of subunit U2 is greater than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is less than or equal to the length L01 of subunit Ui. According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 2 and in which the width La2 of subunit U2 is less than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is greater than or equal to the length L01 of subunit Ui.

[0210] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 2 and in which the width La2 of the subunit U2 is less than or equal to the width Lai of the subunit U1 and the length L02 of the subunit U2 is less than or equal to the length L01 of the subunit Ui.

[0211] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 2 and in which the width La2 of the subunit U2 is greater than or equal to the width Lai of the subunit U1 and the length L02 of the subunit U2 is greater than or equal to the length L01 of the subunit Ui.

[0212] According to a particular embodiment, the present invention relates to a cell as defined above, in which the number of outputs r and m2 of the subunits U1 and U2 is greater respectively than the number of inputs ni and n2 of the subunits U1 and U2; and / or in which the subunits U1 and U2 have width and length dimensions such that the width La2 of subunit U2 is equal to the width Lai of subunit U1 and the length L02 of subunit U2 is equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is greater than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is less than or equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is less than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is greater than or equal to the length L01 of subunit Ui,or in which the width La2 of subunit U2 is less than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is less than or equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is greater than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is greater than or equal to the length L01 of subunit U1.

[0213] Electrophoresis cell with a chamber of 3 subunits.

[0214] According to a particular embodiment, the present invention relates to a cell as defined above, in which s is equal to 3, the electrophoresis chamber comprising three subunits U1, U2 and U3, the subunit U1 comprising a portion hollowed out in a plate X

[0215] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, ci, di) and 2 upper and lower faces (ei, fi),

[0216] ■ said hollowed-out part being of length L01 and width Lai,

[0217] ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0218] ■ the lateral faces (ai , bi) being parallel to each other, the face (ai) being delimited by two edges (A1 i, A2i) of dimension Lai and the face (bi) being delimited by two edges (B11 , B2i) of dimension Lai ,

[0219] ■ the lateral faces (ci, di) being parallel to each other, the face (ci) being delimited by two edges (C11, C2i) of dimension L01 and the face (di) being delimited by two edges (D11, D2i) of dimension L01, o ni successive inputs Ei(1), Ei(2) to Ei(ni-1), Ei(ni), ni being an integer from 4 to 9, distributed on the face (ai) between A11 and A2i and aligned along a direction parallel to A11 and A2i, o mi successive outputs from Si (1), Si(2) to Si(mi-1), Si(mi), mi being an integer from 4 to 12, distributed on the face (bi) between B1i and B2i and aligned along a direction parallel to B1i and B2i, so that Si(1) faces Ei(1) and Si(mi) faces Ei(ni) along a direction parallel to C1 i and D1 i, the subunit U2 comprises o a hollowed-out part in a plate X

[0220] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, b2, C2, d2) and 2 upper and lower faces (e2, f2),

[0221] ■ said hollowed-out part being of length L02 and width La2,

[0222] ■ of height ti2 corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0223] ■ the lateral faces ai, b2) being parallel to each other, the face (32) being delimited by two edges (AI2, A22) of dimension La2 and the face (b2) being delimited by two edges (BI2, B22) of dimension La2,

[0224] ■ the lateral faces (C2, d2) being parallel to each other, face (C2) being delimited by two edges (CI2, C22) of dimension L02 and face (d2) being delimited by two edges (DI2, D22) of dimension L02, o n2 successive inputs E2(0), E2(2) to E2(n2-1), E(n2), n2 being an integer from 4 to 9, distributed on face (32) between AI2 and A2s and aligned along a direction parallel to AI2 and A22, o m2 successive outputs from S2(1), S2(2) to 82(012-1), 82(012), m2 being an integer from 4 to 12, distributed on face (b2) between BI2 and B22 and aligned along a direction parallel to BI2 and B22, such that 82(1 ) faces E2O ) and 82(012) faces £2(02) in a direction parallel to CI2 and DI2, the subunit U3 comprises o a hollowed-out part in a plate X

[0225] ■ in the shape of a rectangular parallelepiped with 4 lateral faces (33, bs, C3, ds) and 2 upper and lower faces (es, fs),

[0226] ■ said hollowed-out part being of length Los and width Las,

[0227] ■ of height hs corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm,

[0228] ■ the lateral faces (33, bs) being parallel to each other, the face (33) being delimited by two edges (Aïs, A2s) of dimension Las and the face (bs) being delimited by two edges (Bis, B2s) of dimension Las,

[0229] ■ the lateral faces (es, ds) being parallel to each other, the face (es) being delimited by two edges (CI3, C2s) of dimension L03 and the face (ds) being delimited by two edges (DI3, D2s) of dimension L03, o ns successive inputs Es(1), Es(2) to Es(ns-1), E(ns), ns being an integer from 4 to 9, distributed on the face (33) between AI3 and A2s and aligned along a direction parallel to AI3 and A23, o m3 successive outputs from 83(1), Ss(2) to Ss(ms-1), Ss(ms), m3 being an integer from 4 to 12, distributed on the face (bs) between Bis and B2s and aligned along a direction parallel to Bis and B2s, so that Ss(1) faces Es(1) and Ss(ms) faces Es(ns) in a direction parallel to CI3 and Dis, characterized in that it comprises an intermediate channel V1, connecting the two subunits U1 and U2 by one of the outputs SVi(Ui) chosen from Si(2) to Si(mi-1) of subunit U1 to one of the inputs EVI(U2) of subunit U2 chosen from Es(2) to E2(n2-1), and an intermediate channel V2,connecting the two subunits U2 and U3 via one of the outputs SV2(U2) chosen from S2(2) to S2(rri2-1) of subunit U2 to one of the inputs EV2OJ3) of subunit U3 chosen from Es (2) to Es(n3-1).

[0230] Electrophoresis device

[0231] Another object of the invention relates to a free-flow electrophoresis cell device comprising: p plates X between two adjacent plates Y: op being an integer from 1 to 50, in particular from 1 to 20, o X being an electrophoresis plate (1) of inert material, o Y being a sealed plate (2) of sapphire or alumina Al2O3 with 99% a-AhCh, f electrophoresis cells as defined above, f is an integer from 1 to 500, clamping means for all the plates enabling the sealing of said device.

[0232] The device of the invention is an arrangement of one to several hundred electrophoresis cells, in particular from 1 to 500 electrophoresis cells, the cells being as defined above. The electrophoresis cell forms a constituent and repeating unit of the device.

[0233] The use of the sapphire Y-plate allows for a robust and watertight device that allows for disassembly and reassembly, facilitating cleaning of the device's components and its maintenance. The modular design of the device allows for the reuse of the plates.

[0234] Advantageously, the mechanical strength of sapphire allows for an assembly system that enables significant pressure from the flows within the device while ensuring excellent sealing between each stage, which is not the case for glass, which scratches and cracks very easily under slight pressure; in the presence of water, the crack in the glass can propagate throughout the cell and induce significant sealing problems.

[0235] Thus, the use of sapphire Y-plates as adjacent plates closing a fluidic circuit ensures robustness and temperature control within the device thanks to the thermal conductivity of sapphire. Consequently, it provides a free-flow electrophoresis device capable of processing industrial volumes, for industrial applications involving the continuous purification and / or separation of samples via free-flow electrophoresis.

[0236] Subunits shared by two electrophoresis chambers.

[0237] According to a particular embodiment, the present invention relates to a device as defined above, comprising at least two electrophoresis cells such that in at least one of their subunits Ut, t being an integer from 1 to (s-1), at least two of their respective outputs lead to the same input of the same successive subunit Ut+1 by their respective channel Vt.

[0238] In this embodiment, for at least two successive subunits, respectively named the first and second subunits, the second subunit has an inlet linked to two intermediate channels V originating from two different first subunits. Thus, a second subunit is distributed by two distinct first subunits, thereby increasing the flow rate of the solution to be purified and / or separated.

[0239] According to a particular embodiment, the present invention relates to a device as defined above, comprising at least one pair of two electrophoresis cells G1 and G2 of identical configuration and each comprising two subunits, in which

[0240] Vi(G1) is the intermediate channel of V1 of cell G1 connecting the subunit U1 of G1, named Ui(G1), to the subunit U2 of G1 named U2(G1), from the output SVi(Ui(G1)) to the input EVI(U2(G1)),

[0241] VI(G2) is the intermediate channel of V1 of cell G2 connecting the subunit U1 of G2, named Ui(G1), to the subunit U2 of G1 named U2(G1), from the output SVi(Ui(G2)) to the input EVI(U2(G2)), and in which the aforementioned channels Vi(G1) and Vi(G2) lead to the same input of the subunit U2. It is understood that the aforementioned inputs EVI(U2(G1)) and EVI(U2(G2)) represent the same input, named EVI(U2(G1 -G2)) and the aforementioned subunits U2(G1) and U2(G2) represent the same subunit, named U2(G1-G2).

[0242] The arrangement of the two electrophoresis chambers and their channels Vi(G1) and Vi(G2) is configured to distribute the purified and / or separated product from Ui(G1) through channel Vi(G1) and the purified and / or separated product from Ui(G2) through channel Vi(G2) to the same subunit U2(G1-G2) via the same inlet EVI(U2(G1-G2)) of the aforementioned subunit EVI(U2(G1-G2)). This allows the flow rate of the solution to be purified and / or separated in the second subunit, which is common to both electrophoresis chambers, to be doubled.

[0243] According to a particular embodiment, the present invention relates to a device as defined above, in which each plate named Xj, j being an integer varying from 1 to p, comprises qj electrophoresis cells as defined above, qj being an integer from 1 to 10.

[0244] In this embodiment, the device comprises p plates X, named Xi to X P, p being an integer from 1 to 50 and each plate contains from 1 to 10 electrophoresis cells, the number of electrophoresis cells on the plate Xj being denoted qj. The sum of the number of cells in each plate qj when j varies from 1 to p is equal to the total number f of cells in the device.

[0245] When the electrophoresis chamber subunits extend over several plates X due to the use of distinct heights of the hollowed parts, the number qj of electrophoresis cells on the plate Xj corresponds to the number of subunits Ui on the plate Xj.

[0246] Subunits of the chamber on the same plate

[0247] According to a particular embodiment, the present invention relates to a device as defined above, in which p is an integer from 1 to 50, in particular from 1 to 10, when j varies from 1 to p, qj is an integer from 2 to 50, in particular from 2 to 10 comprising p electrophoresis plates Xj each comprising from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5, in which for each electrophoresis chamber, the hollowed-out parts of the s subunits are located in the same plate.

[0248] In this embodiment, the subunits Ui to U s The electrophoresis chamber cells are located on the same plate X. The device comprises p plates X, named Xi to X P , p being an integer from 1 to 50 and each plate comprises from 1 to 50 electrophoresis cells with s subunits, s including from 2 to 5.

[0249] Advantageously, the X plates all include the same number of electrophoresis cells.

[0250] Electrophoresis chambers on the same plate

[0251] According to a particular embodiment, the present invention relates to a device as defined above, in which p is equal to 1 and qi is an integer from 2 to 50, in particular from 2 to 10, comprising a single electrophoresis plate Xi having from 2 to 50 electrophoresis chambers, in particular from 2 to 10 chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5.

[0252] In this embodiment, the device comprises a single plate X which has from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each chamber containing s subunits, s representing an integer from 2 to 5.

[0253] Rowing of subunits

[0254] According to a particular embodiment, the present invention relates to a device as defined above, in which, in each plate Xj, the qj electrophoresis chambers are aligned such that the subunits U of the qj electrophoresis chambers are adjacent to each other by faces (c) or (di). In this embodiment, the subunits U of the same index i in each electrophoresis chamber of the same plate are aligned in a row. Thus, the subunits Ui of the electrophoresis chambers form one row, and the subunits U2 form another row. This arrangement advantageously optimizes the cooling of the electrophoresis chambers by the cooling circuit and the use of the supply, recovery, and discharge channels of the device.

[0255] According to a particular embodiment, the present invention relates to a device as defined above, in which p is an integer from 1 to 50, in particular from 1 to 10, when j varies from 1 to p, qj is an integer from 2 to 50, in particular from 2 to 10, comprising p electrophoresis plates Xj each having from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5, in which for each electrophoresis chamber, the hollowed-out parts of the s subunits are located in the same plate.or in which p is equal to 1 and qi is an integer from 2 to 50, in particular from 2 to 10, comprising a single electrophoresis plate X1 having from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5, and / or in which in each plate Xj, the qj electrophoresis chambers are aligned so that the subunits U of the qj electrophoresis chambers are adjacent to each other by faces (G) or (d).

[0256] According to a particular embodiment, the present invention relates to a device as defined above, comprising a supply channel network configured to connect at least one of the inputs Ei(2) to E1 (ni-1) of the subunit U1 of each electrophoresis chamber to an external micro / milli fluidic circuit for supplying an initial solution containing a product to be purified and / or separated.

[0257] In this embodiment, the supply of initial solution to be purified and / or separated in the electrophoresis chambers is carried out in parallel.

[0258] Device with cooling plate.

[0259] According to a particular embodiment, the present invention relates to a device as defined above, comprising cooling means.

[0260] According to a particular embodiment, the present invention relates to a device as defined above, comprising a vertical succession of plates X, Y, Z whose surfaces are stacked according to the sequence YZY(XYZY) P , in which

[0261] X represents an electrophoresis plate made of inert material.

[0262] Y represents a watertight plate made of sapphire or alumina (Al2O3) with 99% a-AhCh.

[0263] Z represents a cooling plate comprising a heat transfer system (4), p, an integer from 1 to 50, in particular from 1 to 20, represents both the number of stages of said device and the number of plates X, each stage being defined:

[0264] - by the following sequence of plates YZYXYZY, in which:

[0265] - Plate X is located between two plates Y,

[0266] - each of the two plates Z being respectively adjacent to a plate Y,

[0267] - and each of the two Y plates located at the ends of the YZYXYZY sequence, respectively covers a Z plate so that each Z plate is located between two Y plates.

[0268] By "vertical succession of plates" we mean a stacking of plates in which the surfaces of the different plates are in contact. Device with membrane

[0269] According to a particular embodiment, the present invention relates to a device as defined above, in which each electrophoresis chamber is configured to contain at least one selective permeability membrane, in particular selective in sizes, positioned so as to be traversed by the solution containing the product to be separated or purified during the operation of the device, preferably said at least one selective permeability membrane is placed in the Us subunit of each electrophoresis chamber.

[0270] The presence of these membranes also allows for selectivity of sizes of the product to be purified and / or separated.

[0271] Advantageously the membrane can be positioned in a subunit parallel to the face (G) and adjacent to an inlet of a buffer solution, so as to be traversed by the initial solution or by the solution to be purified and / or separated during the operation of the device and so that the part of said solution not having crossed said membrane is conveyed to one of the outlets by a buffer solution from an inlet adjacent to said membrane.

[0272] Device with protrusion.

[0273] According to a particular embodiment, the present invention relates to a device as defined above, in which the upper faces ei and / or the lower faces f of the subunits of each of the electrophoresis chambers comprise at least one protrusion configured not to disturb, during the operation of the device, the circulation in the chamber of the product to be purified and / or separated, and configured to improve heat transfer and to maintain each of said electrophoresis chambers at a selected temperature, in particular in which said at least one protrusion is made of thermally conductive material, preferably sapphire or 99% a-AhCh alumina.

[0274] The protrusions extend from faces (e) and / or (f) of the hollowed-out part towards the interior of said hollowed-out part. They may be located only on one face or on both faces.

[0275] The presence of these protrusions promotes heat exchange between the cooling plates along the path, thus optimizing the separation of the desired molecule while avoiding denaturation of the molecules, maintaining the optimal temperature.

[0276] According to a particular embodiment, the present invention relates to a device as defined above, in which each electrophoresis chamber is configured to contain at least one selective permeability membrane, particularly size-selective, positioned so as to be traversed by the solution containing the product to be separated or purified during operation of the device; preferably, said at least one selective permeability membrane is placed in the U subunit sof each electrophoresis chamber; and / or wherein the upper faces ei and / or the lower faces f of the subunits of each of the electrophoresis chambers comprise at least one protrusion configured not to disturb, during the operation of the device, the circulation in the chamber of the product to be purified and / or separated, and configured to enhance heat transfer and to maintain each of said electrophoresis chambers at a selected temperature, in particular wherein said at least one protrusion is made of thermally conductive material, preferably of sapphire or 99% a-AhCh alumina; and / or said device comprises a supply channel network configured to connect at least one of the inlets Ei (2) to Ei (ni-1) of the subunit Ui of each electrophoresis chamber to an external micro / milli fluidic circuit for supplying an initial solution containing a product to be purified and / or separated.

[0277] Use

[0278] Another object of the present invention relates to the use of an electrophoresis cell according to the invention as defined above, or of an electrophoresis cell device according to the invention as defined above, in the implementation of a process for the purification and / or separation by free-flow electrophoresis of a product to be purified and / or separated contained in an initial solution, comprising a first purification and / or separation step in the Ui subunit(s) followed by successive (s-1) polishing steps in the U2 to U1 subunits. s of electrophoresis chamber(s).

[0279] Electrophoresis purification and / or separation process

[0280] Another object of the present invention relates to a method for purifying and / or separating, by free-flow electrophoresis, a product to be purified and / or separated contained in an initial solution, using an electrophoresis cell device as defined above, comprising the following steps: a) a purification and / or separation step of the aforementioned product in the subunits U1 of each electrophoresis chamber, under an electric field CE(1), comprising: in the subunit U1 of each electrophoresis chamber: o the circulation of a liquid cathode CAL(1) from the inlet E1(1) to the outlet Si(1), o the circulation of a liquid anode ANL(1) from the inlet Ei(ni) to the outlet Si(mi), o the circulation of the initial solution containing the product to be separated and / or purified and at least one buffer solution in the subunit U1, between the liquid cathode CAL(1) and the liquid anode ANL(1), of the inlets Ei(2) to Ei(ni-1) at the outputs Si(2) to Si(mi-1),to obtain the purified and / or separated product from subunit U1, b) (s-1) polishing step(s) k, k varying from 1 to (s-1), each step k comprising each: i) in the intermediate channel Vk of each electrophoresis chamber, o recovery at the outlet SVk(Uk) corresponding to the intermediate channel Vk of said purified and / or separated product from subunit Uk; o the circulation, in the absence of an electric field, of the aforementioned purified and / or separated product, from the outlet SVk(Uk) of the subunit Uk corresponding to the intermediate channel Vk to the inlet EVk(Uk+i) of the subunit Uk+1 corresponding to the intermediate channel Vk, ii) in the subunit Uk+1 of each electrophoresis chamber, in the presence of an electric field CE(k+1): o the circulation of a liquid cathode CAL(k+1) from the inlet Ek+1 (1 ) to the outlet Sk+1 (1 ), o the circulation of a liquid anode ANL(k+1) from the inlet Ek+i(nk+i) to the outlet Sk+i(rrik+i),o the circulation of said purified and / or separated product from the outlet of subunit Uk and at least one buffer solution in subunit Uk+1, between the liquid cathode CAL(k+1) and the liquid anode ANL(k+1), from inlets Ek+i(2) to Ek+i(nk+i-1) to outlets Sk+i(2) to Sk+i(mk+i-1), to obtain the product (k+1) times purified and / or separated from subunit Uk+1; c) a recovery step comprising: recovery at one of the outlets S, s (2) to S s (m s -1) of unit U s from each electrophoresis chamber of the product(s) once purified and / or separated.

[0281] Electric fields

[0282] According to a particular embodiment, the present invention relates to a process as defined above, in which each electric field CE(i), generated in the subunit, i an integer varying from 1 to s, during the fluidic circulation in each of the electrophoresis chambers, is from 200V to 4000V.

[0283] According to a particular embodiment, the present invention relates to a method as defined above, in which at least two values ​​of electric fields CE(i), generated respectively in the subunits, i an integer varying from 1 to s, are different.

[0284] According to a particular embodiment, the present invention relates to a process as defined above, in which said process is carried out under a continuous flow of the initial solution containing the product to be purified and / or separated. According to a particular embodiment, the present invention relates to a process as defined above, carried out by a device as defined above, in which s is equal to 2, comprising the following steps: a) a purification and / or separation step of the product to be purified and / or separated in the subunits Ui of each electrophoresis chamber, under an electric field CE(1), comprising: in the subunit Ui of each electrophoresis chamber: o the circulation of a liquid cathode CAL(1) from the inlet Ei(1) to the outlet Si(1), o the circulation of a liquid anode ANL(1) from the inlet Ei(ni) to the outlet Si(mi),a) the circulation of the initial solution containing the product to be separated and / or purified and at least one buffer solution in the subunit Ui, between the liquid cathode CAL(1) and the liquid anode ANL(1), from the inlets Ei(2) to Ei(ni-1) to the outlets Si(2) to Si(mi-1), to obtain the purified and / or separated product from the subunit Ui, b) a polishing step comprising: i) in the intermediate channel Vi of each electrophoresis chamber, o the recovery at the outlet SVi(Ui) corresponding to the intermediate channel Vi of said purified and / or separated product from the subunit Ui; o the circulation, in the absence of an electric field, of said purified and / or separated product, from the outlet SVi(Ui) of the subunit Ui corresponding to the intermediate channel Vi to the inlet EVI(U2) of the subunit U2 corresponding to the intermediate channel V1, ii) in the subunit U2 of each electrophoresis chamber,in the presence of an electric field CE(2): o the circulation of a liquid cathode CAL(2) from the inlet E2(1) to the outlet S2(1), o the circulation of a liquid anode ANL(2) from the inlet E2(n2) to the outlet 82(012), o the circulation of said purified and / or separated product from the outlet of subunit U2 and at least one buffer solution in subunit U2, between the liquid cathode CAL(2) and the liquid anode ANL(2), from the inlets E2(2) to E2(n2-1) to the outlets 82(2) to 82(012-1), to obtain the purified and / or separated and once polished product from subunit U2; c) a recovery step comprising: recovery at one of the outlets 82(2) to 82(012-1) of the U2 unit of each electrophoresis chamber of the purified and / or separated product, once polished.

[0285] Figures and examples

[0286] Figure 1 represents the hollowed-out portion of a subunit of an electrophoresis chamber of an electrophoresis plate X between two adjacent plates Y, one upper and one lower. The hollowed-out portion is inscribed within a rectangular parallelepiped of width Lai, length Loi, and height hi, bounded by the faces (ai, bi, c, di, ei, fi). The faces (ai, bi, c, d) form the lateral walls between the hollowed-out portion and the plate X, with the faces (ai, bi) being parallel to each other and the faces (c, di) also parallel to each other. Face (a) is bounded by the edges (A1i, A2) of dimension Lai, and face (bi) by the edges (B1i, B2) of dimension Lai. Face (c) is bounded by edges (C1i, C2) of dimension Law, face (d) by edges (D1i, D2>) of dimension Law. Edges (A1i, B1i, C1i, D1) bound face (e) formed by the upper plate Y and edges (A2i, B2i, C2i, D2>) bound face (f) formed by the lower plate Y.

[0287] Figure 2 shows the schematic of an electrophoresis chamber with two subunits. The first subunit, U1, has 9 inputs from Ei(1) to Ei(9) and 5 outputs from Si(1) to Si(5), with input Ei(1) facing output Si(1) and input Ei(9) facing output Si(5). The second subunit, U2, has 8 inputs from E2(0) to E2(8) and 12 outputs from E2(1) to E2(12), with input E2(1) facing output E2(1) and input E2(8) facing output S2(12). Subunit U1 is connected to the second subunit U2 by an intermediate channel V1 extending from the output Si(4) of Ui, named SVi(Ui), to the input E2(4) of U2, named EVI(U2). Figure 3 shows the schematic of an electrophoresis chamber with three subunits. Subunit Ui is connected to subunit U2 by an intermediate channel Vi. Subunit U2 is connected to subunit U3 by an intermediate channel V2.

[0288] Figure 4 shows in part a) the exploded view diagram of an electrophoresis cell comprising a plate X between two adjacent plates Y forming a YXY sequence. Plate X comprises a chamber of two subunits U1 and U2 connected by an intermediate channel V1. Subunit U1 has 7 inputs from Ei(1) to Ei(7) and 5 outputs from Si(1) to Si(5), with the output of the intermediate channel SVi(Ui) corresponding to Si(3). Subunit U2 has 7 inputs from E2(1) to E2(7) and 7 outputs from S2(1) to S2(5), with the input of the intermediate channel EVI(U2) corresponding to E2(4). The upper Y plate and the X plate are engraved with the supply and retrieval channels of the outputs Si (1 ), Si(2), Si(4) and Si(5) and of the inputs E2(1), E2(2), E2(3), E2(5), E2(6), E2(7), the output Si(3) and the input E2(4) corresponding to the intermediate channel V1 being excluded.

[0289] In part b), (1) represents an electrophoresis plate comprising a single chamber with two rectangular parallelepiped-shaped subunits (6), including inlets or outlets (7) and supply or return channels (8). (2) represents a synthetic sapphire plate providing fluid separation between two plates but allowing heat exchange. (3) represents a cooling plate Z comprising a cooling system (4) which includes a recess allowing the circulation of a heat transfer fluid from an inlet to an outlet, the inlet of the heat transfer fluid being on the same side as the inlets of the electrophoresis chamber. The device consists of a series of plates YZYXYZY.

[0290] Figure 5 represents the schematic of an electrophoresis chamber comprising two subunits and including a complementary intermediate channel W, in part a) the intermediate channel V1 and the complementary intermediate channel W are adjacent, in part b) the two channels are not adjacent, an outlet between the two channels allows the recovery of a compound from the product to be purified and / or separated.

[0291] Figure 6 represents the two-subunit electrophoresis chamber scheme, in which two U1 subunits distribute the same U2 subunit; in part a), the respective outputs of the two intermediate channels V1 correspond to different outputs of the Ui(1) and Ui(2) subunits; in part b) the respective outputs of the two intermediate channels V1 correspond to the same subunit output.

[0292] Figure 7 represents the schematic of a plate X of a free-flow electrophoresis device comprising 10 electrophoresis chambers of two subunits of Figure 2. The 10 U1 subunits form one row and the U2 subunits form another row.

[0293] Figure 8 represents two devices in exploded view and without representation of the clamping means of all the plates, in part a) a single-stage device comprising a row of 10 electrophoresis chambers and in part b) a two-stage device, each stage comprising a row of 10 electrophoresis chambers.

[0294] (1) represents an electrophoresis plate comprising a row of 10 electrophoresis chambers, each having two rectangular parallelepiped-shaped subunits (6). Each chamber includes inlets or outlets and supply or return channels. Two adjacent subunits are separated by a single wall. (2) represents a synthetic sapphire plate ensuring the separation of fluids between two plates but allowing heat exchange. (3) represents a cooling plate comprising a cooling system (4) which includes a recess allowing the circulation of a heat transfer fluid from an inlet to an outlet. (5) represents a stage consisting of a succession of plates YZYXYZY.

[0295] The device in part a) comprises 1 stage and consists of the following sequence YZYXYZY.

[0296] The device in part b) comprises 2 stages and consists of the following sequence YZYXYZYXYZY in which the central sequence YZY is common to both stages.

[0297] Figure 9 shows a diagram of a device with a plate clamping system. In this particular embodiment, the clamping means consist of two plates (11) that clamp the entire sequence of plates X, Y, and Z, using fastening means (12) connecting the two plates (11), the distance between which can be adjusted. The fastening means (12) are, for example, screws.

[0298] Figure 10 represents the schematic of the system used for the implementation to evaluate the purification and separation yields of a subunit.

[0299] Figure 11 represents the schematic of a chamber with two identical subunits used for implementation to evaluate the purification and separation yield of an initial solution consisting of 3 dyes (fluorescein, rhodamine B and rhodamine 6G).

[0300] Figure 12 is a photograph taken of the Ui subunit of an electrophoresis chamber with 5 inlets and 9 outlets, with a thickness of 100 pm, during a separation of a mixture of three colored compounds (fluorescein, rhodamine B and rhodamine 6G), carried out at 2500 V.

[0301] Figure 13 shows the HPLC spectra of the products at the outputs of the first subunit Ui of an electrophoresis chamber with 5 inlets and 9 outlets, with a thickness of 100 pm, during a separation test of a mixture of three colored compounds (fluorescein, rhodamine B and rhodamine 6G), at 0V in part a) and at 2500 V in part b).

[0302] Figure 14 is a graph representing the output flow rate of each dye at the different outputs of the first subunit Ui of an electrophoresis chamber with 5 inlets and 9 outlets, 100 pm thick, during a separation test of a mixture of three colored compounds (fluorescein, rhodamine B and rhodamine 6G), at 2500 V.

[0303] Figure 15 shows the HPLC spectra of the products at the outputs of the second U2 subunit of an electrophoresis chamber with 5 inlets and 9 outlets, with a thickness of 100 pm, during a separation test of a mixture of three colored compounds (fluorescein, rhodamine B and rhodamine 6G), at 0V in part a) and at 2500 V in part b).

[0304] Figure 16 is a graph representing the output flow rate of each dye at the different outputs of the second U2 subunit of an electrophoresis chamber with 5 inlets and 9 outlets, 100 pm thick, during a separation test of a mixture of three colored compounds (fluorescein, rhodamine B and rhodamine 6G), at 2500 V.

[0305] Example 1: Optimization of the separation of a mixture of Fluorescein, Rhodamine B and Rhodamine 6G in a two-subunit electrophoresis cell

[0306] Materials and methods

[0307] The electrophoresis system

[0308] The electrophoresis system consists of an air supply that feeds a pump connected to flow meters, which control the flow within the electrophoresis chip (Figure 10). This system ensures a stable flow rate for optimal system stability. The electrophoresis chip is interchangeable depending on the separation requirements of different media. Before each experiment, the system requires a start-up and a stabilization period to ensure steady-state operation (stabilized flow rates, pressure, and temperature).

[0309] Model molecules

[0310] The free-flow electrophoresis system was implemented to separate a mixture of three fluorescent molecules: fluorescein, rhodamine B, and rhodamine 6G. These are small dyes with similar molar masses, each with a distinct color that allows for visual identification prior to HPLC analysis. This visual inspection also validates the cell's functionality before the application of the electric field.

[0311] These dyes are easily analyzed by HPLC, they have distinct retention times, and their calibration curves have been defined. By HPLC, the retention time of:

[0312] • Fluorescein, is 5.9 minutes,

[0313] • Rhodamine B, is 7.6 minutes

[0314] • Rhodamine 6G, is 8.6 minutes.

[0315] The chemical structures of the three molecules are shown below.

[0316] Rhodamine B Rhodamine 6G Fluorescein

[0317] Mm = 479.01 g / mol (Purple) Mm = 479.01 g / mol (Red) Mm = 332.3 g / mol (Yellow)

[0318] C28H31CIN2O3 C28H31CIN2O3 C20H12O5

[0319] These dyes also possess different surface charges in aqueous solution, allowing for their migration and separation under the application of an electric field. This surface charge was measured by analyzing the zeta potential with the Malvern Zetasizer. The results obtained (Table 1) show that the three dyes have a significant difference in zeta potential and therefore in surface charge. The zeta potential of each compound was measured beforehand and is reported in Table 1.

[0320] Fluorescein at pH 7.49 exhibited a zeta potential of -23.5 mV, rhodamine B -0.3 mV and rhodamine 6G +9.4 mV.

[0321] Table 1: Measured zeta potential of the three dyes

[0322] It is important to note that rhodamine B is weakly charged and therefore its electrophoretic mobility is very low regardless of the strength of the electric field.

[0323] The separation and purification of a sample containing a mixture of these 3 compounds, notably containing rhodamines B and 6G, was carried out in each of the two subunits constituting the electrophoresis cell, by analyzing the output fluxes in order to demonstrate:

[0324] - the technological feasibility of the cell composed of 2 or more subunits that cooperate.

[0325] - the scientific interest related to the continuous separation of various compounds with similar properties such as the mass of biomolecules.

[0326] The following examples describe the tests set up to separate the mixture of the three dyes in a first subunit U1, followed by polishing to separate a stream of the sample exiting U1 and introduced into a subunit U2, as illustrated in Figure 11. The sample recovered in the second subunit corresponds to the output comprising a Rhodamine B / Rhodamine 6G mixture obtained at the outlet of the first subunit after the first separation step with a 9-output subunit. The liquid electrodes and the separation buffer maintain the same compositions in both subunits.

[0327] Example 2: First electrophoretic separation step in the first subunit U1

[0328] A separation of a sample consisting of a mixture of the 3 dyes (fluorescein, rhodamine B and rhodamine 6G) was implemented with a U1 subunit of EAD-008 configuration which comprises:

[0329] - ni = 5 entries

[0330] - rm = 9 outputs,

[0331] - a height hi of 100 pm

[0332] - width of 3 cm,

[0333] - length L01 of 6.4 cm

[0334] The flow rates of the injected fluids, measured by flow meters at the inlets of subunit U1, are 10, 100, and 20 pL / min for the sample, buffer solutions, and liquid electrode, respectively. The inlet solution setup is shown in Table 2 below.

[0335] The applied electric field voltage CE(1) is set at 2500 V.

[0336] Table 2: Operating conditions in the inputs Ei(1) to Ei(5) of the subunit Ui

[0337] Thus, the inlets Ei(1) and Ei(5) were each supplied with an electrolytic solution for the anode and cathode, respectively. The two electrolytic solutions for the cathode and anode had the same composition. The liquid electrolytic solution had the following composition: 10 mM HEPES, 0.2% (w / v) HPMC, 40% methanol, and 1.5 M KCl. The flow rate was set at 20 pL / min. These electrolytic solutions had a pH of 7.75, a conductivity of 102.6 mS / cm, and a viscosity at 25°C of 2.19 mPa / s. The electric field was generated by the electrolytic solutions in the electrophoresis chamber, which were supplied with solutions containing the carbon electrodes, anode and cathode, respectively. The central inlet Ei(3) was fed with the sample to be purified, composed of 0.156 mg / mL of fluorescein, 0.23 mg / mL of Rhodamine 6G and 0.25 mg / mL of Rhodamine B; The flow rate set for the sample was 10 pl / min throughout the purification.The two inlets Ei(2) and Ei(4) were supplied with a buffer solution composed of 10 mM HEPES, 0.2% (w / v) HPMC, and 0.1% (w / v) Tween 20 in water, at a fixed flow rate of 100 pL / min. The buffer solution had a pH of 7.61, a conductivity of 393.6 pS / cm, and a viscosity of 2.19 mPa / s at 25°C.

[0338] The electric field was set at 2500 V. After stabilization of the flows, i.e. the system was left running for 1 h45 minutes and the products at the outlet of the electrophoresis chamber were collected in tubes and analyzed by HPLC in order to determine the percentage of each compound at each outlet.

[0339] A control test was carried out with an electric field fixed at 0 V. The system was left running continuously for 30 minutes.

[0340] Example 3 - Result of separation by electrophoresis in the Ui subunit

[0341] Visual analysis

[0342] Figure 12 is a photograph taken of the Ui subunit under an electric field of 2500 V during electrophoresis made possible by the use of transparent sapphire plates and colored products to be separated.

[0343] At 2500 V, Figure 12 visually shows an initial sample beam separating into 3 beams of different colors, mainly centered respectively at the output Si(3) with yellow color, at the output Si(5) with violet color, and at the output Si(6) with red color.

[0344] At 0 V, the sample beam composed of the 3 dyes is visible and centered. There is no electric field, therefore no deflection is visible. The dyes exit only at output Si(5), which is the central output facing input Ei(3).

[0345] HPLC Analysis

[0346] At 2500 V, the outputs of the Si(1) to Si(9) electrophoresis subunit were analyzed by HPLC.

[0347] For the control, at 0 V, only the outputs of the electrophoresis subunit Si(4) to Si(6) were analyzed.

[0348] The HPLC spectra of the products exiting at the 0V and 2500V outputs are shown in Figure 13.

[0349] The relative distribution of the concentration of each dye in the outputs by HPLC analysis at 2500 V is reported in Table 3.

[0350] Table 3: Percentage distribution of concentration of each compound in the outputs of the first subunit Ui

[0351] The results of the HPLC analysis at 2500 V reveal:

[0352] Fluorescein emerges pure at the three outputs Si(2), Si(3), and Si(4), primarily at output Si(3), without any signal from rhodamine B and 6G.

[0353] 75.4% of rhodamine B exits pure at the Si (5) outlet

[0354] 55.5% of the rhodamine 6G exits pure at the Si (7) exit

[0355] An output Si(6) stream consisting of a mixture of Rhodamine B (24.6%) and Rhodamine 6G (44.5%). Therefore, to optimize the separation, the output Si(6), composed of Rhodamine B (24.6%) and Rhodamine 6G (44.5%), must be treated in the next second subunit to finalize the separation of the Rhodamines.

[0356] Furthermore, it is also observed that the concentration of each dye is higher at the subunit outlet when the electric field is applied than when it is not. This indicates a concentration of the sample during separation, rather than dilution as previously observed when the electric field is zero. This is also reflected in the visual analysis of the recovered fractions.

[0357] Analysis of outgoing flow rates

[0358] Table 4 below shows the distribution of flow rates of the outgoing flows from subunit Ui.

[0359] Table 4: Flow rates at the outputs of subunit Ui

[0360] Table 5 reports the flow rate values ​​in mg / min of each molecule at each of the outputs of the subunit Ui, corresponding to the concentration multiplied by the flow rate of the outgoing flux, for a zero electric field.

[0361] Table 5: Flow rate in mg / min of each dye at the outputs of the Ui subunit under zero electric field.

[0362] Table 6 and Figure 14 report the flow rate values ​​in mg / min of each molecule at each of the outputs of the U1 subunit, corresponding to the concentration multiplied by the flow rate of the outgoing flux, for a zero electric field and for an electric field of 2500 V.

[0363] Table 6: Flow rate in mg / min of each dye at the outlets of subunit Ui under an electric field of 2500 V. In this case, the flow rate at the outlet of Si(6) of the first subunit is approximately 20 pL / min, therefore:

[0364] - Rhodamine B, present at 0.024 mg / mL, is recovered at a rate of 0.00056 mg / min

[0365] - Rhodamine 6G, present at 0.045 mg / mL, is recovered at a rate of 0.0013 mg / min

[0366] Example 4: Second subunit U2 - Polishing step by electrophoretic separation

[0367] An electrophoretic separation of the outgoing Si(6) flux comprising a mixture of rhodamine 6G and rhodamine B, as the incoming flux at a rate of 20 pL / min, was then implemented in the second subunit U2 of configuration EAD-008, i.e., of the same configuration as that of subunit U1, which comprises:

[0368] - n2 = 5 entries

[0369] - m2 = 9 outlets,

[0370] - a height ti2 of 100 pm

[0371] - width La2 of 3 cm

[0372] - length L02 of 6.4 cm

[0373] The flow rates injected using the flow meters into the inlets of subunit U2 are, for the mixture to be purified and / or separated, the buffer solutions, and the liquid electrode, respectively, 10 / 100 / 20 pL / min. The inlet solution configuration is shown in Table 7 below. The applied electric field voltage CE(2) is set at 2500 V, the same value as that of subunit Ui.

[0374] Table 7: Operating conditions in the E2(1) to E2(5) inputs of the U2 subunit

[0375] Thus, inlets E2(1) and E2(5) were each supplied with an electrolytic solution for the anode and cathode, respectively. The two electrolytic solutions for the cathode and anode had the same composition. This composition was identical to that used for the Ui subunit. The liquid electrolytic solution had the following composition: 10 mM HEPES, 0.2% (w / v) HPMC, 40% methanol, and 1.5 M KCl. The flow rate was set at 20 pL / min. These electrolytic solutions had a pH of 7.75, a conductivity of 102.6 mS / cm, and a viscosity at 25°C of 2.19 mPa / s. The electric field was generated by the electrolytic solutions in the electrophoresis chamber, which were supplied with solutions containing the carbon electrodes, anode and cathode, respectively.

[0376] The central inlet E2(3) was fed by the flow from outlet Si(6), which constituted the solution to be purified and / or separated, comprising the mixture of two rhodamines. The flow rate was set at 20 pL / min throughout the separation.

[0377] The two inlets E2(2) and E2(4) were supplied with a buffer solution composed of 10 mM HEPES, 0.2% (w / v) HPMC, and 0.1% (w / v) Tween 20 in water, at a fixed flow rate of 100 pL / min. The buffer solution had a pH of 7.61, a conductivity of 393.6 pS / cm, and a viscosity of 2.19 mPa / s at 25°C.

[0378] The electric field was fixed in this test at 2500 V. After stabilization of the flows, the products exiting the U2 subunit were collected for 30 minutes in tubes and analyzed by HPLC to determine the percentage of each compound at each exit.

[0379] A control test was carried out with an electric field fixed at 0 V. The system was left running continuously for 30 minutes.

[0380] Example 5 - Result of polishing by electrophoretic separation in the U2 subunit

[0381] At 2500 V, the initially faint beam becomes invisible, preventing visual analysis and photography. Only fraction recovery and HPLC analysis, performed after 30 minutes of sample collection, can determine whether or not the dyes were properly separated. However, the recovered fractions show different coloration in some outputs, particularly in the S2(5), S2(6), and S2(7) fractions.

[0382] For the control, at 0 V, only the outputs of the electrophoresis subunit S2(4) to 82(6) were analyzed.

[0383] The HPLC spectra of the products exiting at the 0V and 2500V outputs are shown in Figure 15. The relative distribution in concentration of each dye in the outputs by HPLC analysis at 2500V is reported in Table 8 below.

[0384] Table 8: Percentage distribution of concentration of each compound in the outputs of the second U2 subunit

[0385] At 2500 V, we again observe a separation of the dyes present at the outputs S2(5) to S2(7): rhodamine B is pure in the fraction S2(5) which represents a gain of 18% of the total rhodamine to be purified.

[0386] We therefore obtain a purification yield of pure rhodamine B of the order of 88.5% with 2 successive subunits.

[0387] Furthermore, by working continuously, it is still possible to recycle the output flux 82(6) of the 2nd subunit which represents approximately 11.5% of the initial quantity of rhodamine B.

[0388] Analysis of outgoing flow rates

[0389] Table 9 below shows the distribution of flow rates of the outgoing flows from subunit U2.

[0390] Table 9: Flow rates at the outputs of subunit U2

[0391] Table 10 reports the flow rate values ​​in mg / min of each molecule at each of the outlets of the second subunit U2, corresponding to the concentration multiplied by the outflow rate, for a zero electric field.

[0392] Table 10: Flow rate in mg / min of each dye at the outlets of the second subunit U2 under zero electric field.

[0393] Table 11 and Figure 16 report the flow rate values ​​in mg / min of each molecule at each of the outputs of the subunit Ui, corresponding to the concentration multiplied by the flow rate of the outgoing flux, for a zero electric field and for an electric field of 2500 V.

[0394] Table 11: Flow rate in mg / min of each dye at the outlets of the second subunit U2 under an electric field of 2500 V.

[0395] Example 6: Summary of the separation

[0396] Table 12 below reports the separation yield of each compound in the mixture and the percentage of compounds still in mixture form.

[0397] Table 12: Separation efficiency

[0398] Thus, the cell comprising two subunits of EAD 008 configuration enabled the purification of the mixture composed of fluorescein, rhodamine B, and rhodamine 6G with excellent separation yields: 100% for fluorescein, 88% for rhodamine B, and 80% for rhodamine 6G. These three compounds are obtained diluted in an aqueous medium and can be concentrated to obtain their pure form.

[0399] In conclusion, the use of a cell comprising a chamber with two successive cooperating subunits allows for optimized separation and purification. In a mixture of three dyes, it was possible to eliminate a greater proportion of the unwanted compounds (fluorescein and rhodamine 6G) in favor of the compound of interest, in this case, rhodamine B. Furthermore, the free-flow electrophoresis cell allows for the adaptation and optimization of operating conditions and configurations to achieve greater purification and / or separation in a continuous flow.

[0400] The adaptability of the electrophoresis cell allows a configuration introducing a third subunit to increase the purification and / or separation yield of rhodamines B and 6G by separating and purifying the remaining percentage to be recycled contained in the output 82(6) of the U2 subunit.

Claims

1. DEMANDS 1. Free-flow electrophoresis cell for purifying and / or separating an initial solution containing a product to be purified and / or separated, comprising at least one plate X between two adjacent plates Y: X being an electrophoresis plate (1) made of inert material, Y being a sealed plate (2) of sapphire or alumina Al2O3 with 99% a-AhOs, an electrophoresis chamber comprising s subunits U, s representing an integer from 2 to 5 and / being an integer ranging from 1 to s, in which each subunit U comprises o a hollowed-out part in a plate X closed between two plates Y, ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, c, di) and 2 upper and lower faces (ei, f), ■ said hollowed-out part being of length Loi and width Lai, ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces (ai, bi) being parallel to each other, the face (a) being delimited by two edges (AT, A2 ) of dimension Lai and the face (bi) being delimited by two edges (B1 i, B2>) of dimension Lai, ■ the lateral faces (c, di) being parallel to each other, the face (c) being delimited by two edges (C1 i, C2 ) of dimension Law and the face (di) being delimited by two edges (D1 i, D2>) of dimension Law, o ni successive entrances Ei(1), Ei(2) to Ei(ni-1), Ei(n), ni being an integer from 4 to 9, distributed on the face (a) between A1 i and A2i and aligned along a direction parallel to A1 i and A2i, o mi successive exits from Si(1), Si(2) to Si(mi-1), Si(rrii), mi being an integer from 4 to 12, distributed on the face (bi) between B1 i and B2i and aligned along a direction parallel to B1 i and B2i, such that Si(1) faces Ei(1) and Si(rrii) faces Ei(n) along a parallel to C1 i and D1 i, a cooling circuit, characterized in that the electrophoresis chamber comprises: (s-1) intermediate channels Vk, k being an integer ranging from 1 to (s-1), each channel Vk connecting the two successive subunits Uk and Uk+1 by one of the outputs SVk(Uk) chosen from Sk(2) to Sk(mk-1) of the subunit Uk to one of the inputs EVk(Uk+i) of the subunit Uk+1 chosen from Ek+1 (2) to Ek+1 (nk+i-1), said input EVk(Uk+i) and said output SVk(Uk) being respectively the input and output connected by the intermediate channel Vk.

2. Cell according to claim 1, comprising: supply channels configured to connect the s inputs Ei(1), i varying from 1 to s, of each subunit of the electrophoresis chamber to at least one external micro / millifluidic circuit for supplying at least one liquid cathode, supply channels configured to connect the s inputs Ei(n) of each subunit of the electrophoresis chamber to at least one external micro / millifluidic circuit for supplying at least one liquid anode, a supply channel configured to connect at least one of the inputs Ei(2) to E1 (ni-1) of the subunit U1 to an external micro / millifluidic circuit for supplying an initial solution containing a product to be purified and / or separated,supply channels configured to connect at least one input of each electrophoresis chamber subunit to at least one external micro / millifluidic supply circuit for at least one buffer solution, at least one recovery channel configured to connect one of the outputs of subunit U, s chosen from S s (2) to S s (m s -1) from the electrophoresis chamber to an external micro / milli fluidic recovery circuit.

3. Cell according to claim 1 or 2, said cell being configured to: o in each subunit Ui, , in the presence of an electric field generated CE(i) between a liquid cathode CAL(i) and a liquid anode ANL(i) parallel to A1 i and perpendicular to C1 i, o and in operation, a) in subunit Ui: o circulate the liquid cathode CAL(1) from inlet Ei (1 ) to outlet Si (1 ), o circulate the liquid anode ANL(1) from inlet Ei(ni) to outlet Si(mi), o circulate the initial solution containing the product to be separated and / or purified and at least one buffer solution in subunit Ui, between the liquid cathode CAL(1) and the liquid anode ANL(1), from inlets Ei(2) to Ei(m-1) to outlets Si(2) to Si(mi-1), to obtain the purified product and / or separated from the subunit Ui, b) in each intermediate channel Vk, k varying from 1 to (s-1), o recover at the output SVk(Uk) the purified and / or separated product from the subunit Uk;to circulate the aforementioned product from the SVk(Uk) output of the Uk subunit through channel Vk, from SVk(Uk) to the EVk(Uk+i) input of the Uk+i subunit; c) in the Uk+i subunit: to circulate the liquid cathode CAL(k+1) from the Ek+i (1) input to the Sk+i (1) output; to circulate the liquid anode ANL(k+1) from the Ek+i(nk+i) input to the Sk+i(rrik+i) output; to circulate the purified and / or separated product from the SVk(Uk) output of the Uk subunit and at least one buffer solution in the Uk+i subunit, between the liquid cathode CAL(k+1) and the liquid anode ANL(k+1), from the Ek+i(2) to Ek+i(nk+i-1) inputs to the outputs Sk+i(2) to Sk+i(mk+i- 1), d) recover to one of the outputs S; s (2) to S s (m s -1) the purified and / or separated product.

4. A cell according to any one of claims 1 to 3, further comprising: at least one complementary intermediate channel W, said complementary intermediate channel W connecting two successive subunits U qand U q +i, q being an integer from 1 to (s-1), said channel W extending: from one of the outputs of unit U q chosen from S q (2) to S q (m q -1), different from the output linked to channel Vq, to ​​the input linked to channel V q of subunit U q +i, the aforementioned channel V input q being connected to at most (m q -3) chosen exit(s) from S q (2) to S q (m q -1).

5. Cell according to any one of claims 1 to 4, comprising a plate X between two adjacent plates Y, said plate X comprising the hollowed-out parts of the subunits Ui of said electrophoresis chamber, the height of the hollowed-out part of each subunit being of identical value, or comprising at least two plates X, each plate X being between two adjacent plates Y, each plate X comprising at least one hollowed-out part of a subunit Ui of said electrophoresis chamber.

6. Cell according to any one of claims 1 to 5, comprising at least one cooling plate Z comprising a heat transfer system, said plate Z being adjacent to one of the two plates Y adjacent to said plate X, and / or comprising at least two plates Z comprising a heat transfer system, said cell comprising the following sequence of plates ZYXYZ, in particular the sequence YZYXYZY, in particular wherein said heat transfer system is a network of pipes configured to permit the circulation of one or more heat transfer fluids parallel to the edges Ci or perpendicular to the edges Ci of the subunits Ui, in particular said network of pipes comprising inlets and outlets of the heat transfer system, engraved in the plate Z.

7. Cell according to any one of claims 1 to 6, wherein the subunits U of the electrophoresis chamber comprise means for channeling the flows opening onto the inlets Ei(1) to Ei(ni) and / or onto the outlets Si(1) to Si(rrii) of the subunits U of the electrophoresis chamber, said channeling means preferably being etched into the electrophoresis plate X, said means in particular being triangular in shape, situated between two adjacent inlets or two adjacent outlets of a subunit, and / or wherein the width Lai of each of the s subunits U is from 1.0 to 8.0 cm, and / or the length Loi of each of the s subunits U of the electrophoresis chamber is from 2.0 to 20.0 cm, and / or wherein the electrophoresis plate X is made of a material selected from polytetrafluoroethylene (PTFE), perfluoroalkoxy (PFA) and fluoroethylene propylene (FEP), in particular Teflon™, Teflon™-PFA and Teflon™-FEP sheets.

8. Cell according to any one of claims 1 to 7, wherein s is equal to 2, the electrophoresis chamber comprising two subunits Ui and IL, the subunit Ui comprising a portion hollowed out in a plate X ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, ci, di) and 2 upper and lower faces (ei, fi), ■ said hollowed-out part being of length Loi and width Lai, ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces (ai , bi) being parallel to each other, the face (ai) being delimited by two edges (A1 i, A2i) of dimension Lai and the face (bi) being delimited by two edges (B11 , B2i) of dimension Lai , ■ the lateral faces (ci, di) being parallel to each other, the face (ci) being delimited by two edges (C1 i, C2i) of dimension Law and the face (di) being delimited by two edges (D11 , D2i) of dimension Law , o ni successive entries Ei(1), Ei(2) to Ei(ni-1), Ei(ni), ni being an integer from 4 to 9, distributed on the face (ai) between A11 and A2i and aligned along a direction parallel to A11 and A2i, or successive exits from Si (1 ), Si(2) to Si(mi-1), Si(mi), r being an integer from 4 to 12 , distributed on the face (bi) between B1 i and B2i and aligned along a direction parallel to B1 i and B2i, such that Si(1) faces Ei(1) and Si(mi) faces Ei(ni) along a direction parallel to C1 i and D1 i, the subunit U2 comprises a hollowed-out portion in a plate X, possibly in the aforementioned plate X of the aforementioned subunit U1, ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, b2, C2, d2) and 2 upper and lower faces (e2, f2), ■ said hollowed-out part being of length L02 and width La2, ■ of height ti2 corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces ai, b2) being parallel to each other, the face (32) being delimited by two edges (AI2, A22) of dimension La2 and the face (b2) being delimited by two edges (BI2, B22) of dimension La2, ■ the lateral faces (C2, d2) being parallel to each other, the face (C2) being delimited by two edges (CI2, C22) of dimension L02 and the face (d2) being delimited by two edges (DI2, D22) of dimension L02, o n2 successive entries E2O), E2(2) to E2(n2-1), E(n2), n2 being an integer from 4 to 9, distributed on the face (32) between AI2 and / 2i and aligned along a direction parallel to AI2 and A22, o m2 successive outputs of S2(1), S2(2) to S2(rri2-1), S2(rri2), m2 being an integer from 4 to 12, distributed on the face (b2) between BI2 and B22 and aligned along a direction parallel to BI2 and B22, so that S2O) faces E2O) and S2(rri2) faces E2(n2) along a direction parallel to CI2 and DI2, characterized in that it comprises an intermediate channel Vi, connecting the two subunits U1 and U2 by one of the outputs SVi(Ui) chosen from Si(2) to Si(mi-1) of the subunit U1 to one of the inputs EVI(U2) of the subunit U2 chosen from E2(2) to E2(n2-1).

9. Cell according to claim 8, comprising: a supply channel configured to connect the inlet Ei(1) of the electrophoresis chamber subunit U1 to an external micro / millifluidic circuit for supplying a first liquid cathode CAL(1), a supply channel configured to connect the inlet Ei(ni) of the electrophoresis chamber subunit U1 to an external micro / millifluidic circuit for supplying a first liquid anode ANL(1), a supply channel configured to connect at least one of the inlets Ei(2) to E1 (ni-1) of the subunit U1 to an external micro / millifluidic circuit for supplying an initial solution containing a product to be purified and / or separated, at least one supply channel configured to connect at least one remaining inlet of the unit U1 to at least one external micro / millifluidic circuit for supplying at least one buffer solution,a supply channel configured to connect the inlet E2(O) of the electrophoresis chamber subunit U2 to an external micro / millifluidic circuit supplying a second liquid cathode CAL(2), a supply channel configured to connect the inlet E2(n2) of the electrophoresis chamber subunit U2 to an external micro / millifluidic circuit supplying a second liquid anode ANL(2), at least one supply channel configured to connect at least one remaining inlet of the U2 unit to at least one external micro / millifluidic circuit supplying at least one buffer solution, at least one recovery channel configured to connect one of the outputs of the U2 subunit selected from 82(2) to S2(rri2-1) of the electrophoresis chamber to an external micro / millifluidic recovery circuit, said cell being configured to,in the presence of: o an electric field generated CE(1) in subunit U1 between liquid cathode CAL(1) and liquid anode ANL(1) parallel to A1 i and perpendicular to C11, o an electric field generated CE(2) in subunit U2 between liquid cathode CAL(2) and liquid anode ANL(2) parallel to AI2 and perpendicular to CI2, o and during operation, a) in subunit U1: o circulate liquid cathode CAL(1) from inlet E1 (1 ) to outlet Si (1 ), o circulate liquid anode ANL(1) from inlet Ei(ni) to outlet Si(mi), o circulate the initial solution containing the product to be separated and / or purified and at least one buffer solution in subunit Ui, between liquid cathode CAL(1) and liquid anode ANL(1), from inlets Ei(2) to Ei(ni-1) at the outputs Si(2) to Si(mi-1), b) in the intermediate channel V1,to recover at the SVi(Ui) outlet the purified and / or separated product contained in said initial solution; to circulate said purified and / or separated product from the SVi(Ui) outlet of subunit Ui in channel V1, from SVi(Ui) to the EVI(U2) inlet of subunit U2, c) in subunit U2: to circulate the liquid cathode CAL(2) from the E2O inlet to the 82(1) outlet, o circulate the liquid anode ANL(2) from the inlet E2(n2) to the outlet 82(012), o circulate said purified and / or separated product from the outlet SVi(Ui) of the subunit U1 and at least one buffer solution in the subunit U2, between the liquid cathode CAL(2) and the liquid anode ANL(2), from the inlets E2(2) to E2(n2-1) to the outlets 82(2) to 82(012-1), d) recover at one of the outlets 82(2) to 82(012-1) the purified and / or separated product.

10. Cell according to any one of claims 8 or 9, wherein the number of outputs r of unit U1 is greater than or equal to 5, further comprising an intermediate channel W1 connecting subunit U1 to subunit U2, extending: o from one of the outputs SWi(Ui) of unit U1 selected from Si(2) to Si(mi-1), different from the output SVi(Ui) of channel V1, o to the input EVI(U2) of subunit U2.

11. Cell according to any one of claims 8 to 10, wherein the number of outputs r and m2 of subunits U1 and U2 is greater respectively than the number of inputs ni and 02 of subunits U1 and U2; and / or in which the subunits U1 and U2 have width and length dimensions such that the width La2 of subunit U2 is equal to the width Lai of subunit U1 and the length L02 of subunit U2 is equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is greater than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is less than or equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is less than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is greater than or equal to the length L01 of subunit Ui,or in which the width La2 of subunit U2 is less than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is less than or equal to the length L01 of subunit Ui, or in which the width La2 of subunit U2 is greater than or equal to the width Lai of subunit U1 and the length L02 of subunit U2 is greater than or equal to the length L01 of subunit U1.

12. Cell according to any one of claims 1 to 7, wherein s is equal to 3, the electrophoresis chamber comprising three subunits U1, U2 and U3, subunit U1 comprising a portion hollowed out in a plate X ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, bi, ci, di) and 2 upper and lower faces (ei, fi), ■ said hollowed-out part being of length L01 and width Lai, ■ of height hi corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces (ai , bi) being parallel to each other, the face (ai) being delimited by two edges (A1 i, A2i) of dimension Lai and the face (bi) being delimited by two edges (B11 , B2i) of dimension Lai , ■ the lateral faces (ci, di) being parallel to each other, the face (m) being delimited by two edges (C11, C2i) of dimension L01 and the face (di) being delimited by two edges (D11 , D2i) of dimension L01 , neither successive entries Ei(1), Ei(2) to Ei(ni-1), Ei(ni), nor being an integer from 4 to 9, distributed on the face (ai) between A11 and A2i and aligned along a direction parallel to A11 and A2i, o mi successive outputs from Si (1 ), Si(2) to Si(mi-1), Si(mi), where mi is an integer from 4 to 12, distributed on the face (bi) between B1 i and B2i and aligned along a direction parallel to B1 i and B2i, such that Si(1) faces Ei(1) and Si(mi) faces Ei(ni) along a direction parallel to C1 i and D1 i, the subunit U2 comprises o a hollowed-out part in a plate X ■ in the shape of a rectangular parallelepiped with 4 lateral faces (ai, b2, C2, d2) and 2 upper and lower faces (e2, f2), ■ said hollowed-out part being of length L02 and width La2, ■ of height i2 corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces ai, b2) being parallel to each other, the face (32) being delimited by two edges (AI2, A22) of dimension La2 and the face (b2) being delimited by two edges (BI2, B22) of dimension La2, ■ the lateral faces (C2, d2) being parallel to each other, face (C2) being delimited by two edges (CI2, C22) of dimension L02 and face (d2) being delimited by two edges (DI2, D22) of dimension L02, o n2 successive inputs E2(0), E2(2) to E2(n2-1), E(n2), n2 being an integer from 4 to 9, distributed on face (32) between AI2 and / 2i and aligned along a direction parallel to AI2 and A22, o m2 successive outputs from S2(1), S2(2) to 82(012-1), 82(012), m2 being an integer from 4 to 12, distributed on face (b2) between BI2 and B22 and aligned along a direction parallel to BI2 and B22, such that 82(1) faces E2O and 82(012) faces £2(02) in a direction parallel to CI2 and DI2, subunit U3 comprises a hollowed-out portion in a plate X ■ in the shape of a rectangular parallelepiped with 4 lateral faces (33, bs, C3, ds) and 2 upper and lower faces (es, fs), ■ said hollowed-out part being of length Los and width Las, ■ of height hs corresponding to the thickness of the X-ray electrophoresis plate, from 25 µm to 20 mm, ■ the lateral faces (33, bs) being parallel to each other, the face (33) being delimited by two edges (Aïs, A2s) of dimension Las and the face (bs) being delimited by two edges (Bis, B2s) of dimension Las, ■ the lateral faces (es, ds) being parallel to each other, the face (es) being delimited by two edges (CI3, C2s) of dimension L03 and the face (ds) being delimited by two edges (DI3, D2s) of dimension L03, o ns successive inputs Es(1), Es(2) to Es(ns-1), E(ns), ns being an integer from 4 to 9, distributed on the face (33) between Aïs and A2s and aligned along a direction parallel to AI3 and A23, o m3 successive outputs from 83(1), Ss(2) to Ss(ms-1), Ss(ms), m3 being an integer from 4 to 12, distributed on the face (bs) between Bis and B2s and aligned along a direction parallel to Bis and B2s, so that Ss(1) faces Es(1) and Ss(ms) faces Es(ns) in a direction parallel to CI3 and Dis, characterized in that it comprises an intermediate channel V1, connecting the two subunits U1 and U2 by one of the outputs SVi(Ui) chosen from Si(2) to Si(mi-1) of subunit U1 to one of the inputs EVI(U2) of subunit U2 chosen from Es(2) to E2(n2-1), and an intermediate channel V2,connecting the two subunits U2 and U3 via one of the outputs SV2(U2) chosen from 82(2) to 82(012-1) of subunit U2 to one of the inputs EV2(U3) of subunit U3 chosen from Es (2) to Es (ns-1).

13. Free-flow electrophoresis cell device comprising: p plates X between two adjacent plates Y: op being an integer from 1 to 50, in particular from 1 to 20, o X being an electrophoresis plate (1) of inert material, o Y being a sealing plate (2) of sapphire or alumina Al2O3 with 99% a-AhCh, f electrophoresis cells according to any one of claims 1 to 12, t is an integer from 1 to 500, clamping means for all the plates allowing the sealing of said device.

14. Device according to claim 13, comprising at least two electrophoresis cells such that in at least one of their subunit Ut, t being an integer from 1 to (s-1), at least two of their respective outputs open into the same input of the same successive subunit Ut+1 by their respective channel Vt.

15. Device according to any one of claims 13 or 14, wherein each plate X, named Xj, j being an integer ranging from 1 to p, comprises qj electrophoresis cells according to any one of claims 1 to 12, qj being an integer from 1 to 10.

16. Device according to claim 15, wherein p is an integer from 1 to 50, in particular from 1 to 10, when j varies from 1 to p, qj is an integer from 2 to 50, in particular from 2 to 10, comprising p electrophoresis plates Xj each having from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5, wherein for each electrophoresis chamber, the hollowed-out parts of the s subunits are located in the same plate.or in which p is equal to 1 and qi is an integer from 2 to 50, in particular from 2 to 10, comprising a single electrophoresis plate X1 having from 2 to 50 electrophoresis chambers, in particular from 2 to 10 electrophoresis chambers, each electrophoresis chamber comprising s subunits, s representing an integer from 2 to 5, and / or in which in each plate Xj, the qj electrophoresis chambers are aligned such that the subunits of the qj electrophoresis chambers are adjacent to each other by faces (G) or (d).

17. Device according to any one of claims 13 to 16, comprising a vertical succession of plates X, Y, Z whose surfaces are stacked according to the sequence YZY(XYZY) P , in which X represents an electrophoresis plate made of inert material. Y represents a sealed plate made of sapphire or alumina (Al2O3) with 99% a-AhCh, Z represents a cooling plate including a heat transfer system, p, an integer from 1 to 50, in particular from 1 to 20, represents both the number of stages of said device and the number of plates X, each stage being defined: - by the following sequence of plates YZYXYZY, in which: - Plate X is located between two plates Y, - each of the two plates Z being respectively adjacent to a plate Y, - and each of the two Y plates located at the ends of the YZYXYZY sequence, respectively covers a Z plate so that each Z plate is located between two Y plates.

18. A device according to any one of claims 13 to 17, wherein each electrophoresis chamber is configured to contain at least one selectively permeable membrane, in particular selectively permeable in size, positioned so as to be traversed by the solution containing the product to be separated or purified during operation of the device, preferably said at least one A selective permeability membrane is placed in the U subunit sof each electrophoresis chamber; and / or wherein the upper faces ei and / or the lower faces f of the subunits U of each of the electrophoresis chambers comprise at least one protrusion configured not to disturb, during the operation of the device, the circulation in the chamber of the product to be purified and / or separated, and configured to enhance heat transfer and to maintain each of said electrophoresis chambers at a selected temperature, in particular wherein said at least one protrusion is made of thermally conductive material, preferably of sapphire or 99% a-AhCh alumina; and / or said device comprises a supply channel network configured to connect at least one of the inlets Ei(2) to Ei(ni-1) of the subunit Ui of each electrophoresis chamber to an external micro / milli fluidic circuit for supplying an initial solution containing a product to be purified and / or separated.

19. A method for purifying and / or separating, by free-flow electrophoresis, a product to be purified and / or separated contained in an initial solution by implementing an electrophoresis cell device according to any one of claims 13 to 18, and comprising the following steps: a) a purification and / or separation step of said product in the subunits Ui of each electrophoresis chamber, under an electric field CE(1), comprising: in the subunit Ui of each electrophoresis chamber: o the circulation of a liquid cathode CAL(1) from the inlet Ei(1) to the outlet Si(1), o the circulation of a liquid anode ANL(1) from the inlet Ei(ni) to the outlet Si(mi), o the circulation of the initial solution containing the product to be separated and / or purified and at least one buffer solution in the subunit Ui, between the liquid cathode CAL(1) and the liquid anode ANL(1), from inputs Ei(2) to Ei(ni-1) to outputs Si(2) to Si(mi-1),to obtain the purified and / or separated product from the subunit Ui, b) (s-1) polishing step(s) k, k varying from 1 to (s-1), each step k comprising each: i) in the intermediate channel Vk of each electrophoresis chamber, o recovery at the outlet SVk(Uk) corresponding to the intermediate channel Vk of said purified and / or separated product from the subunit Uk; o the circulation, in the absence of an electric field, of the aforementioned purified and / or separated product, from the outlet SVk(Uk) of the subunit Uk corresponding to the intermediate channel Vk to the inlet EVk(Uk+i) of the subunit Uk+i corresponding to the intermediate channel Vk, ii) in the subunit Uk+i of each electrophoresis chamber, in the presence of an electric field CE(k+1): o the circulation of a liquid cathode CAL(k+1) from the inlet Ek+i (1 ) to the outlet Sk+i (1 ), o the circulation of a liquid anode ANL(k+1) from the inlet Ek+i(nk+i) to the outlet Sk+i(rrik+i),o the circulation of said purified and / or separated product from the outlet of subunit Uk and at least one buffer solution in subunit Uk+i, between the liquid cathode CAL(k+1) and the liquid anode ANL(k+1), from inlets Ek+i(2) to Ek+i(nk+i-1) to outlets Sk+i(2) to Sk+i(mk+i-1), to obtain the product (k+1) times purified and / or separated from subunit Uk+i; c) a recovery step comprising: recovery at one of the outlets S, s (2) to S s (m s -1) of unit U s from each electrophoresis chamber of the product(s) once purified and / or separated.

20. Use of an electrophoresis cell according to any one of claims 1 to 12 or of a device according to any one of claims 13 to 19, in the implementation of a process for the purification and / or separation by free-flow electrophoresis of a product to be purified and / or separated contained in an initial solution, comprising a first purification and / or separation step in the Ui subunit(s) followed by successive (s-1) polishing steps in the U2 to U subunits s of electrophoresis chamber(s).

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