Method for selective separation of amphoteric substances

Abstract

The present invention relates to a method for the selective isolation of amphoteric substances, in particular biomolecules or biomolecule complexes (e.g., molecule-containing carriers), based on the isoelectric points of the amphoteric substances. For selective separation, an electrically conductive and / or electrically charged porous surface comprising an electrically conductive coating or layer is provided. The fluid containing the amphoteric substance is in contact with the surface, thereby adsorbing the substance to the surface due to the attractive interaction force. Thereafter, at least a portion of the adsorbent substance is released by applying the selected release voltage such that the net interaction force between the surface and the adsorbent substance transforms from an attractive force to a repulsive force. Selectivity is achieved by selective adsorption (so that at least one substance is not adsorbed) and / or selective desorption (so that some previously adsorbed substances are selectively released while other substances still bind to the surface, preferably only one previously adsorbed substance is selectively released).

Description

[0001] The present invention relates to a method for the selective separation of amphoteric species by means of selective (electro)adsorption and electrodesorption, in particular (bio)molecules such as proteins, amino acids or nucleic acids, or molecular complexes such as vectors (in particular capsids, plasmids or vesicles), respectively, molecules such as genomes contained in the molecular complexes (in particular vectors) with respect to their isoelectric points, in particular for the preparation of high-purity samples of the desired amphoteric species (in particular vectors and / or biomolecules).

[0002] Generally, biomolecule is the material that is highly sought after in medical applications, for example, as the protein of medicine or analogue. Therefore, it is necessary to produce the required biomolecule with high purity, therefore it is necessary to separate it from the impurities produced in these biomolecule formation processes. Usually this separation is carried out by combining-elution process, wherein the required molecule is bound to for example exchange resin, gel or film, subsequently eluted from the resin, gel or film by the mode of pH value change (for example, acid elution is carried out using acetate buffer solution of pH 3), and / or eluted from the resin, gel or film using saline solution with the biomolecule required for salt ion replacement. Both of these elution methods may cause severe conditions, which may be harmful to any biomolecule, for example, may cause aggregation or denaturation of required substance (particularly with regard to protein). The example of this method is isoelectric focusing chromatography or ion exchange chromatography.

[0003] Particularly interesting substances in the biomedical field are vectors. Such vectors are often used as a means for delivering molecules, particularly biologically active substances, to cells. Therefore, vectors play a key role in gene therapy and vaccines, or in other therapeutic areas, including, for example, personalized cancer therapy. To this end, vectors containing molecules, particularly molecules in the form of biologically active substances, such as genomes in the form of deoxyribonucleic acid (DNA), single-stranded deoxyribonucleic acid (ssDNA), ribonucleic acid (RNA), or single-stranded ribonucleic acid (ssRNA), are prepared.

[0004] Typically, a vector such as a retrovirus, slow virus, adenovirus or adeno-associated virus is used as a viral vector. For all of these, capsid protein forms a shell in which molecules, particularly bioactive substances such as drugs and / or the genomes of DNA, RNA, ssDNA or ssRNA forms, can be placed. Therefore, capsid is used, for example, as a mode for delivering bioactive substances, particularly as a protective shell for molecules to be delivered, particularly capsid can be used as a vector for artificially carrying bioactive substances (such as nucleic acid sequences, typically DNA or RNA) to cells, for example, for expression or replication. Plasmids are typically small, circular, mainly double-stranded DNA or RNA molecules found primarily in bacteria, and plasmids are also commonly used as vectors. Regarding gene therapy, plasmids are primarily used by integrating the desired genomic sequence into the plasmid itself. Then, plasmids can be migrated into the cell and can participate in the replication or similar processes of the integrated genomic sequence.

[0005] For gene therapy and other medical applications, it is desirable to use highly pure vectors. However, when preparing vectors containing molecules, the molecules contained in the vectors may generally vary, particularly in terms of molecular content and / or the molecules contained. For example, a specific molecule may be contained or only a portion of such a molecule, or some vectors may not contain any molecules at all. For example, when preparing capsids containing genomes, vectors are generally obtained that contain complete genomes or partial genomes, or even no genome at all. In order to use such drugs in therapy, it is necessary to selectively separate vectors containing the correct (particularly complete) genome or molecule in the correct amount from vectors that do not meet (particularly not fully meet) these conditions.

[0006] In order to separate vectors containing a complete genome from vectors containing a partial genome or no genome, several techniques are known, such as anion exchange high performance liquid chromatography (AEX HPLC), capillary isoelectric focusing (cIF), ultracentrifugation (UCF), or gel isoelectric focusing (gIF). Among them, AEX HPLC, cIF, and gIF are used as preparation techniques in the manufacture of vectors for gene therapy, etc., while UCF is only commonly used in analysis. In particular, these techniques are not limited to vectors, but can be applied to general amphiphilic substances, especially biomolecules or biomolecule complexes.

[0007] Both AEX HPLC and IF methods are based on the overall charge of the capsids. The overall charge of the capsids is a function of the amount of genome they contain relative to the pH of the surrounding environment. However, the differences in the isoelectric points of intact, partial, and empty capsids are small, corresponding to differences in the range of, for example, 0.1.

[0008] In the AEX HPLC method, the capsid, which has a total cationic charge and therefore a net total charge of positive polarity, is adsorbed to a chemically negatively charged surface of a resin, for example, containing anionic binding sites. To elute the vector bound to the anionic binding sites, the pH of the environment is typically altered to change the net total charge of the vector from negative to neutral or positive, or salts are used to displace the bound vector by removing it from the binding site and effectively blocking these bindings due to higher affinity, thereby releasing the vector. For example, MgCl2 and / or NaCl solutions can be used as eluents. However, the vector released with such salt solutions must then be separated from the salts, as higher concentrations of such salts can be harmful in medical applications. In addition, elution by altering the pH of the environment can damage the vector itself, as the capsid protein can flocculate, especially if the capsid protein becomes neutral.

[0009] Similarly, isoelectric focusing techniques such as gel isoelectric focusing or capillary isoelectric focusing face the same challenges and limitations compared to AEX HPLC methods. In this technique, for example, a gel consisting of several regions with different pH values is prepared. A solution containing amphiphilic biomolecules with different isoelectric points is brought into contact with the gel. The regions of varying pH values are arranged in an increasing gradient along a single axis. An electric field is applied within the axis. The amphiphilic biomolecules then migrate along the electric field according to their charge. When the molecules reach a region with a pH equal to their isoelectric point, their charge becomes neutral and they cease movement along the axis, thus separating the biomolecules according to their isoelectric points. However, due to the neutralization of the biomolecules' charge, flocculation may occur, potentially damaging the biomolecules.

[0010] Furthermore, these techniques are often limited in terms of purity due to slight differences in isoelectric points. Typically, both AEXHPLC and IF methods produce preparations with, for example, capsids containing intact genomes (70% with the remainder mostly consisting of capsids with partial genomes or empty capsids). Therefore, achieving a higher degree of separation between these vectors is desirable to obtain capsid preparations with higher purity.

[0011] Other methods for separating biomolecules generally include filtration by electrosorption methods. Among them, EP3115099B1 discloses a method for enhancing the retention of biomolecules by a metal-coated polymer membrane by applying a voltage to the membrane. In addition, WO2021 / 084080A1 discloses a method for electrodesorbing molecules adsorbed to a charged membrane coated with a metal coating by applying a voltage of opposite polarity to the membrane charge to the coating. However, for biomolecules that are only slightly different, in particular for separating vectors containing complete genomes, partial genomes, or no genome, this method does not provide the desired selectivity.

[0012] Based on this, the task to be accomplished is to provide a suitable method for separating amphiphilic substances, in particular biomolecules or biomolecular complexes, in particular supports, which may differ slightly but still provide a high-purity separation, wherein desorption by changing the pH value and / or by applying salt solutions is avoided, thereby eliminating the potential hazards of degradation of the separated biomolecules and biomolecular complexes, in particular if degradation by any means could represent a health risk in medical applications or similar applications.

[0013] The object of the invention is achieved by a method according to claim 1. The subsequent claims 2 to 25 provide advantageous embodiments of the method according to the invention.

[0014] The present invention provides a method for selectively separating amphoteric substances, particularly based on their isoelectric points. The substances to be separated consist of at least two specific substances. Each specific substance has a specific isoelectric point. Therefore, particularly based on their specific isoelectric points, each specific substance has a specific net total charge at a given pH value. The given pH value can particularly be a predetermined pH value.

[0015] The given pH value is preferably in the range of pH 6 to pH 9, preferably in the range of pH 6.5 to pH 8.5, particularly preferably in the range of pH 7 to pH 8, which generally minimizes the risk of substances that may be used therapeutically or happen to be naturally present in the human body.

[0016] In this context, the term "species" may refer to any (bio)molecule or (bio)molecule complex. (Bio)molecules that may be isolated using the present invention may include, but are not limited to, amino acids, proteins, nucleic acids such as RNA or DNA.

[0017] A (bio)molecular complex may include, but is not limited to, a carrier containing a molecule, in particular a carrier containing a biomolecule.

[0018] The term "molecule-containing carriers" refers to carriers in which at least some of the carriers contain molecules, and for some of the carriers, the molecules are different, for example, in terms of content and / or type, thereby providing at least two groups of carriers with different contained molecules as at least two specific substances to be separated.

[0019] In this context, the term "having different contained molecules" includes different molecules and / or different amounts of molecules, in particular different amounts of the same molecule and / or different parts of the same molecule and / or different specific molecules. In particular, the term "different amounts of molecules" may include zero molecular content. Thus, some vectors may not contain any molecules at all, and thus "empty" vectors may form a group of vectors or may be part of a group of vectors.

[0020] In particular, the vectors consist exclusively of vectors of the same type and / or have the same base structure, in particular, contain no molecules. In particular, the vectors consist of capsids and / or plasmids. In particular, the vectors are viral vectors, for example based on retroviral, lentiviral, adenoviral or adeno-associated viral vectors.

[0021] Any vector has a basic isoelectric point and therefore a basic net total charge at a given pH value, in particular at the pH value of the surrounding environment. In particular, the basic isoelectric point corresponds to the isoelectric point of an empty vector, and therefore is a vector without molecules. Similarly, the basic net total charge is the net total charge of an empty vector. The basic isoelectric point and the basic net total charge of any particular vector containing molecules are shifted by the molecules contained in the particular vector. Thus, due to the molecules contained in the particular vector, a shifted isoelectric point that is different from the basic isoelectric point and a shifted net total charge at a given pH value that is different from the basic total charge at the given pH value are generated. Thus, in particular based on the molecules contained in each group of vectors, in particular for different molecules contained and / or different amounts of molecules contained, at least two groups of vectors are defined. In particular, the different molecules can be different types of molecules and / or different parts of the same molecule and / or molecules of the same type.

[0022] In particular, in at least two groups of carriers, the first group of carriers has a first isoelectric point and / or a first isoelectric point range and / or a first total charge and / or a first net total charge range at a given pH value, and the second group of carriers has a second isoelectric point and / or a second isoelectric point range and / or a second net total charge and / or a second net total charge range at a given pH value. In particular, the groups defined by the isoelectric point ranges and / or net total charge ranges can be non-homogeneous, in particular, they can be composed of more than one type of carrier and / or carriers containing different molecules and / or different amounts of molecules. In particular, the electronic structure of the carriers can be affected by the presence of molecules. Such an effect can result in a shift in the isoelectric point and / or net total charge of the carriers containing the molecules. In particular, the shifted isoelectric point and / or shifted net total charge can be grouped into ranges, wherein it is desirable to separate the carriers into groups of carriers falling within these ranges. In particular, separation into several groups may be unnecessary for undesirable impurities. Thus, if applicable, a range of isoelectric points and / or net total charges can be defined, which consist of a variety of different impurities (e.g., empty carrier and partially filled carrier).

[0023] In particular, by carrying out the method according to the invention, the at least two groups of carriers are selectively separated from one another.

[0024] In another embodiment, the method according to the present invention is used to separate groups of substances. In these groups of substances, each group can be composed of more than one specific substance. In particular, this substance can be a similar substance with only slight differences, for example, with the same medical applicability or similar biomolecules, for example, an enantiomer without enantiomeric specific application, but in particular can be a protein variant or analog as a part of a protein family, in particular if these substances do not need to be used separately in medical applications. For this group, the specific isoelectric point and / or specific net total charge of the substance of this group can be a range of isoelectric points and / or a range of net total charge, in particular a disjunct range, for example, one group has a first range, and a second group has different ranges, in particular a disjunct range. In particular, this specific substance group can be limited by this range of isoelectric points and / or net total charge.

[0025] The method comprises the following steps:

[0026] a) providing a charged porous surface, in particular a chemically and / or electrically charged porous surface, wherein the charged porous surface is electrically conductive and / or comprises an electrically conductive coating or an electrically conductive layer, wherein in particular the electrically conductive layer can preferably be integrated into the porous surface and / or formed integrally in the porous surface, or it can be a different layer, in particular a non-integrated layer;

[0027] b) bringing a fluid having a given pH value into contact with a charged porous surface, the fluid containing a substance (particularly a carrier containing molecules), thereby adsorbing at least a first substance of the at least two substances or at least a first group of substances (particularly a first carrier or a first group of carriers of the at least two groups of carriers) to the charged porous surface, the adsorption being due to an interaction force between the net total charge of the adsorbed substance (particularly the carrier) and the charged porous surface, the interaction force being an attractive force;

[0028] c) preferably passing at least a portion of the fluid over and / or through the charged porous surface; and

[0029] d) applying a release voltage to the electrically conductive charged porous surface and / or the electrically conductive coating or layer, the release voltage being selected such that the net interaction force between the charged surface and at least a portion of the adsorbed species (particularly the carrier) changes from an attractive force to a repulsive force, thereby releasing said portion of the adsorbed species.

[0030] Advantageously, the porous surface can be formed, in particular at at least one site of one or more filter membranes (in particular polymer membranes), by one or more filter membranes (in particular polymer membranes) or one or more non-woven substrates having a conductive coating or conductive layer (in particular a metal coating or metal layer or a corresponding coating or layer formed from another conductive material). In particular, the metal coating or metal layer is also porous. Alternatively, the porous surface can be provided by at least one metal membrane. Preferably, gold or platinum is used as the material for the conductive metal coating and / or metal layer. Preferably, the metal used is pure and / or the coating or layer consists of only a single metal. In particular, the polymer membrane itself is non-conductive and only the coating is conductive. In particular, only one conductive material and / or metal is used for the charged porous surface. In particular, the conductive coating or conductive layer is not formed by a metal mesh, in particular no metal mesh is coated with gold or platinum. This is particularly advantageous for pleating or winding using at least one membrane as the porous surface (as described later in the present invention), said at least one membrane being coated, in particular coated with a metal, most preferably a metal coating using gold or platinum as the metal.

[0031] The term "charged surface" refers to a surface having an electric charge, in particular an electric charge of different polarities. In this context, a chemically charged surface can be achieved, for example, by placing ionic groups and / or compounds on the surface, in particular by chemical bonding of, for example, quaternary ammonium groups (in particular for achieving a positively charged surface), or carboxymethyl groups (in particular for achieving a negatively charged surface), or by chemical bonding of compounds containing such groups to the surface. For this purpose, an anion exchange membrane or a cation exchange membrane can be used as a charged porous surface, in particular with an additional conductive coating or conductive layer, in particular, for example, a metallic coating or metallic layer, on the surface. In particular, if the membrane is conductive, for example, if a metallic membrane is used, a charged porous surface can be achieved by applying a voltage (in particular a base voltage) applied to the conductive (in particular metallic) coating or layer or the membrane itself. In particular, charging the surface can be used to adjust the charge of the chemically charged membrane, thereby enhancing or reducing the total charge of the charged porous surface. In particular, the porous surface is not formed by a gel or resin. In particular, the charged surface can also be chemically charged and electrically charged. In this respect, a chemically charged surface, such as an ion exchange membrane, is provided in step a), on which a conductive coating is provided (in particular a membrane), to which a base voltage is applied. Thus, the total charge of the charged surface can be constituted by a combination of the provided chemical charge and the applied charge. Thus, the charge applied by the base voltage can be used to shape, influence or change the chemical "pre-charge" of the surface. In this respect, the applied charges can have the same polarity, thereby enhancing the "pre-charge". If opposite polarities are used, the "pre-charge" may be reduced or even ineffective, or the polarity of the total charge of the charged surface may be transformed into a polarity opposite to the "pre-charge".

[0032] In some embodiments, the present invention provides a plurality of electrodes that are provided on the porous surface of the present invention.Particularly, for applying voltage, particularly releasing voltage and / or basic voltage, using the charged porous surface of conduction and / or the conductive coating or the conductive layer of the charged porous surface can be a part of the charged porous surface as the first electrode and / or the first electrode, in addition, a second electrode is also provided as a counter electrode.The second electrode also can be formed by another conductive coating or conductive layer.Particularly, the second electrode can preferably be arranged on the second side relative to the first side of the porous surface formed by the conductive coating or the conductive layer of the first electrode.In this respect, the first electrode and the second electrode can be arranged on different porous surfaces (particularly films).For example, the first electrode can be set up by the conductive coating of the first porous membrane, and the second electrode can be set up by the conductive coating of the second porous membrane.

[0033] In particular, the applied voltage, in particular the base voltage and / or the release voltage, is in the range of 0.1 V to 50 V, preferably in the range of 0.1 V to 3 V, in particular 2.5 V and / or above 2 V. In particular, a voltage exceeding 3 V (in particular a voltage exceeding 2 V in some embodiments) is applied only for a short time (in particular typically less than 2 seconds).

[0034] In particular, during step d), in particular before step d), a further fluid is brought into contact with the charged porous surface, which further fluid may be of the same or different kind, in particular having a given pH value and / or a pH value which differs from the given pH value by less than 1, in particular by 0.5, in particular by 0.2, in particular by 0.1, and / or does not contain any substance. At least part of the further fluid may pass on and / or through the porous surface. Furthermore, the fluid collected in step d), in particular the fluid passed on and / or through the porous surface, and which contains the substances released in step d), in particular (bio)molecules or carriers containing (bio)molecules, in particular only one specific substance, in particular (bio)molecules of the same or identical type or carriers of a single carrier group, is selectively separated from the remaining specific substances, in particular from the biomolecules of the single type from different biomolecules or from the carriers of the single carrier group from other carriers. A single specific substance, in particular a single vector group, in particular a group of vectors that is homogeneous and / or contains a specific molecule or has a specific isoelectric point, in particular different isoelectric points, in particular not to be described as having an isoelectric point range and / or not being an isoelectric point range, and therefore in particular consists of the same substance, in particular the same molecule or vectors containing the same type of molecule (containing the same molecule and / or containing the molecule in the same amount). In this context, the term same does not necessarily mean completely identical, but also includes substantially identical, in particular including molecules / vectors that are considered to be the same by a person skilled in the art, for example due to the same molecular weight, the same functionality and / or functional groups, for example different enantiomers of the same molecule for medical applications where enantiomeric purity is not required.

[0035] pH values deviating from the given pH are preferably in the range of pH 6 to pH 9, preferably in the range of pH 6.5 to pH 8.5, particularly preferably in the range of pH 7 to pH 8, which generally poses the least risk to substances which may be used therapeutically or which occur naturally in the human body.

[0036] Particularly, can repeat step b) and step c) in the form of multiple circulations, particularly before carrying out step d).Carry out this multiple circulation and can be used for enriching the desired substance adsorbed on the membrane.Usually, in the preparation process of material, the concentration in the fluid is lower.Therefore, the amount of the fluid containing material used is usually very high, to combine the material of required amount, but can by using less other fluid in step d) and / or in step c), allow the major part of the used fluid to pass and / or pass through the surface, remove the fluid that contacts, passes through and / or passes through simultaneously and be discharged into the less-volume fluid in step d) subsequently and reduce the amount of used fluid.

[0037] Furthermore, all steps b) to d) can be repeated in multiple cycles. This can be advantageous if various impurities are present during the preparation of the desired substance. For example, in this context, carriers containing molecules with an undesirable molecular content, carriers completely devoid of molecules, or carriers containing undesirable molecules can be considered impurities. These impurities are separated from the desired carrier containing the molecules. In general, pure products are particularly desirable for (bio)molecules, especially those used in medical applications.

[0038] By performing multiple cycles, for example, in a first cycle, a first impurity can be selectively separated from a desired carrier, while a second impurity is not separated. Then, in a second cycle with adjusted process conditions, such as at another given pH and / or with an adjusted voltage, the desired carrier can be separated from the second impurity. Furthermore, if the selective separation is incomplete, performing multiple cycles can achieve a higher purity separation.

[0039] In particular, for vectors containing molecules, at least two groups of vectors may contain different amounts of the molecules, so that the vectors of a first group contain a first amount of molecules and the vectors of a second group contain a second amount of molecules.

[0040] In one advantageous embodiment (particularly from a gene therapy perspective), the molecule contained in the vector is a genome, particularly RNA or DNA. Advantageously, one of the at least two groups of vectors can consist of vectors containing a complete genome, while at least another of the at least two groups of vectors can consist of vectors containing partial genomes and / or vectors containing no genome. In particular, vectors containing partial genomes and vectors containing no genome can form separate groups of vectors, but can also be considered as a single group, for which the isoelectric point range and net total charge must be selected accordingly.

[0041] In an alternative embodiment, the contained molecule may be a drug or a protein or another biologically active substance.

[0042] In terms of pH value, acidic pH value or alkaline pH value may be harmful to biomolecules in general. For example, proteins may aggregate or similar phenomena under acidic pH value, for example, under pH value less than pH 5, particularly using buffer solutions with these pH values. For example, the pH value of human blood is about 7.4, which is close to neutral pH value. Therefore, neutral pH value is preferably used for all fluids used in the method of the present invention, particularly between pH 6 and pH 8, most preferably pH 7.4. Advantageously, the pH value remains constant and / or remains unchanged throughout step b) to step d), particularly for all fluids used in the method and / or on charged porous surface and / or around charged porous surface and / or fluids on charged porous surface and / or around charged porous surface. In this context, the pH value with negligible difference is referred to as constant value, for example, pH value accurate to the decimal point. In this regard, any voltage applied can, for example, cause local pH value changes due to the substance being locally protonated or deprotonated. With respect to the overall pH value of the fluid, this effect is negligible and is therefore still referred to as a constant value. However, when cycling, in particular when comprising a complete cycle from step b) to step d), the pH value can be kept constant within a single cycle, but can be adjusted between cycles, for example by cycling at pH 8 for the first cycle and cycling at pH 6 for the second cycle. In this respect, the change in pH value is not used to effect any release of previously adsorbed substances. However, the pH value can be selected so that it is very close to the isoelectric point of any adsorbed amphoteric substance (in particular a carrier), but does not cause release, and in particular the pH value can be adjusted accordingly between cycles, in particular for different substances (in particular a carrier) released within any cycle. In this respect, a pH value very close to the isoelectric point of any specific substance to be released (in particular a carrier) (in particular the difference between the pH value and the isoelectric point is less than 1, in particular less than 0.5), but does not cause release, is selected, which can generally particularly reduce the release voltage required for initiating the release of the specific substance (in particular a carrier) in step d).

[0043] In particular, saline solution (especially high concentration saline solution) is not used to release previously adsorbed substances, especially if the substance is a biomolecule such as a carrier. In particular, all fluids used are "salt-free" fluids. In this context, liquids with negligible salt ion concentrations are considered to be "salt-free". In particular, in the context of the present invention, common pH buffer solutions of pH 6 to pH 8 are considered to be "salt-free". In this respect, the liquid (especially all liquids) introduced in the method preferably can be or include such a buffer solution for maintaining a constant pH value and / or a small amount of ions and / or a small amount of salt, preferably less than 150 mM. Therefore, these buffer solutions contain a small amount of salt, and these contents are lower than the concentration of the saline solution commonly used for ordinary salt elution. In particular, these buffer solutions (such as phosphate buffered saline (PBS)) can be particularly isotonic, therefore providing an osmotic pressure and ion concentration that match the human body, which may be particularly beneficial to medical-related applications, especially for biomolecules (such as carriers) as substances. In particular, PBS used in common concentrations generally provides isotonic conditions, in particular with an osmotic pressure and ion concentration that match those of the human body. In particular, the substance release in step d) is performed solely by applying the release voltage. However, changing the pH value can be used for cleaning purposes, for example if, after separation and / or recovery of the desired substance (in particular the carrier), any unwanted impurities remain bound to the surface, in particular after step d), in particular if the method is cyclically carried out after step d) of the last cycle.

[0044] According to the invention, the selective separation of at least two substances is carried out in particular by the following steps:

[0045] I. in step b), selectively adsorbing at least a portion of a first substance of at least two substances, in particular selectively adsorbing a first group of at least two groups of carriers, while adsorbing at least another substance of at least two substances, in particular adsorbing another group of at least two groups of carriers, in particular not adsorbing other substances, in particular not adsorbing other groups of at least two groups; and / or

[0046] II.i) in step b), adsorbing a first substance and at least part of at least one second substance of at least two substances, in particular a first group of at least two carriers and at least one second group of carriers, in particular all substances of at least two substances, in particular a group of at least two groups of carriers, and

[0047] ii.) In step d), a first substance, in particular a first group of supports, is selectively released, wherein at least one further substance, in particular a group of supports, adsorbed in step b) remains, in particular all other substances, in particular all other groups of supports, adsorbed in step b) remain adsorbed.

[0048] In this regard, the selectivity of the separation according to Option I is not limited to the adsorption of only the substance as the first substance, but multiple substances can be adsorbed in step b). In this regard, at least one of the at least two substances must not be adsorbed in step b) to achieve the selectivity according to Selectivity Concept I. Therefore, Option I specifically includes the adsorption of a group of substances consisting of more than one substance, in particular the adsorption of a first substance group, while not adsorbing at least another substance, in particular a second substance group consisting of one or more other substances different from the first substance group. In this regard, the first substance group and the second substance group are selectively separated. However, in order to selectively separate the substances contained in the first group, this method can additionally use selective desorption according to Concept II. In step d) and / or can be carried out in multiple cycles, for example, the general process conditions can be adjusted between cycles (for example, cycles using different predetermined pH values or the like).

[0049] In particular, regarding the separation of substance groups (in particular carrier groups) of at least two substance groups (in particular carriers), the first substance group (in particular carriers) has a first isoelectric point and / or a first isoelectric value range and / or a first net total charge and / or a first net total charge range at a given pH value and the second substance group (in particular carriers) has a second isoelectric point and / or a second isoelectric value range and / or a second net total charge and / or a second net total charge range at a given pH value.

[0050] In a particularly advantageous embodiment, the given pH value is selected such that, at the given pH value selected, the net total charge of the first and second substances of the at least two substances or two groups of substances, or the first and second groups of substances (in particular the support), is of opposite polarity to the charge of the charged porous surface, and the net total charge of the other of the first and second groups is zero or has the same polarity as the charge of the charged porous surface, so that, in particular, only the first and second substances, or one of the first and second groups of substances (in particular the support) is adsorbed in steps b) and c). In this embodiment, in particular, the selective separation is performed according to option I. Advantageously, only the desired substance (in particular the support containing the desired molecule or the desired molecule content) is adsorbed, or alternatively, all substances except the desired substance (in particular the support containing the desired molecule or the desired molecule content) are adsorbed, so that the desired substance (in particular the support containing the desired molecule or the desired molecule content) is the only support not adsorbed to the surface in step b).

[0051] Advantageously, the given pH value is selected such that it is between the isoelectric point of the first substance (particularly the group of carriers) and the isoelectric point of the second substance (particularly the group of carriers), such that the given pH value is within the range of the first isoelectric point and the second isoelectric point, such that the given pH value is within the range of the first isoelectric point and the second isoelectric point, and / or between the second isoelectric point and the range of the first isoelectric point. In particular, at this selected given pH value, the first substance and the second substance (particularly the group of substances (particularly the carriers)) differ in the polarity of their net total charge. Thus, in step b), since one specific substance or group of substances (particularly the group consisting of the carriers) has a polarity opposite to the surface charge and the other specific substance or group of substances (particularly the group consisting of the carriers) has a polarity identical to the surface charge, only one specific substance or group of substances (particularly a single group of carriers from the first and second groups) is adsorbed in step b). Since only substances with polarity opposite to the surface charge can be adsorbed, one of the substances (particularly a group of substances (particularly carriers)) is selectively adsorbed to the surface, while the other of the first substance and the second substance, or the other of the first substance group and the second substance group, is not adsorbed to the surface, and is particularly removed in step c) together with the fluid that contacts, flows over and / or passes through the surface.

[0052] In another particularly advantageous embodiment, both the first and second substances, the first and second substance groups (particularly the carrier (particularly all substances)), and the substance (particularly the substance (particularly the carrier) group) are adsorbed onto a charged porous surface. This can correspond to a selective separation according to Option II. Advantageously, the given pH value is selected such that the selected pH value is above or below the isoelectric point or isoelectric point range of the first and second substances, the first and second substance groups (particularly all substances, all substance (particularly the carrier) group). In particular, at the given pH value, the net total charge of the first and second substances, the first and second substance groups (particularly the carrier (particularly all substances, substance groups) for the first and second groups) has the same polarity, which is opposite to the polarity of the charge of the charged porous surface.

[0053] Advantageously, the release voltages of the first and second substances, the first and second substance groups (particularly the carriers adsorbing the first and second groups) used for adsorption in step d) are different. By applying a first release voltage to the conductive surface and / or the conductive coating or conductive layer, the net interaction force of one of the first and second substances, one of the first substance (particularly the carrier) group and the second substance group (particularly a single group of all substances (particularly the carrier) group) is converted into a repulsive force, while the net interaction force of the other of the first and second substances, another of the first and second substance groups (particularly all groups) still adsorbed on the surface is maintained as an attractive force, thereby selectively desorbing only the first or second substance, the first and second substance groups (particularly the carrier) group, while other substances, other substance groups (particularly all other groups) remain adsorbed on the charged porous surface.

[0054] Subsequently, before desorbing the previously adsorbed one of the first and second substances, one of the first substance (particularly carrier) group and the second substance group, or one of the first carrier group and the second carrier group by applying a first release voltage, the applied voltage is advantageously adjusted to a second release voltage, wherein the net interaction force between the other of the first and second substances, the first substance (particularly carrier) group and the second substance group is converted from an attractive force to a repulsive force by applying the second release voltage, thereby desorbing the other of the first and second substances, the first substance (particularly carrier) group and the second substance group. In particular, if more than two substances, or more than two substance (particularly carrier) groups, are adsorbed to the surface, the first and second release voltages are selected such that a third substance, or a third substance (particularly carrier) group, can remain adsorbed while desorbing the first and second groups of carriers.

[0055] In another advantageous embodiment, the porous surface is charged by applying a base voltage. The desorption in step d) can then be initiated by changing (in particular reducing) the base voltage to a release voltage, in particular by gradually changing (e.g. reducing) the voltage to a first release voltage and then to a second release voltage. "Changing or reducing the voltage" can include a change in polarity, thus, for example, in particular gradually changing from a negative voltage to a positive voltage or vice versa.

[0056] In particular, it may be advantageous to combine options I and II. To achieve a selective separation, in step b), in the presence of two or more substances or two or more groups of substances (in particular carriers), a specific substance or a specific group of substances (in particular carriers) is selectively adsorbed, while in step b), the remaining substances or the remaining group of substances are selectively not adsorbed, and subsequently, in step d), at least one of the specific substances previously adsorbed in step b) is selectively released, in particular, all substances are released gradually in step d).

[0057] In another advantageous embodiment, at least two charged porous surfaces are provided in step a), in particular surfaces formed by polymer films. These at least two charged porous surfaces comprise at least a first charged porous surface and a second charged porous surface, in particular polymer films. In addition, the first charged porous surface is electrically conductive and / or comprises an electrically conductive coating or a conductive layer. Optionally, the second charged porous surface is also electrically conductive and / or also comprises an electrically conductive coating or a conductive layer. For example, by having more than one charged porous surface, the overall porous surface can be enhanced, thus in particular if combined with a stacked, in particular pleated, arrangement, in particular according to the disclosure to be found further below, resulting in a higher capacity for binding substances to the surface.

[0058] Preferably, at least two charged porous surfaces (in particular if these surfaces are formed by polymer films) are arranged as a stack formed by stacked layers. In this arrangement, the stack is formed so that the first charged porous surface and / or the conductive coating or conductive layer of the first charged porous surface is placed on the first end of the stack. The counter electrode is preferably arranged at the second end opposite to the first end of the stack. In addition, in particular, a plurality of second charged porous surfaces are preferably placed between the first porous surface and the counter electrode. Particularly advantageously, the second charged porous surface is preferably non-conductive and does not include a conductive coating or conductive layer, in particular chemically charged. Alternatively, the second charged porous layer can be conductive and / or include a conductive coating or conductive layer. The counter electrode is then preferably formed by the second charged porous surface itself or the conductive coating or conductive layer of the second charged porous surface. Therefore, in this arrangement, the second charged porous surface is placed at the second end of the stack.

[0059] Advantageously, between the first charged porous surface and the second charged porous surface, in particular between all charged porous surfaces and / or surfaces forming the stacked layer, no further electrodes are placed, only the surface placed at the first end or the second end is electrically conductive and / or comprises a conductive layer. In particular, at least one polymer film can act as an insulator between the conductive coating or the conductive layer. Alternatively, in addition, a different insulator (for example an insulator that can be formed by another polymer film) is placed between the conductive coating or the conductive layer. Thus, the insulator can in particular be formed by at least one, in particular a plurality, in particular up to 30, preferably 5 to 15 additional polymer films and / or a non-conductive side of a polymer film comprising a conductive coating, in particular forming the non-conductive side of the film of the first surface and the second surface.

[0060] In particular, a stacked arrangement of charged porous surfaces, in particular polymeric membranes, can provide a higher surface area relative to the surface area of the space required for providing a surface (in particular a membrane) in any suitable device for performing the method according to the invention. In particular, this arrangement can also provide a fluid-permeable channel between surfaces (in particular membranes), for example, by a surface (in particular a membrane) being slightly spaced apart from another surface. This arrangement can particularly enable a fluid to flow over and through a surface simultaneously.

[0061] In particular, the disclosed stacking arrangement may preferably consist of a plurality of surfaces, in particular membranes, in particular of at least 2 membranes, in particular up to 50 membranes, preferably 5 to 20 membranes.

[0062] In another advantageous embodiment, the first porous surface (in particular a polymer membrane) has or carries a first polarity. In addition, the second porous surface has or carries a second polarity. Preferably, the second polarity is particularly opposite to the first polarity. The polarity of the first charge and the second charge is the polarity of the corresponding total charge, which can be provided and / or defined by the sum of the chemical charge and the electrical charge applied to the porous surface by means of a base voltage. In particular, the use of multiple charged porous surfaces can provide several beneficial effects, in particular charged porous surfaces with different properties, in particular different charges (in particular charges of different polarity).

[0063] A particularly advantageous embodiment is based on the use of two charged surfaces, wherein the first charged porous surface and the second charged porous surface are preferably both formed by chemically charged polymer membranes, in particular ion exchange membranes (e.g. in particular cation exchange membranes and anion exchange membranes). Preferably, both polymer membranes (in particular ion exchange membranes) have a conductive coating or a conductive layer at at least one site of each polymer membrane (in particular ion exchange membrane). In addition, each membrane can have a further conductive coating or a conductive layer at at least a second site of the membrane, in particular the second site being opposite the first site. By using two charged surfaces with different polarity, substances that are anionic at a given pH value can be adsorbed to one of the surfaces (in particular the anion exchange membrane) while substances that are cationic can be adsorbed to the other surface (in particular the cation exchange membrane). If the given pH value is chosen such that the given pH value is between the isoelectric points of at least two substances to be separated, in particular for carrying out subsequent methods such as concentration enhancement, detection enhancement, this may be particularly advantageous, in particular if it is desired to separate the two substances or, in particular, for example in the purification of blood or the like, in particular in plasma purification or plasma protein separation, both substances are understood to be impurities.

[0064] In this method using surfaces with different polarities (particularly anion exchange membranes and cation exchange membranes), in carrying out step b) of the method of the present invention, a substance with a second polarity and a specific net total charge is adsorbed onto the first porous surface, a substance with a first polarity and a specific net total charge is adsorbed onto the second porous surface, for example, at a given pH, a substance with a total cationic charge is adsorbed onto the cation exchange membrane and at a given pH, a substance with a total anionic charge is adsorbed onto the anion exchange membrane. In a subsequent step d), a release voltage is preferably applied to the first charged porous surface while a release voltage is not applied to the second charged porous surface. Therefore, only the substance previously adsorbed onto the first charged porous surface is desorbed, while the substance adsorbed onto the second porous surface remains in an adsorbed state. Optionally, in particular after collecting and / or removing the eluted fluid, it may be beneficial to reclaim other substances still adsorbed onto the second surface by subsequently applying a release voltage to the second charged porous surface, thereby desorbing the substance previously adsorbed onto the second charged porous surface. Likewise, the fluid from which such material elutes may also be collected and / or removed.

[0065] In particular, if more than two substances are to be selectively separated, combining several of the previously described embodiments may be beneficial. For example, a method using an anion exchange membrane and a cation exchange membrane can combine Option I and Option II to achieve selective separation. In this regard, a given pH can be selected so that two substances are adsorbed to the same porous surface, while a third substance is adsorbed to another porous surface. Therefore, if the surfaces are eluted separately, the third substance can be effectively separated from the other two substances. This method uses Option I for selective processing. For the separation of the remaining two substances bound to the same porous surface, Option II for achieving selective separation can be used. Therefore, the release voltage can be adjusted stepwise to ensure gradual elution of the two bound substances by selectively desorbing one of the substances at a first release voltage that is insufficient to trigger desorption of the other substance bound to the surface, thereby selectively separating that substance from the other. Alternatively, another cycle of the method can be performed to separate two substances bound to the same surface. In this regard, the given pH can be varied between cycles, particularly to change the net charge of the substances. Preferably, the pH is then selected so that the net charges of the substances have different polarities. By doing this, in the second cycle, these substances can be adsorbed to a different surface in step b) of the second cycle. Thus, option I of selective separation is used again.

[0066] Furthermore, in such methods using differently charged porous surfaces, it can be particularly advantageous to provide a conductive coating or layer as an electrode for applying the release voltage between each porous surface (particularly a membrane). Thus, the release voltage can be applied to only a portion of the surface. In particular, the porous surface (particularly a polymer membrane) can then be used as an insulator between the conductive coating or layer.

[0067] In particular, each portion of the fluid flows over each of the at least two charged porous surfaces, in particular if these surfaces are differently charged porous surfaces, in particular a first charged porous surface having a first polarity and a second charged porous surface having a second polarity opposite to the first polarity. When a plurality of identical charged porous surfaces are used, each portion of the fluid may be flowed over each of the charged porous surfaces or only over some of the charged porous surfaces.

[0068] Preferably, for each elution, fluid is collected and removed, typically before the next elution is performed.

[0069] In another aspect, an advantageous arrangement of surfaces (particularly membranes) is provided. This advantageous arrangement is a corrugated arrangement of surfaces (particularly membranes) and / or a coiled arrangement of these surfaces. This arrangement is particularly useful for chromatography, including but not limited to the methods according to the above invention. In particular, this arrangement can be used as part of any suitable chromatography device.

[0070] In this respect, pleated arrangement is produced by folding the stacking body on flat surface or alternatively surface into multiple pleats (pleat).About winding arrangement, the stacking body on flat surface or surface is wound into a series of stacked layers of winding.Pleated arrangement or winding arrangement are placed in shell.Usually, shell can be selected to accommodate pleated arrangement or winding arrangement so that fluid can flow through and / or pass through surface (preferably film).Shell can provide different flow directions, and particularly shell can provide at least one inlet and at least one outlet for inputting (inlet) and removing (outlet) fluid.Then, pleated arrangement or winding arrangement can be arranged in shell, so that it blocks the free passage from entrance to exit and / or ensures that any flow from entrance to exit will flow through and / or pass through film at least once (preferably repeatedly) and / or flow through and / or pass through film at multiple parts or regions (preferably separated parts or regions) of stacking body and / or film.Pleated arrangement and / or winding arrangement can be arranged in shell so that the main extension of this arrangement is parallel, vertical or tilted to described different flow directions.

[0071] Tubular structure can be used as the shell for these arrangements.This tubular structure can have any form of substrate area, including but not limited to rectangular or circular substrate area.In addition, suitable shell can be the shell of box-like structure form, provides inlet and outlet between the charged porous surface being placed.The charged porous surface between these inlet and outlet can be fed laterally.In this respect, charged porous surface is arranged to block the free passage from inlet to outlet, and makes the fluid of outlet feeding (fed) must flow through at least once and / or preferably pass through charged porous surface (preferably film).Therefore, inlet must be towards the first side of charged porous surface, and outlet is arranged at the second site relative to the first side of charged porous surface, preferably the second site relative to the first side diagonal line.

[0072] However, in other usage scenarios, a circular shell consisting of an inner core and an outer cage is preferred, with a gap between the core and the cage. In this aspect, the gap has a gap distance defined by the shortest distance between the core and the cage. The pleated surface or the wound surface (in particular a membrane) is placed in the gap. Preferably, the inner core and the outer cage are permeable to fluid and / or include inlets / outlets for feeding fluid to the surface and / or removing fluid / eluent. Preferably, the fluid needs to pass radially through the arrangement in the gap to flow from the inlet to the outlet. Typically, only one of the inner core and the outer shell has an inlet and the other has an outlet. In this aspect, the inlet can describe any channel for feeding, in particular a channel permeable to fluid, and the outlet can describe any channel for removal. In particular, the volume within the core can be used for feeding or removal.

[0073] With or without pleating and / or coiling, when surfaces with different polarities are used, preferably the fluid flows over (in particular sequentially over) various types of surfaces with different polarities, and / or they can be arranged so that the fluid flows over (in particular sequentially over) various types of surfaces with different polarities from the inlet to the outlet.

[0074] Preferably, a polymer film is used to provide and / or serve as a surface in a pleated and / or wound arrangement. Most preferably, the pleated and / or wound arrangement is formed by one or more polymer films. In these polymer films, at least some or all are preferably coated with a metal coating or metal layer on at least one side of the film. Typically, the pleated and / or wound arrangement (preferably a stack of films) includes at least two conductive coatings or conductive layers, preferably a metal coating.

[0075] In a particularly advantageous embodiment, the pleated arrangement and / or wound arrangement is composed of at least two polymer films, the at least two polymer films having a metal coating as a conductive coating or conductive layer on one side of the respective films, forming a stack, the at least two polymer films having a non-conductive polymer film between the coated films, respectively. Preferably, the metal coating is arranged so that at least two are placed on the outermost side of the stack, in particular on the outer side of the outermost polymer film in the stack, one on each side, in particular with respect to the flow direction in the housing. In this respect, it is particularly beneficial to pleat and / or wind the film and the conductive coating together. In particular, the polymer film with the metal coating (in particular gold or platinum) can be pleated or wound, in particular pleated or wound together. Preferably, the wound and / or pleated stack consists of only one type of film, which is uncoated, coated and / or has a binding site for attachment, for example, only different in terms of coated / uncoated and / or binding site.

[0076] In another advantageous embodiment, each polymer film of the stack may have a metal coating as a conductive coating or conductive layer on one or both sides of the respective film. Preferably, the film or films themselves are non-conductive and act as an insulator between the metal coatings or metal layers. In particular, the release voltage may be applied only to a selected metal coating and / or metal layer at a time, in particular, the release voltage may be applied to at least one other metal coating and / or to different metal coating pairs, each coating pair possibly sharing a single metal coating.

[0077] Preferably, the stack is formed of "same films", i.e., films of the same type. At least some of them preferably include a conductive coating, or all of them preferably include a conductive coating. Preferably, some of them may include a conductive coating, while others may preferably not include a conductive coating.

[0078] Yet, in another advantageous embodiment, can use " different membranes " in combination, particularly membranes with different charge polarities on charged surfaces, for example cation exchange membrane and anion exchange membrane. In this respect, it may be useful to provide an arrangement in which a voltage (particularly a release voltage) can be applied to a type of membrane to separate it from other types of membranes. In this respect, for any type of membrane, at least one of the membranes of said type particularly has a conductive coating or a conductive layer. Between the conductive coatings or conductive layers of said type, at least one insulator, particularly an insulator formed by any polymer film, is provided. Said arrangement can be used for selectively applying a release voltage to selected membrane or selected type of membrane.

[0079] Preferably, for all embodiments of a pleated and / or rolled arrangement formed by a stack, the stack as a whole is pleated and / or rolled, in particular all films and the conductive coating are pleated and / or rolled together.

[0080] The use of a pleating or rolling aid and / or promoter may also be advantageous, in particular for achieving easier production of the pleats / rolling and for maintaining the shape of the pleats / rolling produced by pleating, or of the stack of layers formed by rolling after such pleating / rolling. Such an aid and / or promoter may consist of one or more thermoplastic layers which, during the production of the pleats / rolling, are heated, in particular to a temperature of 30° C. to 100° C. and / or above the glass transition temperature, and which, after such production, maintains the pleats and / or rolls (in particular their shape), in particular at a temperature below the aforementioned temperatures. Preferably, the pleating and / or rolling aid and / or promoter is also porous and / or permeable to fluids.

[0081] Particularly in the case of housings, by pleating or rolling a surface (particularly a membrane) a higher surface area per unit volume can be provided compared to a non-pleated or non-rolled and therefore flat surface (particularly a membrane).

[0082] The surface area to volume ratio can be influenced by the choice of pleat pattern, particularly with respect to the outer shell. In this aspect, the pleats consist of flat areas located between localized folds, each having a length, preferably an M-shaped pleat. For each pleat, the first length is preferably selected to exceed at least one extension (particularly width, diameter, etc.) of the outer shell (particularly for a circular outer shell with a gap between the outer cage and the inner core). In this aspect, because the core circumference is smaller than the cage circumference, pleats with regular (at least approximately uniform) surface areas will result in a lower, or at least non-optimal, surface area to volume ratio. Therefore, pleats with irregular M-shapes are preferred, as these shapes yield a higher surface area to volume ratio (particularly an optimized surface area to volume ratio). Thus, such irregular M-shapes include flat areas having at least one second length different from the first length. Most preferably, a pleat group consisting of more than one pleat is used. The pleats in any pleat group preferably include pleats having at least two different irregular M-shapes. In this aspect, at least one second length can be less than the gap distance or the width of the channel. Most preferably, the pleats are arranged in groups (particularly regularly) within the shell. In particular, for a circular shell, the pleats and / or the pleats are arranged in groups to form a circle (particularly a complete circle) within the shell.

[0083] A circular housing is also advantageous with respect to the coil arrangement. The coil may be formed by wrapping a surface or a stack of surfaces around an inner core of the housing, whereby the inner core in particular supports the coil formed by the wrapping.

Claims

1. A method for selectively separating amphoteric substances, wherein the substances consist of at least two specific substances, each of which has a specific isoelectric point and thus a specific net total charge at a given pH value; the method comprising the steps of: a) providing a charged porous surface, in particular a chemically and / or electrically charged porous surface, wherein the charged porous surface is electrically conductive and / or comprises an electrically conductive coating or layer; b) contacting a fluid having a given pH value with the charged porous surface, the fluid containing the amphiphilic substance, thereby adsorbing at least a first substance of at least two substances to the charged porous surface, the adsorption being due to an interaction force between the net total charge of the adsorbed substance and the charged porous surface, the interaction force being an attractive force; c) preferably passing at least part of the fluid over and / or through the charged porous surface; d) applying a release voltage to the electrically conductive charged porous surface and / or the electrically conductive coating or layer, the release voltage being selected such that the net interaction force between the charged surface and at least a portion of the adsorbed species changes from an attractive force to a repulsive force, thereby releasing the portion of the adsorbed species; The method is characterized in that the separation is carried out by the following steps: I. in step b), selectively adsorbing at least a portion of the first substance while not adsorbing at least one other substance of the at least two substances, in particular not adsorbing other substances of the at least two substances; and / or II.i) in step b), adsorbing at least part of the first substance and at least part of at least one second substance of the at least two substances, in particular of all substances of the at least two substances, and ii) in step d), selectively releasing the first substance, wherein at least one second substance adsorbed in step b) remains adsorbed, in particular all other substances adsorbed in step b) remain adsorbed.

2. The method according to claim 1 , wherein during step d), a further fluid is brought into contact with the charged porous surface, the further fluid in particular having a given pH value and / or having a pH value which differs from the given pH value by less than 1, in particular less than 0.5, in particular less than 0.2, in particular less than 0.1, and / or being free of any substance, wherein at least part of the further fluid passes over and / or through the porous surface, wherein the fluid which passes over and / or through the porous surface in step d) is collected and contains the substance released in step d).

3. The method according to any one of the preceding claims, characterized in that Throughout steps b) to d), in particular for all fluids, the pH values are kept and / or maintained constant and / or differ by less than 1, in particular less than 0.5, in particular less than 0.2, in particular less than 0.

1.

4. The method according to any one of the preceding claims, characterized in that The substance is a carrier, wherein at least some of the vectors contain molecules, thereby providing at least two groups of vectors with different contained molecules, in particular vectors with different molecules and / or different molecule contents, wherein the carrier has a basic isoelectric point and a basic net total charge at a given pH value, The at least two groups of vectors are defined, in particular, based on the molecules contained in each group of vectors, wherein the basic isoelectric point and the basic net total charge of any specific vector of the vectors containing molecules are shifted due to the molecules contained in the specific vector, resulting in a shifted isoelectric point that is different from the basic isoelectric point, and a shifted net total charge at the given pH value that is different from the basic net total charge at the given pH value.

5. The method according to claim 4, characterized in that The vectors of the first set of vectors contain a first amount of the molecule, and the vectors of the second set of vectors contain a second amount of the molecule.

6. The method according to claim 5, characterized in that The molecule contained in at least some of the vectors is a genome, in particular RNA, DNA, wherein one of the at least two groups of vectors each contains a complete genome, and wherein at least another group of vectors in the at least two groups of vectors each contains a partial genome and / or does not contain a genome.

7. The method according to any one of the preceding claims, characterized in that The given pH value is selected so that the given pH value is between the isoelectric points of the first substance and the second substance, and at the selected given pH value, the polarity of the net total charge of the first substance is opposite to the polarity of the charge of the charged porous surface, and the net total charge of the second substance, in particular all other substances, is zero or has the same polarity as the charge of the charged porous surface, so that only the first substance is adsorbed in step b), in particular no other substances are adsorbed.

8. The method according to any one of the preceding claims, characterized in that The given pH value is selected so that the selected pH value is higher or lower than the isoelectric point of the first substance and the second substance, and the polarity of the net total charge of the first substance and the second substance is the same, which is opposite to the polarity of the charge of the charged porous surface, thereby adsorbing both the first substance and the second substance to the charged porous surface in step b).

9. The method according to claim 8, characterized in that In step d), for the adsorbed first substance and the second substance, the release voltage is different and / or the selection of the release voltage is different, wherein by applying a first release voltage to the conductive surface and / or the conductive coating or the conductive layer, the net interaction force of one of the first substance and the second substance is converted into a repulsive force, while the net interaction force of the other of the first substance and the second substance is maintained as an attractive force, thereby selectively desorbing only the first substance or the second substance, while the other substance remains adsorbed on the charged porous surface.

10. The method according to claim 9, characterized in that After desorbing one of the first substance and the second substance by applying the first release voltage, the applied voltage is adjusted to a second release voltage, wherein by applying the second release voltage, the net interaction force between the first group of carriers and the other group of the second group of carriers is changed from an attractive force to a repulsive force, thereby desorbing the other group of the first group of substances and the second group of substances.

11. The method according to any one of the preceding claims, characterized in that The porous surface is electrically charged by applying a base voltage, wherein the desorption in step d) is initiated by changing the base voltage to a release voltage, in particular lowering it to a release voltage, in particular gradually changing the voltage to, in particular lowering it to, a first release voltage and subsequently a second release voltage.

12. The method according to any one of the preceding claims, characterized in that In step a) at least two charged porous surfaces are provided, wherein the at least two charged porous surfaces comprise at least a first charged porous surface and a second charged porous surface, wherein the first charged porous surface is electrically conductive and / or comprises an electrically conductive coating or layer.

13. The method according to claim 12, characterized in that The second charged porous surface is electrically conductive and / or comprises an electrically conductive coating or layer.

14. The method according to claim 12 or claim 13, characterized in that The at least two charged porous surfaces are arranged as a stack of stacked layers.

15. The method according to claim 14, characterized in that forming the stack such that the first charged porous surface and / or the conductive coating or conductive layer of the first charged porous surface is positioned on a first end of the stack, wherein the counter electrode is arranged on a second end opposite to the first end of the stacked layer, The second charged porous surface, in particular a plurality of second charged porous surfaces, is placed between the first porous surface and the counter electrode, wherein the second charged porous surface is in particular non-conductive and does not comprise a conductive coating or layer, in particular is chemically charged.

16. The method according to claim 15, wherein no further electrodes are placed between the first charged porous surface and the second charged porous surface, in particular between all charged porous surfaces, and / or wherein among the surfaces forming the stacked layers, only the surface placed at the first end or the second end is conductive and / or includes a conductive layer, and / or wherein at least one charged surface, in particular a non-conductive surface, acts as an insulator between the conductive coating or layer and the corresponding counter electrode, and / or An insulator is placed between the conductive coating or layer and the corresponding counter electrode.

17. The method according to any one of the preceding claims, characterized in that The charged porous surface is formed by at least one polymer film having a conductive coating or layer disposed on at least one side thereof, and / or the at least two surfaces each comprise at least one polymer film.

18. The method according to any one of claims 16 or 17, characterized in that The insulation is formed by at least one, in particular a plurality, in particular up to 30, preferably 5 to 15, additional polymer films and / or non-conductive sides of polymer films comprising a conductive coating, in particular the non-conductive sides of the films forming the first surface and the second surface.

19. The method according to any one of the preceding claims, characterized in that The conductive coating or conductive layer is a metal coating or metal layer, preferably gold or platinum, in particular consisting of a single metal, preferably gold or platinum, and / or the conductive coating is formed on a non-conductive support, in particular on a polymer film of the first charged porous surface and / or the second charged porous surface.

20. The method according to any one of claims 12 to 19, characterized in that The first porous surface has or is imparted with a first polarity, and the second porous surface has or is imparted with a second polarity, the second polarity being opposite to the first polarity.

21. The method according to claim 20, characterized in that The first charged porous surface and the second charged porous surface are both formed by chemically charged polymer membranes, in particular ion exchange membranes, both polymer membranes, in particular ion exchange membranes, in particular each polymer membrane, in particular ion exchange membrane, has a conductive coating or conductive layer on at least one side.

22. The method according to claim 20 or claim 21, characterized in that In step b), a substance having a specific net total charge of the second polarity is adsorbed onto the first porous surface, and a substance having a specific net total charge of the first polarity is adsorbed onto the second porous surface.

23. The method according to claim 23, characterized in that In step d), a release voltage is applied to the first charged porous surface, while no release voltage is applied to the second charged porous surface, thereby desorbing the substance previously adsorbed on the first charged porous surface, while the substance adsorbed on the second porous surface remains adsorbed, and optionally, a release voltage is subsequently applied to the second charged porous surface, thereby desorbing the substance previously adsorbed on the second charged porous surface.

24. The method according to any one of claims 18 to 23, characterized in that The polymer film and the conductive coating or layer and / or the stack are corrugated.

25. A method according to any of the preceding claims, wherein the release voltage, in particular the first release voltage and / or the second release voltage, and / or the base voltage, in particular the applied voltage, in particular the base voltage and / or the release voltage, is in the range of 0.1 V to 50 V, in particular in the range of 0.1 V to 3 V.