A single-step process for preparing pure immunoglobulin from plasma.
A single-step electroseparation process with pH adjustment and filtration effectively addresses the limitations of conventional methods, producing high-purity immunoglobulins with preserved functionality and efficient yield, bypassing ethanol and chromatography.
Patent Information
- Authority / Receiving Office
- JP · JP
- Patent Type
- Applications
- Current Assignee / Owner
- Filing Date
- 2024-03-13
- Publication Date
- 2026-03-16
AI Technical Summary
Conventional plasma fractionation processes for immunoglobulin purification face challenges such as high cost, low yield, unsuitability for automation, exposure to harsh conditions, and difficulty in preserving biological functionality, failing to meet the demand for high-purity and high-recovery immunoglobulins.
A single-step electroseparation process using a specified pore size separation membrane and electric field to selectively move charged immunoglobulin molecules across the membrane, combined with pH adjustment and filtration to achieve at least 95% purity.
The process achieves high-purity immunoglobulins with preserved biological functionality, eliminating the need for ethanol and chromatography, while maintaining a similar IgG subclass distribution to plasma, and achieving yields of at least 80%.
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Abstract
Description
Technical Field
[0001] This technology relates to the separation of at least 95% pure immunoglobulin from plasma using a single electrical separation step. Cross-reference to related applications
[0002] This application claims priority based on Australian Provisional Patent Application No. 2023900670 filed on March 13, 2023, the content of which is incorporated herein by reference in its entirety.
Background Art
[0003] Plasma contains many proteins including coagulating substances (mainly fibrinogen), albumin and globulin to maintain a colloid osmotic pressure of about 25 mmHg, electrolytes, immunoglobulins, and various other enzymes, hormones, and vitamins.
[0004] Plasma or plasma components have many clinical uses. Plasma can help fight infections and treat inflammation and autoimmune diseases, and can also promote blood clotting to prevent shock and assist in postoperative recovery. For example, whole plasma is indicated for the treatment of hemorrhagic shock. Coagulation factors can be used in the treatment of hemophilia and von Willebrand disease. Albumin infusions can be used in the treatment of burns, cirrhosis, and the management of hepatorenal syndrome. In addition, human plasma-derived α-1 antitrypsin is used to treat α-1 antitrypsin deficiency.
[0005] Immunoglobulins derived from pooled human plasma are useful in the treatment of a very large number of autoimmune diseases, inflammatory diseases, hematological diseases, and viral diseases including COVID-19. Immunoglobulins are typically administered as intravenous immunoglobulin (IVIG).
[0006] A common method for preparing immunoglobulins from plasma uses the Cohn / Oncley fractionation procedure, which separates plasma proteins by differential precipitation using cold ethanol. Plasma fractionation involves a series of processing steps at specified ethanol concentrations, related to shifts in pH, temperature, and ionic strength, resulting in the selective precipitation of immunoglobulin G (IgG).
[0007] Further purification is achieved using at least two chromatographic steps, most schemes involving size exclusion, anion / cation exchange, or affinity chromatography. Chromatography offers superior selectivity in protein purification, with recovery typically exceeding 70% when proteins are isolated from the plasma protein fraction. However, the chromatographic approach has drawbacks, including the high cost of resins and columns, and the need for purification between uses.
[0008] Conventional plasma fractionation processes have many drawbacks, including their unsuitability for automation, the use of ethanol which impacts plant design, the exposure of immunoglobulins to harsh conditions that negatively affect their function, the often difficult solubilization of precipitated proteins, and the low yield of immunoglobulins, typically around 50%. [Overview of the project] [Problems that the invention aims to solve]
[0009] The shortcomings of conventional purification schemes result in an inability to meet the high demand for plasma-derived therapeutic molecules such as IVIG. Therefore, the plasma fractionation industry is working together to find improved and novel methods for the rapid fractionation of plasma, as well as for the purification of IgG with high recovery and purity, while preserving the biological functionality of the target protein. [Means for solving the problem]
[0010] In a first embodiment, a process for producing immunoglobulin with a purity of at least 95% from plasma; the following: a. An electroseparation system, the following: A separation membrane disposed between a cathode and an anode, the separation membrane having a specified pore size; A first limiting membrane, positioned between the cathode and the separation membrane, and defining the first fluid path of the first fluid flow (preventing molecules in the first fluid flow from coming into contact with the cathode); A second limiting membrane, positioned between the anode and the separator membrane, and defining the second fluid flow path of the second fluid flow (preventing molecules in the second fluid flow from coming into contact with the anode); Plasma is applied to the first fluid flow of the electroseparation system, including; and, b. Apply an electric field to the fluid flow to selectively move positively charged immunoglobulin molecules toward the cathode, across the separation membrane in the second fluid flow; or a. Apply plasma to the second fluid flow of the electroseparation system; and, b. Apply an electric field to the fluid flow to selectively move negatively charged molecules in the plasma toward the anode, across the separation membrane in the first fluid flow. The process, including the above, is provided.
[0011] The process described in the claim may further include the step of recovering immunoglobulin with a purity of at least 95% from the first or second stream.
[0012] In one embodiment, the process described in the claim further includes preparing the plasma by lowering the pH of the stored plasma to about 4.5 to 7.5.
[0013] The pH can be lowered by adding a buffer, such as a buffer containing MES, Bis-tris, and glycine.
[0014] The process may further include removing precipitates from the plasma, for example, by filtration or centrifugation, preferably by filtration.
[0015] In one embodiment, the first fluid and / or second fluid flow includes a buffer, such as a buffer containing MES, Bis-tris, and glycine.
[0016] In one embodiment, the pore size of the separation membrane is approximately 200 kDa to approximately 1500 kDa, preferably approximately 1000 kDa.
[0017] In one embodiment, immunoglobulin with a purity of at least 95% is free of bacteria, viruses, or prions when collected from a first or second stream.
[0018] The process may further involve the enrichment of immunoglobulin with a purity of at least 95%.
[0019] In a second embodiment, a preparation is provided comprising immunoglobulin with a purity of at least 95%, produced by the process described in the first embodiment. In one embodiment, the immunoglobulin produced by the process has substantially the same IgG subclass distribution as the original plasma.
[0020] definition Throughout this specification, unless the context clearly requires a different meaning, variations of the word "include" or "include" are understood to mean the inclusion of the elements, integers, or steps, or groups of elements, integers, or steps, but not the exclusion of other elements, integers, or steps, or groups of elements, integers, or steps.
[0021] Throughout this specification, the term "consisting of" means consisting solely of that.
[0022] The description of the documents, acts, materials, devices, articles, or the like contained in this application is solely for the purpose of providing the background of the technology of the present invention. It should not be construed that any or all of these matters formed part of the basis of the prior art because they existed before the priority date of each claim of this application, nor should it be construed that they were common general knowledge in the field related to the technology of the present invention.
[0023] Unless the context requires otherwise, or unless specifically stated to the contrary, the integers, steps, or elements of the technology described herein as a single integer, step, or element clearly encompass both the singular and plural forms of the recited integer, step, or element.
[0024] In the context of this specification, the words "a" and "an" refer to the fact that the grammatical object of the article is one or more (that is, at least one). By way of example, a reference to an "element" is intended to mean one element or more than one element.
[0025] In the context of this specification, the word "about" when referring to a number or value should not be regarded as an absolute number or value, but rather encompasses the margin of variation above and below that number or value that would be understood by a person skilled in the art in accordance with the relevant art, within the range of typical margins of error or device limitations. In other words, the use of the word "about" is understood to refer to a range or approximation such that a person skilled in the art would consider the value described to be equivalent from the perspective of achieving the same function or result.
[0026] It will be understood by those skilled in the art that variations and modifications other than those specifically described are possible for the technology described herein. It is to be understood that the technology encompasses all such variations and modifications. For the sake of clarity, it is hereby stated that the technology also encompasses individually or collectively all of the steps, features, and compounds mentioned or indicated herein, and also any and all combinations of any two or more of the above steps, features, and compounds.
[0027] To enable a clearer understanding of the technology of the present invention, preferred embodiments will be described with reference to the following drawings and examples. [Modes for carrying out the invention]
[0028] Description of the Embodiment The process described herein uses plasma, typically stored human plasma, as the source of immunoglobulins and involves a single electrolysis step to prepare immunoglobulins of at least 95% purity from the stored plasma.
[0029] plasma storage Plasma may be provided or prepared.
[0030] In some embodiments, the preparation involves adjusting the pH of the plasma before electrolysis.
[0031] In one embodiment, the pH of the plasma may be adjusted by any means known in the art, such as diafiltration.
[0032] In a preferred embodiment, plasma is prepared by diluting the stored plasma with a buffer so that the pH of the diluted plasma is in the range of approximately pH 4.5 to approximately pH 7.5. For example, the pH of the diluted plasma is approximately 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, or 7.5.
[0033] Naturally, as the pH is lowered, plasma proteins with a pI close to the pH of the diluted plasma will precipitate. In this regard, the process may optionally include a precipitate removal step, for example, continuous stirring followed by filtration, centrifugation, or filtration, preferably filtration, to remove the precipitated proteins from the diluted plasma.
[0034] Any physiological buffer that can be formulated to lower the pH of diluted plasma can be used in the current process. Selecting and formulating the appropriate buffer is within the realm of the knowledge of those skilled in the art.
[0035] Suitable physiological buffers include MES, Bis-tris, ADA, ACES, PIPES, MOPSO, coramine hydrochloride, Bis-tris propane, BES, MOPS, BES, TES, HEPES, DIPSO, MOBS, TAPSO, acetamidoglycine, Tris, HEPPSO, POPSO, TEA, HEPPS, HEPPSO, tricine, glycine, glycinamide, glycylglycine, bicine, HEPBES, TAPS, and combinations thereof.
[0036] In some embodiments, the buffer does not contain a salt (e.g., NaCl).
[0037] The pH of the buffer may differ from the final pH of the diluted plasma.
[0038] To prepare plasma, the buffer is mixed with the stored plasma to form diluted plasma.
[0039] In some embodiments, the buffer:retained plasma ratio is about 0.5:1 to about 4:1, for example, about 0.5:1 (0:5:1), 1:1, 1.5:1, 2:1, 2.5:1, 3:1, 3.5:1, or about 4:1. Preferably, the ratio is about 2.5:1, 2:1, or 2.5:1, for example, about 2:1.
[0040] In one embodiment, the buffer contains a combination of MES hydrate, Bis-tris, and glycine. The pH of this buffer is approximately 3.5 to 8.5. For example, the pH of the buffer could be 3.5, 3.6, 3.7, 3.8, 3.9, 4.0, 4.1, 4.2, 4.3, 4.4, 4.5, 4.6, 4.7, 4.8, 4.9, 5.0, 5.1, 5.2, 5.3, 5.4, 5.5, 5.6, 5.7, 5.8, 5.9, 6.0, 6.1, 6.2, 6.3, 6.4, 6.5, 6.6, 6.7, 6.8, 6.9, 7.0, 7.1, 7.2, 7.3, 7.4, 7.5, 7.6, 7.7, 7.8, 7.9, 8.0, 8.1, 8.2, 8.3, 8.4, or approximately 8.5.
[0041] In one embodiment, the buffer contains 15.86 kg of MES hydrate, 1.66 kg of Bis-tris, and 11.56 kg of glycine in a total volume of 668 L. In this embodiment, the buffer is mixed with approximately 320 L of thawed stored plasma to form diluted plasma. Those skilled in the art can change the buffer volume to accommodate various amounts of stored plasma.
[0042] Precipitates in diluted plasma are optionally removed. Removal of precipitated proteins may include allowing the precipitate to settle in a waste collection tank, such as a bottom drain tank. Removal of precipitate may additionally or selectively include centrifugation and / or filtration. Filtration is generally preferred because it is suitable for use in large volumes.
[0043] In some embodiments, plasma is subjected to a buffer exchange step before electrolysis. Buffer exchange can be achieved by any means known in the art, such as filtration (e.g., diafiltration), dialysis, or chromatography. Filtration (e.g., diafiltration) is generally preferred because it can be modified for use in large volumes.
[0044] In one embodiment, the buffer is replaced with another buffer using a filtration step, for example, a tangential flow filtration step. In this embodiment, the buffer is replaced with analogues of different physiological buffers. The selection of the buffer in such situations is within the realm of the knowledge of those skilled in the art.
[0045] In one embodiment, the MES hydrate, Bis-tris, and glycine buffer used to dilute plasma is replaced with a slightly different composition of MES hydrate, Bis-tris, and glycine buffer. In one embodiment, the replacement buffer contains 12.29 kg of MES hydrate, 2.85 kg of Bis-tris, and 16.69 kg of glycine in a total volume of 1058 L. In this embodiment, the buffer is mixed with approximately 988 L of diluted plasma. Those skilled in the art can adjust the buffer volume to accommodate various starting amounts of plasma.
[0046] In some embodiments, the exchange buffer:plasma ratio is 0.5:1 to about 2:1, for example, about 0.5:1 (0:5:1), 0.75:1, 1:1, 1.25:1, 1.5:1, 1.75:1, or about 2:1. Preferably, the ratio is about 0.75:1, 1:1, 1.25:1, for example, about 1:1.
[0047] In some embodiments where it is advantageous to minimize the contribution of the buffer species to the ionic strength of the buffer, this can be achieved by using an amphoteric buffer species (e.g., so-called Good's buffers such as MES or Tris) rather than salts such as sodium phosphate or ammonium chloride.
[0048] Single electrical isolation step Immunoglobulins are separated from plasma using a single electrolysis step.
[0049] Electrical separation is as follows: A separation membrane disposed between a cathode and an anode, the separation membrane having a specified pore size; A first limiting membrane, positioned between the cathode and the separation membrane, and defining the first fluid path of the first fluid flow (preventing molecules in the first fluid flow from coming into contact with the cathode); A second limiting membrane, positioned between the anode and the separator membrane, and defining the second fluid flow path of the second fluid flow (preventing molecules in the second fluid flow from coming into contact with the anode); This is achieved using the aforementioned electro-separation system, including the following:
[0050] The electroseparation system may include a first fluid channel and inlets and outlets that communicate with the fluid, as well as a second fluid channel and inlets and outlets that communicate with the fluid.
[0051] During operation, the first fluid flow moves along the first channel, and the second fluid flow moves along the second channel. An electric field is applied to the fluid flow in the channels, causing negatively charged molecules in the fluid flow to move toward the anode and positively charged molecules in the fluid flow to move toward the cathode. In general, molecules with a size (or hydrodynamic radius) smaller than the pore size of the separation membrane will cross the separation membrane from one fluid flow to the other. The shape of the molecule also affects the passage of charged molecules across the membrane. For example, a rod-shaped protein may have a hydrodynamic radius that prevents the protein from passing through the membrane when moving in solution.
[0052] Typically, one of the fluid flows (e.g., the first or second fluid flow) is a supply flow into which plasma is applied, and the other flow is a product flow that receives molecules that have passed through the separation membrane due to the application of an electric field.
[0053] In some embodiments, the flow path is an independent linear path, a series of linear paths connected by bends. Alternatively, the flow path may be zigzag or helical. In some embodiments, the flow path is as long as possible to maximize the time the fluid flow is exposed to the electric field, thereby reducing or eliminating the need to recirculate the fluid flow to achieve effective separation of immunoglobulins.
[0054] Typically, a limiting membrane is a molecular barrier with a defined pore size smaller than that of a separation membrane. The separation membrane functions to prevent proteins and other macromolecules in a fluid flow from coming into contact with the electrodes (i.e., the anode and cathode).
[0055] A suitable electroseparation system is the Aegros HaemaFrac® system, which uses the mobility of charged particles in an electric field to transport proteins across a membrane in a collection chamber or fluid flow.
[0056] In one embodiment, the fluid of the first and / or second fluid flow is a buffer, for example, the physiological buffer described above.
[0057] In one embodiment, the buffer contains a combination of MES hydrate, Bis-tris, and glycine. For example, the buffer contains 5.8 kg of MES hydrate, 1.3 kg of Bis-tris, and 9.3 kg of glycine in a total volume of 500 L. Those skilled in the art can change the buffer volume to accommodate various starting amounts of plasma.
[0058] Plasma is applied to a first fluid flow. For example, plasma can be pumped into the first fluid flow through an inlet. When plasma enters the first fluid flow, an electric field is applied to that fluid flow.
[0059] If the first fluid flow exits the first fluid channel, for example, through an outlet, it may be recirculated to the inlet of the first fluid channel or it may be recovered.
[0060] In some embodiments, the first or second fluid flow may be cooled.
[0061] If the second fluid flow exits the second fluid channel, for example, through an outlet, it can be recirculated to the inlet of the second fluid channel, or it may be recovered.
[0062] The typical flow rate used in the process for each fluid flow is independently selected from approximately 5 to 100 ml / min. For example, the flow rate may be any one of 5, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 ml / min. In one embodiment, the flow rate is 5 to 50 ml / min. In a further embodiment, the flow rate is approximately 20 ml / min.
[0063] In some embodiments, a voltage of approximately 50V is applied.
[0064] In one embodiment, the electric field causes positively charged immunoglobulin molecules to traverse the separation membrane toward the cathode into a second fluid flow.
[0065] In an alternative embodiment, the electric field causes negatively charged molecules to traverse the separation membrane toward the anode into a second fluid flow.
[0066] The separation membrane contains pores of a specified size sufficient to allow immunoglobulins to pass across the membrane. Plasma contains various immunoglobulin isotypes, although immunoglobulin G (IgG) accounts for 70% to 75% of the total immunoglobulins in the plasma. IgG has a molecular weight of approximately 150 kDa, but the pore size of the membrane is larger than this to allow immunoglobulins to traverse the membrane.
[0067] In one embodiment, the pore size of the membrane is at least 200 kDa to about 1500 kDa, for example, 200, 250, 300, 250, 400, 450, 500, 550, 600, 650, 700, 750, 800, 850, 900, 950, 1000, 1050, 1100, 1150, 1200, 1250, 1300, 1350, 1400, 1450, or about 1500 kDa.
[0068] In one embodiment, the pore size of the membrane is at least about 800, 850, 900, 950, 1000, 1050, 1100, 1150, or 1200 kDa, preferably 900, 950, 1000, 1050, or 1100 kDa, and more preferably about 1000 kDa.
[0069] Pore size is either the average pore size or the median pore size.
[0070] The separation membrane may be a polysulfone, polypropylene, cellulose acetate, polylactic acid, or polyacrylamide-based membrane.
[0071] In some embodiments, the membrane is a hydrogel membrane, for example, a polyacrylamide-based hydrogel. In other embodiments, a polyethylene glycol or chitosan-based hydrogel membrane may be used.
[0072] In some embodiments, the separation membrane prevents mixing of fluid flows but allows charged molecules to pass from one fluid flow to the other.
[0073] When an electric field is applied to a fluid flow, all positively charged proteins are expected to traverse the separation membrane toward the cathode into the second fluid flow if they are smaller than the pore size of the membrane. As a result, at least 95% of the proteins entering the second fluid flow become immunoglobulins, and therefore, the process is a process for producing immunoglobulins with at least 95% purity from stored plasma.
[0074] Alternatively, when an electric field is applied to the fluid flow, all negatively charged proteins are expected to traverse the separation membrane toward the anode into the second fluid flow if their size is smaller than the membrane's pore size, while positively charged immunoglobulins remain in the first fluid flow. As a result, at least 95% of the proteins remaining in the first fluid flow will be immunoglobulins from the stored plasma.
[0075] In some embodiments, the immunoglobulins produced by the process have a purity of at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%. In this context, purity % refers to the total amount of immunoglobulins (of all IgG subtypes) expressed as a percentage of the total amount of all proteins present in the second fluid stream.
[0076] Immunoglobulin purity can be measured by any means known in the art, such as natural or modified polyacrylamide gel electrophoresis (e.g., SDS-PAGE), analytical HPLC, or mass spectrometry.
[0077] In some embodiments, the yield of immunoglobulins produced by the process is at least 80% and at most 95% of the total immunoglobulins in the plasma or stored plasma. For example, the yield may be at least about 80%, e.g., at least 80%, 82%, 84%, 86%, 88%, 90%, 92%, 94%, 95%, 96%, 97%, 98%, or at least 99%. In this context, the % yield refers to the amount of immunoglobulins (of all IgG subtypes) in a second fluid stream, expressed as a percentage of the total amount of immunoglobulins present in the plasma or stored plasma.
[0078] Since the pore size of the separation membrane is substantially smaller than that of bacteria and viruses, the electrolysis step is also a viral and bacterial filtration / removal step.
[0079] Immunoglobulin with a purity of at least 95% in the second flow can be collected, for example, by recovering the second fluid flow in a container.
[0080] Processes involving ethanol fractionation are known to potentially lead to the selective loss of several IgG subtypes, specifically IgG3. This is problematic because the IgG profile is key to the plasma's ability to neutralize viral infections such as SARS-CoV-2. For example, Kober et al. (2022) PLOS ONE 17(1): e0262162 found that convalescent plasma donation with hyperimmune immunoglobulin or high IgG3 concentrations may be a highly effective treatment. Generally, viral infections lead to the initial appearance of IgG3 during the course of the infection, followed by the production of IgG1 and IgG3. IgG3 is particularly effective in inducing effector function.
[0081] It is also known that individuals with low IgM or IgG3 levels are at higher risk of developing post-acute coronavirus disease 2019 (COVID-19) syndrome (PACS) or long-term COVID (see, for example, Cervia et al, Nature Communications (2022) 13:446).
[0082] In this respect, immunoglobulin preparations manufactured by processes involving ethanol precipitation often show a drastic reduction in IgG3 (see, for example, Table 1). Caprylate precipitation does not remove IgG and leaves it in solution, thus providing a distribution of IgG subclasses that reflects the distribution in plasma. However, bioprocessing is more complex and still requires gentler purification methods to ensure the preservation of antibody activity. Table 1: IgG subclass profiles of commercially available immunoglobulin preparations [Table 1]
[0083] Immunoglobulin preparations prepared by the processes described herein (i.e., without using cold ethanol precipitation or chromatographic purification) have substantially the same IgG subclass distribution as those of stored plasma, plasma, or normal plasma (IgG1 55%, IgG2 36%, IgG3 5%, IgG4 4% - Miles J & Riches P. Ann Clin Biochem 1994; 31: 245-248). It is understood that there may be variations in the IgG subclass profile between batches of stored plasma. For example, if some plasma donors have a viral infection or have recently had a viral infection, this may cause an increase in IgG3 levels, for example, IgG3 levels in stored plasma, plasma, or immunoglobulin preparations may be approximately 4%, 5%, 6%, 7%, 8%, 9%, or approximately 10%.
[0084] The process does not involve chromatography, ethanol, or caprylate precipitation. The process described herein is used to prepare pure immunoglobulin preparations from plasma. (When preparing plasma or in the separation step) the process does not use ethanol precipitation (e.g., cold ethanol precipitation), chromatographic purification, the use of harsh chemical precipitants such as caprylates, or any combination of these techniques.
[0085] In one embodiment, the process does not involve ethanol precipitation, for example, cold ethanol precipitation.
[0086] In one embodiment, the process does not include any steps involving caprylate precipitation, or caprylate or its salts, derivatives, or analogs.
[0087] In one embodiment, the process does not involve chromatography, such as anion exchange chromatography, cation exchange chromatography, size exclusion chromatography, or affinity chromatography (e.g., using proteins A, G, A / G, and / or L).
[0088] Further processing In some embodiments, the collected immunoglobulin solution is further processed. For example, it may be concentrated or subjected to buffer exchange. Any method for concentrating the collected immunoglobulin solution may be, for example, ultrafiltration.
[0089] In some embodiments, it may be desirable to additionally or selectively perform buffer exchange, for example, to replace the buffer of the collected immunoglobulin solution with a buffer more suitable for use in therapeutics or for long-term storage of the immunoglobulin solution.
[0090] In one embodiment, the buffer of the collected immunoglobulin solution is replaced with glycine buffer. The glycine buffer may contain 3.86 kg of glycine and 0.18 kg of 5 M HCl in a total volume of 206 L. Those skilled in the art can change the buffer volume.
[0091] Those skilled in the art will understand that numerous modifications and / or alterations can be made to the present invention, as shown in the specific embodiments, without departing from the broadly described spirit or scope of the invention. Therefore, these embodiments should be considered illustrative in all respects and not limiting.
Claims
1. A process for producing immunoglobulins with a purity of at least 95% from plasma; the following: a. An electroseparation system, the following: A separation membrane disposed between a cathode and an anode, the separation membrane having a specified pore size; A first limiting membrane, positioned between the cathode and the separation membrane, and defining the first fluid path of the first fluid flow (preventing molecules in the first fluid flow from coming into contact with the cathode); A second limiting membrane, positioned between the anode and the separator membrane, and defining the second fluid flow path of the second fluid flow (preventing molecules in the second fluid flow from coming into contact with the anode); Plasma is applied to the first fluid flow of the electroseparation system, including; and, b. Apply an electric field to the fluid flow to selectively move positively charged immunoglobulin molecules toward the cathode, across the separation membrane in the second fluid flow; or a. Apply plasma to the second fluid flow of the electroseparation system; and, b. Apply an electric field to the fluid flow to selectively move negatively charged molecules in the plasma toward the anode, across the separation membrane in the first fluid flow. The process including the process described above.
2. The process according to claim 1, further comprising the step of recovering immunoglobulin with a purity of at least 95% from a first or second stream.
3. The process according to claim 1 or 2, further comprising preparing plasma by lowering the pH of the stored plasma to approximately 4.5 to 7.
5.
4. The process according to claim 3, wherein the pH is lowered by the addition of a buffer.
5. The process according to claim 4, wherein the buffer comprises MES, Bis-tris, and glycine.
6. The process according to claim 5, further comprising the removal of precipitates from plasma.
7. The process according to claim 6, wherein the removal of the precipitate includes filtration or centrifugation, preferably filtration.
8. The process according to any one of claims 1 to 7, wherein the first and / or second fluid flow includes a buffer.
9. The process according to claim 8, wherein the buffer comprises MES, Bis-tris, and glycine.
10. The process according to any one of claims 1 to 9, wherein the defined pore size of the separation membrane is about 200 kDa to about 1500 kDa, preferably 1000 kDa.
11. The process according to any one of claims 1 to 10, wherein the immunoglobulin, when collected from the first or second stream, is free of bacteria, viruses, or prions.
12. The process according to any one of claims 1 to 11, further comprising the concentration of the immunoglobulin with a purity of at least 95%.
13. The process according to any one of claims 1 to 12, wherein the process does not include one or more of cold ethanol precipitation, caprylate precipitation, or chromatography.
14. A preparation comprising immunoglobulin with a purity of at least 95%, manufactured by the process described in any one of claims 1 to 13.
15. The formulation according to claim 14, wherein the distribution of IgG subclasses is substantially the same as the distribution of IgG subclasses in plasma.