Deep filtration process for removing pigment in protein purification and application of deep filtration process

Abstract

The invention provides a deep filtration process for removing pigment in protein purification and application thereof, and the process comprises the following steps: respectively introducing a deep filtration membrane in the filtration step of protein purification, and simultaneously realizing filtration and pigment removal; the filtering step comprises any one or a combination of at least two of a clarification filtering step, an intermediate product deep filtering step or a virus removal filtering pre-filtering step; the deep filter membrane is selected from an Emphaze deep filter membrane and / or an X0SP deep filter membrane. According to the invention, a pigment removal process flow is introduced into a protein purification process flow, and a new solution and strategy are provided for pigments which cannot be controlled and removed by upstream and chromatography.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and in particular relates to a deep filtration process for removing pigments in protein purification and an application thereof. Background Art

[0002] The appearance of APIs for injection is a test item required by the International Council for Harmonization of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) Q6B release specification, which usually includes color, transparency and turbidity. Abnormal color means that there may be contamination or the properties of the product may have changed. Therefore, the control and removal of pigments is an important part of the production process of APIs for injection. Mastering the control and removal methods of pigments in advance can improve the safety and effectiveness of drugs.

[0003] Pigments include macromolecules, small molecules, positively charged, negatively charged, hydrophobic and hydrophilic pigments, etc. They are generally introduced from the upstream culture medium and bind to the antibody protein through covalent or non-covalent bonds, showing the color of the molecule itself or causing the color produced by protein denaturation. Among them, vitamin B and iron ions have the greatest impact on color. Since they form color substances with covalent bonds with proteins, they are generally controlled at the upstream stage by optimizing cell lines or culture conditions. For solutions where the nature of the pigment is unclear, different chromatography methods are generally tried to remove the color.

[0004] CN201610749377.6 discloses a pretreatment method for improving the clarity of Panax notoginseng total saponins raw material for injection, comprising the following steps: removing impurities in Panax notoginseng total saponins by water precipitation, obtaining a Panax notoginseng extract after water precipitation; filtering the supernatant after separation and precipitation, mainly to remove impurities that are not completely precipitated; then filtering the impurities through a centrifuge, passing the extract through a quartz sand column, to remove small molecule impurities such as pigments; passing the extract from which small molecule impurities such as pigments are removed through an activated carbon column, to remove large molecule impurities such as pigments; the pretreated liquid is continued to be produced according to the process for producing Panax notoginseng total saponins injection, the clarity of the Panax notoginseng total saponins for injection obtained will be greatly improved, and the quality requirements of the raw material for Panax notoginseng total saponins for injection can be met.

[0005] CN202310368838.5 discloses a method for removing pigments during the purification of recombinant human serum albumin; comprising: centrifuging the fermentation broth and filtering, concentrating, and adjusting the pH; then sequentially passing through an anion exchange chromatography column and a reversed-phase polymer filler chromatography column for pigment adsorption; adding L-cysteine, sodium octanoate, and EDTA to the chromatographic product, and then heating and decolorizing. By combining anion exchange chromatography and reversed-phase polymer filler chromatography, negatively charged and weakly polar pigment impurities in the fermentation broth are removed, and by adding L-cysteine, sodium octanoate, and EDTA, heating and decolorizing are performed to remove the pigment bound to the inside of human serum albumin.

[0006] However, due to the complexity of the pigment's own properties, conventional chromatography in-situ cleaning may not be thorough. As the number of times the chromatography filler is used increases, the deposition of pigments may not only lead to changes in filler color, but also blockage and collapse of the filler, which may even affect the load and quality, thereby affecting the performance and life of the filler and increasing production costs. Therefore, developing an effective and robust method to remove pigments from raw materials has important application value in the development and production of injectable drugs. Summary of the invention

[0007] In view of the shortcomings of the prior art, the purpose of the present invention is to provide a deep filtration process for removing pigments in protein purification and its application. The present invention provides an effective and robust method for removing pigments, creatively introduces a combined deep filter membrane to effectively remove pigments, and the filter membrane used in the present invention is a disposable deep filter membrane. Compared with the conventional color removal scheme (the conventional scheme is to use fillers for chromatography, the fillers are expensive, and the life cycle of the fillers needs to be considered), the method of the present invention is lower in cost, and there is no need to worry about the impact of pigments on the life cycle of the fillers. It can also effectively remove pigments and has strong robustness.

[0008] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0009] In a first aspect, the present invention provides a deep filtration process for removing pigments in protein purification, the pigment removal process comprising: introducing a deep filtration membrane into the filtration step of protein purification to achieve filtration and pigment removal simultaneously;

[0010] The filtration step includes: any one or a combination of at least two of a clarification filtration step, an intermediate product deep filtration step, or a pre-filtration step for virus removal filtration;

[0011] The deep filtration membrane is selected from: Emphaze deep filtration membrane and / or X0SP deep filtration membrane.

[0012] The present invention provides a kind of effective and robust method for the removal of pigment, the generation of pigment is more difficult to find specific root cause, when the existing technology of upstream and downstream can not effectively control or downstream chromatography causes life-span decline and increases production cost because of the introduction of pigment, the present invention creatively introduces the deep filter membrane of combination to effectively remove pigment. Mainly introduce deep filter membrane respectively in the pre-filtration step of clarifying filtration step, intermediate product deep filtration and virus removal filtration, for example XOSP or Emphaze. Deep filter membrane is set respectively at three positions, increases the removal efficiency and robustness of pigment. Research shows that the film adopted by the present invention is a disposable deep filter membrane, not only need not worry about the influence of pigment on the life cycle, but also can effectively remove pigment and have stronger robustness.

[0013] In the present invention, when the deep filtration membrane is introduced in the clarification filtration step and the intermediate product deep filtration step, the deep filtration membrane is directly used to replace the original conventional filter membrane, and the deep filtration membrane achieves the purpose of filtration and pigment removal at the same time.

[0014] In the present invention, when a deep filter membrane is introduced in the pre-filtration step of virus removal filtration, the deep filter membrane is used to replace the original conventional pre-filtration membrane, and the deep filter membrane achieves the purpose of pre-filtration and pigment removal at the same time.

[0015] In the present invention, the function of removing pigments is added to three steps respectively to improve the robustness of pigment removal; mainly by introducing it into three steps of using the membrane package in the downstream process, namely the clarification filtration step, the intermediate product deep filtration step and the pre-filtration step of virus removal filtration. The three steps support and cooperate with each other. Even if one of the membranes fails to remove the color, the other two membranes can still take on the role of removing the color.

[0016] In the present invention, various types of deep filter membranes can be introduced in the three steps to increase the robustness of pigment removal; the same type of membrane package can be used in the three steps, or two or three types of membrane packages can be used in the three steps. Mainly, by introducing a variety of effective membranes for removing colors that have been screened out, they are placed in different positions, for example, the Emphaze membrane package can be placed in the clarification filtration step, and the XOSP membrane can be used in the intermediate product deep filtration and virus removal prefiltration. In this way, different membranes remove different types of colors, which can also improve robustness.

[0017] In the present invention, a deep filter membrane series combination strategy is adopted in the three steps to increase the robustness of pigment removal; it can be a simple series connection of the same membrane package in each of the three steps or in a certain step, or multiple membrane packages can be connected in series.

[0018] In the present invention, the pigment removal process can reduce costs, remove pigments efficiently, and is simpler and faster than conventional chromatography.

[0019] Preferably, at least one deep filtration membrane is provided in any filtration step.

[0020] Preferably, when the same type of deep filtration membrane is arranged in any filtration step, the same type of deep filtration membrane is arranged in parallel or in series.

[0021] Preferably, when different types of deep filtration membranes are arranged in any one filtration step, the different types of deep filtration membranes are arranged in series.

[0022] In the present invention, when the color pollution comes from different components, different types of membrane packages remove different colors, and all different types of membrane packages are combined to become the first choice. However, when the color is only contributed by various sources, it is better to choose one membrane package for production.

[0023] In the present invention, when the loading capacity of a single membrane package is not high, parallel connection is adopted, and when the loading capacity of a single membrane package is high, series connection is adopted.

[0024] Preferably, the same type of deep filtration membrane is used in the clarification filtration step, the intermediate product deep filtration step, and the pre-filtration step for virus removal filtration.

[0025] Preferably, any two of the clarification filtration step, the intermediate product deep filtration step or the pre-filtration step for virus removal filtration are provided with the same type of deep filter membrane, and the other step is provided with another type of deep filter membrane.

[0026] In a specific embodiment of the present invention, the deep filtration membranes used in the clarification filtration step and the intermediate product deep filtration step are the same type of deep filtration membranes, such as Emphaze depth filtration membranes; and another type of deep filtration membrane is used in the pre-filtration step for virus removal filtration, such as X0SP depth filtration membranes.

[0027] In a specific embodiment of the present invention, the deep filtration membranes provided in the clarification filtration step and the pre-filtration step of virus removal filtration are the same type of deep filtration membranes, for example, Emphaze depth filtration membranes; and another type of deep filtration membrane is provided in the intermediate product deep filtration step, for example, X0SP depth filtration membranes.

[0028] In a specific embodiment of the present invention, the deep filtration membranes set in the intermediate product deep filtration step and the pre-filtration step of virus removal filtration are the same type of deep filtration membranes, for example, Emphaze depth filtration membranes; and another type of deep filtration membrane is set in the clarification filtration step, for example, X0SP depth filtration membrane.

[0029] Preferably, the process flow of protein purification includes: clarification filtration, affinity capture, intermediate product deep filtration, virus inactivation, purification step, virus removal filtration and concentration and liquid exchange; the virus removal filtration includes prefiltration and nanofiltration.

[0030] Preferably, the polishing step comprises a first polishing step and a second polishing step which are arranged in series.

[0031] In one embodiment of the present invention, the purification step comprises two chromatography steps arranged in sequence, including a first chromatography step and a second chromatography step.

[0032] In the present invention, the process flow of protein purification comprises the following steps:

[0033] Step (1), a clarification filtration step, is used to remove cell debris.

[0034] Step (2), the capture step, is used for initial purification and capture of the target protein.

[0035] Step (3), the intermediate product deep filtration step, is used to remove impurities.

[0036] Step (4), the virus inactivation step, is used to inactivate viruses.

[0037] Step (5), a purification step, is used to remove impurities and includes a chromatography step using different fillers.

[0038] Step (6), the virus removal filtration step, includes prefiltration and nanofiltration, which are used to remove viruses.

[0039] Step (7), ultrafiltration concentration step, is used to concentrate and replace the liquid.

[0040] Preferably, the processing steps after setting the deep filtration membrane include: rinsing the deep filtration membrane with pure water and balancing the deep filtration membrane with a balancing liquid; starting the deep filtration step to load the sample to be processed; eluting the filter with a rinsing liquid, and ending the deep filtration step.

[0041] Preferably, the flow rate of the pure water flushing is 270-600LMH, for example, it can be 270LMH, 300LMH, 400LMH, 500LMH or 600LMH, and the flushing volume is greater than or equal to 100L / m 2 .

[0042] Preferably, the flow rate of the balancing liquid is 100-600LMH, for example, 100LMH, 200LMH, 300LMH, 400LMH, 500LMH or 600LMH, and the balancing volume is greater than or equal to 20L / m 2 .

[0043] Preferably, the flow rate of the sample loading is 100-300LMH, for example, 100LMH, 200LMH or 300LMH.

[0044] Preferably, the flow rate of the elution is 100-600LMH, for example, 100LMH, 200LMH, 300LMH, 400LMH, 500LMH or 600LMH, and the elution volume is greater than or equal to 30L / m 2 .

[0045] Preferably, the pH of the loaded sample is 6.5-7.5, for example, it can be 6.5, 7 or 7.5.

[0046] Preferably, the conductivity of the loaded sample is 0-100 mS / cm, for example, it can be 1 mS / cm, 10 mS / cm, 20 mS / cm, 40 mS / cm, 60 mS / cm, 80 mS / cm or 100 mS / cm, etc.

[0047] Preferably, the loading capacity of the sample is 2000 g / m 2 Within.

[0048] The present invention also provides a method for developing a pigment removal process, including: using colored intermediates to screen deep filter membranes, screening the ability of different functional deep filter membranes on the market to remove color, and obtaining deep filter membranes with color removal capabilities. The screened deep filter membranes are optimized and their functions are understood, including the pH, conductivity, loading capacity, flow rate, and elution conditions of the sample. The screened deep filter membranes and process conditions are applied to the clarification filtration step and then fine-tuned. The clarified product needs to be captured in one step to evaluate the color removal effect. The screened deep filter membranes and process conditions are applied to the virus removal filtration step and then fine-tuned.

[0049] In the present invention, the color determination needs to be concentrated to the same concentration as DS through an ultrafiltration concentration tube to effectively determine whether the color removal effect can achieve effective results. The comparison of color removal effects needs to be further distinguished by detecting the spectrum to further distinguish the difference in removal effects.

[0050] In a second aspect, the present invention provides application of the deep filtration process for removing pigments in protein purification described in the first aspect in a drug purification process.

[0051] The pigment removal process of the present invention is particularly suitable for the removal of pigments in the purification processes of antibodies, FC fusion proteins, bispecific antibodies and recombinant proteins, and has important application value in the preparation process of protein drugs.

[0052] The numerical range described in the present invention not only includes the point values ​​listed above, but also includes any point values ​​between the above numerical ranges that are not listed. Due to space limitations and for the sake of simplicity, the present invention no longer exhaustively lists the specific point values ​​included in the range.

[0053] Compared with the prior art, the present invention has the following beneficial effects:

[0054] (1) The present invention introduces a pigment removal process flow, providing a new solution and strategy for pigments that cannot be controlled and removed by upstream and chromatography.

[0055] (2) utilize the deep filter combination technology to remove the pigment that upstream and chromatography technology cannot control and remove. Specifically, mainly in the clarification step, the intermediate product deep filtration and the virus removal pre-filtration step respectively introduce the deep filtration membrane package, such as XOSP or Emphaze. Put the membrane package in three positions respectively to increase the removal efficiency and robustness of the pigment. Studies have shown that the present invention adopts disposable membrane package, not only does not need to worry about the influence of pigment on the life cycle of filler, but also can effectively remove pigment and have stronger robustness.

[0056] (3) The filter membrane of the present invention is disposable, and there is no need to worry about the problem of repeated service life caused by color contamination. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] Figure 1 It is the result of the pigment removal capacity of the same depth filter combination.

[0058] Figure 2 It is the result of the pigment removal capacity of different depth filter combinations. DETAILED DESCRIPTION

[0059] The technical solution of the present invention is further described below by specific implementation methods. It should be understood by those skilled in the art that the embodiments are only to help understand the present invention and should not be regarded as specific limitations of the present invention.

[0060] If no specific techniques or conditions are specified in the examples, the techniques or conditions described in the literature in the field or the product instructions are used. If no manufacturer is specified for the reagents or instruments used, they are all conventional products that can be purchased through regular channels.

[0061] The sources of the materials and raw materials used in the following examples are shown in Table 1.

[0062] Table 1

[0063] Serial number factory Representative membrane package types 1 Solventum <![CDATA[3M TM Emphaze TM Composite clarification membrane]]> 2 Merck Millistak+Pro Pod Depth Filter,X0SP Series

[0064] Example 1

[0065] This example investigates the color removal effect of the same type of deep filter membrane applied to different processes. X0SP deep filter membranes are applied to clarify filtration, intermediate product deep filtration, and pre-filtration steps of nanofiltration.

[0066] (1) Clarification and Filtration Steps

[0067] First, rinse the deep filter membranes with pure water. The flow rate is set to 600LMH and the flushing volume needs to be greater than or equal to 100L / m 2 Secondly, balance the deep filter membrane with a balance solution at a flow rate of 600LMH, and the balance volume is greater than or equal to 20L / m 2 The supernatant was obtained after centrifugation at 10000g for 30 minutes for sample loading, flow rate 300LMH, sample loading capacity 100L / m 2 Finally, the filter was eluted with eluent at a flow rate of 600 LMH and a volume greater than or equal to 30 L / m 2 The product is concentrated by one-step affinity capture and then color detected.

[0068] (2) Capture Steps

[0069] First, the chromatography column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution before loading the sample, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally the captured product is eluted.

[0070] (3) Intermediate product deep filtration step

[0071] First, rinse the deep filter membranes with pure water. The flow rate is set to 600LMH and the flushing volume needs to be greater than or equal to 100L / m 2 Secondly, balance the deep filter membrane with a balance solution at a flow rate of 600LMH, and the balance volume is greater than or equal to 20L / m 2 The captured product was loaded with a flow rate of 300LMH and a loading capacity of 1000g / m 2 Finally, the deep filter membrane is eluted with eluent at a flow rate of 600LMH, with a volume greater than or equal to 30L / m 2 The product is concentrated and its color is tested.

[0072] (4) Virus inactivation and purification step 1

[0073] The chromatographic column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution, and then the completed virus sample is loaded, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally a pure product 1 is eluted.

[0074] (5) Purification 2 steps

[0075] First, the chromatography column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution, and then the purified product 1 is loaded, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally the purified product 2 is eluted.

[0076] (6) Virus removal pre-filtration step

[0077] First, rinse the deep filter membranes with pure water. The flow rate is set to 600LMH and the flushing volume needs to be greater than or equal to 100L / m 2 Secondly, balance the deep filter membrane with a balance solution at a flow rate of 600LMH, and the balance volume is greater than or equal to 20L / m 2 The product of purified 2 was loaded with a flow rate of 300LMH and a loading capacity of 5000g / m 2 Finally, the deep filter membrane is eluted with eluent at a flow rate of 600LMH, with a volume greater than or equal to 30L / m 2 The product is concentrated and its color is tested.

[0078] The X0SP deep filter membrane was used in the clarification filtration, intermediate product deep filtration, and pre-filtration steps of nanofiltration, showing effective color removal effects, as shown in Table 2 and Figure 1 As shown; Table 2 shows the color change of the solution, Figure 1 Results for pigment removal capacity of the same depth filter combination.

[0079] Table 2

[0080]

[0081] From Table 2, it can be seen that after the sample material is filtered through the corresponding membrane package, the color is removed to a certain extent.

[0082] Example 2

[0083] This example investigates the color removal effects of different membrane packages, using the Emphaze deep filter membrane for clarifying filtration and the X0SP for intermediate deep filtration and pre-filtration steps of nanofiltration.

[0084] (1) Clarification and Filtration Steps

[0085] Use the balance solution to balance the depth filter membrane at a flow rate of 100LMH, and the balance volume is greater than or equal to 80L / m 2 The supernatant was obtained after centrifugation at 10000g for 10 minutes for sample loading, flow rate 100LMH, sample loading capacity 200L / m 2 Finally, the deep filter membrane is eluted with eluent at a flow rate of 100LMH, with a volume greater than or equal to 30L / m 2 The product is concentrated by one-step affinity capture and then color detected.

[0086] (2) Capture Steps

[0087] First, the chromatography column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution before loading the sample, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally the captured product is eluted.

[0088] (1) Intermediate product deep filtration step

[0089] First, rinse the deep filter membranes with pure water. The flow rate is set to 270LMH and the flushing volume needs to be greater than or equal to 100L / m 2 Secondly, balance the deep filter membrane with a balance solution at a flow rate of 100LMH, and the balance volume is greater than or equal to 20L / m 2 The affinity captured product was loaded with a flow rate of 100LMH and a loading capacity of 1000g / m 2 Finally, the deep filter membrane is eluted with eluent at a flow rate of 100LMH, with a volume greater than or equal to 30L / m 2 The product is concentrated and its color is tested.

[0090] (4) Virus inactivation and purification step 1

[0091] The chromatographic column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution, and then the completed virus sample is loaded, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally a pure product 1 is eluted.

[0092] (5) Purification 2 steps

[0093] First, the chromatography column is treated with more than 2 column volumes of water, sodium hydroxide and equilibrium solution, and then the purified product 1 is loaded, and impurities are removed by eluting with more than 2 column volumes of elution buffer, and finally the purified product 2 is eluted.

[0094] (3) Virus removal pre-filtration step

[0095] First, rinse the deep filter membranes with pure water. The flow rate is set to 270LMH and the flushing volume needs to be greater than or equal to 100L / m 2 Secondly, balance the deep filter membrane with a balance solution at a flow rate of 100LMH, and the balance volume is greater than or equal to 20L / m 2 The hydrophobic chromatography product was loaded with a flow rate of 100LMH and a loading capacity of 5000g / m 2 Finally, the deep filter membrane is eluted with eluent at a flow rate of 100LMH, with a volume greater than or equal to 30L / m 2 The product is concentrated and its color is tested.

[0096] The application of Emphaze depth filter membrane in clarifying filtration and X0SP in intermediate product depth filtration and pre-filtration steps of nanofiltration showed effective color removal, as shown in Table 3 and Figure 2 As shown in Table 3, the color change of the solution is shown. Figure 2 Results for pigment removal capacity of different depth filter combinations.

[0097] Table 3

[0098]

[0099]

[0100] From Table 3, it can be seen that after the sample material is filtered through the corresponding membrane package, the color is removed to a certain extent.

[0101] In summary, the present invention provides a method for effectively removing pigments, and provides a new solution for pigments that cannot be controlled and removed by upstream and chromatography. Compared with conventional color removal solutions, the method of the present invention is lower in cost and more convenient, and has important application value in protein purification.

[0102] The applicant declares that the above is only a specific implementation mode of the present invention, but the protection scope of the present invention is not limited thereto. Those skilled in the art should understand that any changes or substitutions that can be easily thought of by those skilled in the art within the technical scope disclosed by the present invention shall fall within the protection scope and disclosure scope of the present invention.

Claims

1. A deep filtration process for removing pigments in protein purification, characterized in that: The process comprises: introducing a deep filter membrane into the filtration step of protein purification to achieve filtration and pigment removal simultaneously; The filtration step includes: any one or a combination of at least two of a clarification filtration step, an intermediate product deep filtration step, or a pre-filtration step for virus removal filtration; The deep filtration membrane is selected from: Emphaze deep filtration membrane and / or X0SP deep filtration membrane.

2. The deep filtration process for removing pigments in protein purification according to claim 1, characterized in that: At least one of the deep filtration membranes is provided in any filtration step; Preferably, when the same type of deep filtration membrane is arranged in any filtration step, the same type of deep filtration membrane is arranged in parallel or in series; Preferably, when different types of deep filtration membranes are arranged in any one filtration step, the different types of deep filtration membranes are arranged in series.

3. The deep filtration process for removing pigments in protein purification according to claim 1 or 2, characterized in that: The same type of deep filtration membrane is used in the clarification filtration step, the intermediate product deep filtration step, and the virus removal pre-filtration step.

4. The deep filtration process for removing pigments in protein purification according to claim 1 or 2, characterized in that: Any two of the clarification filtration step, the intermediate product deep filtration step or the pre-filtration step for virus removal filtration are provided with the same type of deep filter membrane, and the other step is provided with another type of deep filter membrane.

5. A deep filtration process for removing pigments in protein purification according to any one of claims 1 to 4, characterized in that: The process flow of protein purification includes: clarification filtration, affinity capture, intermediate product deep filtration, virus inactivation, purification step, virus removal filtration and concentration and liquid replacement; the virus removal filtration includes prefiltration and nanofiltration; Preferably, the polishing step comprises a first polishing step and a second polishing step which are arranged in series.

6. A deep filtration process for removing pigments in protein purification according to any one of claims 1 to 5, characterized in that: The processing steps after the deep filter membrane is set include: washing the deep filter membrane with pure water and balancing the deep filter membrane with a balancing liquid; starting the deep filtration step to load the sample to be processed; eluting the filter with an eluting liquid, and ending the deep filtration step.

7. The deep filtration process for removing pigments in protein purification according to claim 6, characterized in that: The flow rate of the pure water flushing is 270-600LMH, and the flushing volume is greater than or equal to 100L / m 2 ; Preferably, the flow rate of the balance liquid is 100-600LMH, and the balance volume is greater than or equal to 20L / m 2 .

8. The deep filtration process for removing pigments in protein purification according to claim 6 or 7, characterized in that: The flow rate of the sample loading is 100-300LMH; Preferably, the flow rate of the elution is 100-600LMH, and the volume of the elution is greater than or equal to 30L / m 2 .

9. A deep filtration process for removing pigments in protein purification according to any one of claims 6 to 8, characterized in that: The pH of the loaded sample is 6.5-7.5; Preferably, the conductivity of the loaded sample is 0-100 mS / cm; Preferably, the loading capacity of the sample is 2000 g / m 2 Within.

10. Use of the deep filtration process for removing pigments in protein purification according to any one of claims 1 to 9 in a drug purification process.

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