CN111068360A - Combined chromatographic device and continuous flow chromatographic method - Google Patents
Combined chromatographic device and continuous flow chromatographic method Download PDFInfo
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- 238000002360 preparation method Methods 0.000 claims description 22
- 238000005406 washing Methods 0.000 claims description 13
- 229960000397 bevacizumab Drugs 0.000 claims description 9
- 239000003124 biologic agent Substances 0.000 claims description 7
- 238000000746 purification Methods 0.000 claims description 6
- 229960002964 adalimumab Drugs 0.000 claims description 4
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- 229960003876 ranibizumab Drugs 0.000 claims description 4
- 229960004641 rituximab Drugs 0.000 claims description 4
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- B01D—SEPARATION
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- B01D15/08—Selective adsorption, e.g. chromatography
- B01D15/10—Selective adsorption, e.g. chromatography characterised by constructional or operational features
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Abstract
The invention provides a combined chromatography device and a continuous flow chromatography method. This combined chromatography device includes a plurality of basic chromatography equipment, a plurality of chromatography columns and a plurality of multi-way valve, basic chromatography equipment is used for controlling fluid inflow or outflow the chromatography column, basic chromatography equipment includes post position valve and outlet valve, wherein, every the import of chromatography column passes through the multi-way valve is with a plurality of two at least in the basic chromatography equipment link to each other, every the export of chromatography column with at least one basic chromatography equipment links to each other, combined chromatography device can realize the continuous flow chromatography operation of multithread way. By using the combined chromatography device, the continuous flow chromatography of the multi-flow path multi-chromatography column can be realized on the basis of the existing basic chromatography equipment and the existing chromatography column.
Description
Reference to related applications
The present invention claims priority from an invention patent application entitled "Combined chromatography device and continuous flow chromatography method", application number 201811228757.0, filed in China on 10/22/2018, which is incorporated herein by reference in its entirety.
Technical Field
The present invention relates to the field of biotechnology, in particular to chromatography devices, and more particularly to a combined chromatography device and a method for continuous flow chromatography thereof.
Background
The existing chromatographic device is generally composed of a chromatographic apparatus and a chromatographic column. For example, a common single-column chromatography apparatus, which is composed of a single chromatography device connected to a single chromatography column, can perform purification only by operating one flow path at a time, and in an actual experimental stage and a pilot-scale and production stage, continuous flow chromatography is often performed by using multiple chromatography columns.
Continuous flow chromatography devices have multiple modes of operation, primarily to achieve simultaneous operation of two flow paths, one for continuous loading and the other for pre-and post-loading processing operations.
Two flow paths connecting two chromatography columns are the simplest mode; further, three columns may be connected in two flow paths, two for loading and one for pre and post treatment. In addition, there are also devices using four or more columns, but in such devices, only two (or three, with separate pre and post treatments) flow paths are actually operating simultaneously.
Currently mainstream continuous flow chromatography devices have five of the following:
AKTApc is a continuous flow chromatography device from general electric company that supports two-pass three-column, or three-pass four-column operation. But it cannot realize the liquid phase direction switching of the chromatographic column and has higher price.
BioSMB is a continuous flow chromatography device from Pall corporation that supports a two-stream multi-column mode of operation. However, it cannot realize gradient of buffer solution, has no collector, cannot collect elution products of different chromatographic columns respectively, and does not have a function of collecting products according to an ultraviolet absorption value.
BioSC is a continuous flow chromatography device from Novasep, which supports two flow paths and three columns operating modes. However, it cannot realize gradient of buffer solution, has no collector, cannot collect elution products of different chromatographic columns respectively, and cannot realize switching of liquid phase direction of the chromatographic columns.
Contichrom CUBE is a continuous flow chromatography device from ChromaCon, which has an operation mode of two columns with two flow paths, and the device has a simple structure and is not prone to errors. However, it requires the purchase of a complete set of equipment from the laboratory stage to the production stage, is less flexible and in use can sometimes only achieve semi-continuous flow chromatography.
There are also some improvements in the art, for example, chinese patent application 201510568307.6 discloses a method of retrofitting a chromatograph for perfusion culture purification and a solution to retrofit AKTAexplorer to a continuous flow chromatography device. Chinese patent application 201710594337.3 discloses an improved chromatographic apparatus and a method for continuous flow chromatography, which discloses a scheme of a two-flow path three-chromatographic column continuous flow chromatographic apparatus composed of an improved AKTApure chromatographic apparatus. However, the above scheme needs to change the use of the valve and the component, which is easy to cause the discrepancy between the instruction and the actual function and the confusion; meanwhile, the original functions of the equipment are easily influenced, and the improved technical scheme can only be used in a laboratory scale, cannot be amplified, cannot meet the requirements of a pilot plant test or production, and still needs to purchase the equipment meeting the requirements of the pilot plant test and the production scale.
The continuous flow chromatography device needs to provide equipment meeting the corresponding scale in different stages, which causes that different equipment needs to be provided in the experimental stage and the production stage to meet the operation requirements of different stages, the equipment investment cost is high, and the equipment idle rate is high. Moreover, different equipment can only meet the requirements of the current stage, and the equipment can be provided with a plurality of unique operation methods, so that the requirement on operators is high, and errors are easy to occur in the operation process of the equipment at different stages.
Disclosure of Invention
In view of the above-mentioned problems, the present invention provides a combined chromatography device constructed using existing resources.
According to a first aspect of the present invention, there is provided a combined chromatography apparatus comprising a plurality of basic chromatography devices for controlling the flow of fluid into or out of a chromatography column, a plurality of chromatography columns and a plurality of multi-way valves, wherein the basic chromatography devices comprise a column position valve and an outlet valve,
the inlet of each chromatographic column is connected with at least two of the basic chromatographic devices through the multi-way valve, the outlet of each chromatographic column is connected with at least one basic chromatographic device, and the combined chromatographic device can realize continuous flow chromatographic operation of multiple flow paths.
In at least one embodiment, the outlet of each chromatography column is connected to the inlet of another chromatography column via the multi-way valve.
In at least one embodiment, the number of the plurality of basic chromatography devices is two or three, the number of the plurality of chromatography columns is two, three or more, and the multi-way valve is a three-way valve, a four-way valve, a five-way valve or a six-way valve.
In at least one embodiment, the base chromatography device comprises two column valves, the inlet of the chromatography column is connected with one of the column valves or the outlet valve of the base chromatography device through the multi-way valve, and the outlet of the chromatography column is connected with the other column valve of the base chromatography device.
In at least one embodiment, the number of the basic chromatography devices is the same as the number of the chromatography columns, and is N, N is a positive integer greater than or equal to 2, the inlet of the first chromatography column is connected with one column position valve of the first basic chromatography device and the outlet valve of the Nth basic chromatography device through a multi-way valve, the outlet of the first chromatography column is connected with the other column position valve of the first basic chromatography device, the inlet of the second chromatography column is connected with one column position valve of the second basic chromatography device and the outlet valve of the first basic chromatography device through a multi-way valve, the outlet of the second chromatography column is connected with the other column position valve of the second basic chromatography device, and by analogy, the inlet of the Nth chromatographic column is connected with one column position valve of the Nth basic chromatographic equipment and the outlet valve of the N-1 th basic chromatographic equipment through the multi-way valve, and the outlet of the Nth chromatographic column is connected with the other column position valve of the Nth basic chromatographic equipment.
In at least one embodiment, the number of the chromatographic columns is larger than that of the basic chromatographic equipment, the inlet and the outlet of each chromatographic column are connected with the column position valves of at least two basic chromatographic devices through the multi-way valves, and the inlet of each chromatographic column is also connected with the outlet of another chromatographic column through the multi-way valve.
In at least one embodiment, the number of the basic chromatography devices and the number of the chromatography columns are both two, the inlet of the first chromatography column is connected with one of the column-level valves of the first basic chromatography device and the outlet valve of the second basic chromatography device through a first three-way valve, and the outlet of the first chromatography column is connected with the other column-level valve of the first basic chromatography device; the inlet of the second chromatographic column is connected with one column position valve of the second basic chromatographic device and the outlet valve of the first basic chromatographic device through a second three-way valve, and the outlet of the second chromatographic column is connected with the other column position valve of the second basic chromatographic device.
In at least one embodiment, the number of the basic chromatography devices and the number of the chromatography columns are three, the inlet of the first chromatography column is connected with one of the column-level valves of the first basic chromatography device and the outlet valve of the third basic chromatography device through a first three-way valve, and the outlet of the first chromatography column is connected with the other column-level valve of the first basic chromatography device; the inlet of the second chromatographic column is connected with one column position valve of the second basic chromatographic equipment and the outlet valve of the first basic chromatographic equipment through a second three-way valve, and the outlet of the second chromatographic column is connected with the other column position valve of the second basic chromatographic equipment; the inlet of the third chromatographic column is connected with one column position valve of the third basic chromatographic device and the outlet valve of the second basic chromatographic device through a third three-way valve, and the outlet of the third chromatographic column is connected with the other column position valve of the third basic chromatographic device.
In at least one embodiment, the number of the chromatography columns is three, the number of the basic chromatography devices is two, two four-way valves are connected to two column position valves of each of the two basic chromatography devices, one four-way valve is connected to an inlet and an outlet of each of the three chromatography columns, an inlet of each chromatography column is connected to one column position valve of the two basic chromatography devices and an outlet of the other chromatography column through one four-way valve, and an outlet of each chromatography column is connected to the other column position valve of the two basic chromatography devices and an inlet of the other chromatography column through one four-way valve.
In at least one embodiment, any one of the branch flow paths of the multi-way valve is provided with a check valve.
According to a second aspect of the present invention, there is provided a method of continuous flow chromatography of a fluid using a combined chromatography apparatus according to the present invention, the method comprising switching the passage of the multi-way valve to selectively perform a single-column continuous flow chromatography operation or a multi-column continuous flow chromatography operation.
In at least one embodiment, the method comprises,
and switching the multi-way valve connected with the inlet of one of the plurality of chromatographic columns to a mode connected with the column position valve, and switching the multi-way valve connected with the outlet of the chromatographic column to a mode connected with the multi-way valve connected with the inlet of the other chromatographic column, thereby continuously loading at least two chromatographic columns.
In at least one embodiment, the method comprises,
and switching the multi-way valve connected with the inlet of one of the plurality of chromatographic columns to a mode connected with the column position valve and switching the multi-way valve connected with the outlet of the chromatographic column to a mode connected with the column position valve, thereby washing and eluting the chromatographic column.
According to a third aspect of the present invention there is provided a method of purifying a biological agent, the method comprising: the combined chromatographic device according to the invention is used to perform continuous flow chromatography on biological agents to obtain purified products.
In at least one embodiment, the biological agent is an antibody agent; further, the antibody is any one of the group consisting of certolizumab, adalimumab, bevacizumab, apraxizumab, infliximab, nimotuzumab, pertuzumab, ranibizumab, rituximab, and trastuzumab.
By using the combined chromatography device, the continuous flow chromatography of a plurality of chromatography columns with multiple flow paths can be realized on the basis of the existing basic chromatography equipment and the existing chromatography columns.
Drawings
FIG. 1 is a schematic flow path diagram of a combined chromatography device according to a first embodiment of the invention.
FIG. 2 is a schematic flow path diagram of the combined chromatography device of FIG. 1 with continuous flow loading controlled by the first base chromatography apparatus P1.
FIG. 3 is a schematic flow diagram of the first column C1 elution by washing and the second column C2 loading of the combined chromatography device of FIG. 1.
FIG. 4 is a schematic flow path diagram of the combined chromatography device of FIG. 1 with continuous flow loading controlled by the second basic chromatography apparatus P2.
FIG. 5 is a schematic flow diagram of the flow path for washing and eluting the second chromatography column C2 and loading the first chromatography column C1 in the combined chromatography device of FIG. 1.
FIG. 6 is a schematic flow path diagram of a combined chromatography device according to a second embodiment of the invention.
FIG. 7 is a schematic flow path diagram of a combined chromatography device according to a third embodiment of the invention.
Fig. 8 is a continuous flow chromatography test chromatogram for performing an affinity chromatography experiment on bevacizumab using a combined chromatography device according to a first embodiment of the present invention.
Wherein the dashed lines in fig. 1 and 7 represent alternative flow paths; the two-dot chain lines in fig. 2 to 5 indicate the unconnected flow paths.
Detailed Description
Exemplary embodiments of the present invention are described below with reference to the accompanying drawings. It should be understood that the detailed description is intended only to teach one skilled in the art how to practice the invention, and is not intended to be exhaustive or to limit the scope of the invention.
The invention relates to a combined chromatographic device which combines a plurality of basic chromatographic devices and a plurality of chromatographic columns into a multi-flow path. Among them, the basic chromatography apparatus, also called a chromatograph, can be connected to a chromatography column to provide a function of controlling a flow path for inflow and outflow of a preparation to the chromatography column, and can be further equipped with corresponding components to perform functions of preparation mixing, UV detection, conductivity detection, pH detection, and the like, as required. The chromatography device formed by connecting a chromatography column with a basic chromatography device can realize functions such as automatic solution configuration, multiple buffer and preparation (sample) inlets, gradient pulling, bubble trap, automatic collection, automatic peak cutting, program editing, program operation, manual control and the like according to needs.
First embodiment
Referring to fig. 1 to 5, a combined chromatography apparatus and a method for continuous flow chromatography using the same according to a first embodiment of the present invention will be described.
The chromatography apparatus according to the first embodiment, as shown in fig. 1, is a continuous flow chromatography apparatus comprising two flow paths and two chromatography columns, comprising two basic chromatography devices, two chromatography columns and two three-way valves.
The basic chromatographic equipment, chromatographic column and three-way valve can be selected from those conventional in the artDevices, for example, first basic chromatography device P1 and second basic chromatography device P2 can be selected from the group consisting of AKTA avant 150 type chromatography devices available from GE corporation, which include pumps, mixing cells, pump inlet valves, column level valves, UV detectors, conductivity detectors, pH detectors, and outlet valves; the first chromatographic column C1 and the second chromatographic column C2 can be VL11 type chromatographic columns of millipore company; first three-way valve V1And a second three-way valve V2Related accessories provided by GE corporation may be selected.
First three-way valve V1Are respectively defined as
Second three-way valve V2Are respectively defined as
As shown in fig. 1, for the first three-way valve V1Which is
Port connected to the upper column valve V of the first basic chromatography device P1aAnd it
The port was connected to the inlet of the upper part of the first chromatographic column C1, which
The port is connected to the outlet valve V of the second basic chromatography device P2c(ii) a For the second three-way valve V2Which is
A column valve V with a port connected to the upper part of the basic chromatography device P2aAnd it
An inlet connected to the upper part of the chromatographic column C2, which
Mouth connecting pieceOutlet valve V connected to the first basic chromatography device P1c。
For two basic chromatography apparatuses, the lower column position valve V of the first basic chromatography apparatus P1bA column position valve V connected to the lower outlet of the chromatography column C1 and the lower part of the second basic chromatography device P2bConnected to the outlet of the lower part of the chromatography column C2. Optionally, each basic chromatography device is provided with a gradient-adjustable buffer configuration, and a plurality of collectors are arranged to collect elution products of different chromatography columns respectively.
The present embodiment is configured by arranging two three-way valves (V)1、V2) Two basic chromatographic devices (P1, P2) and two chromatographic columns (C1, C2) are connected to form a two-flow two-column continuous chromatographic flow device. The continuous flow chromatography device has a continuous flow chromatography running mode of continuous flow sample loading, sample loading of one chromatographic column and elution of the other chromatographic column, and can realize an independent single-column chromatography running mode.
The continuous flow chromatography method of the two-flow path two-column combined chromatography apparatus will be described with reference to FIGS. 2 to 5.
(i) Continuous flow loading controlled by a first basic chromatography device P1
Referring to fig. 2, a first three-way valve V is provided1Is/are as follows
Mouth and
the ports are communicated, i.e., the column position valve V of the upper part of the first basic chromatography device P1aIs communicated with the inlet of the first chromatographic column C1; a second three-way valve V2Is/are as follows
Mouth and
the ports are communicated, i.e., the outlet valve V of the first basic chromatography device P1cIs communicated with the inlet of a second chromatographic column C2. FromMeanwhile, the two chromatographic columns (C1, C2) are loaded under the control of a first basic chromatographic device P1, and the two basic chromatographic devices (P1, P2) operate simultaneously; wherein the first basic chromatography device P1 is loaded, the second basic chromatography device P2 opens the flow path, and the preparation passes through the upper column position valve V of the first basic chromatography device P1aEnters a first chromatographic column C1 and then passes through a lower column position valve V of a first basic chromatographic device P1bOutlet valve VcEnters a second chromatographic column C2 and then passes through a lower column position valve V of a second basic chromatographic device P2bAnd an outlet valve VcAnd (4) discharging waste liquid. In this way, the preparation was loaded continuously via the first C1 and second C2 chromatography columns.
(ii) Eluting with a first C1 eluent, and loading with a second C2 chromatographic column
Referring to fig. 3, a first three-way valve V1The inlet of the first chromatographic column C1 was connected to a first basic chromatographic apparatus P1, a second three-way valve V2The inlet of the second chromatography column C2 was connected to a second basic chromatography device P2. In this way, the first basic chromatography device P1 and the first chromatography column C1 operate as a single unit, and the second basic chromatography device P2 and the second chromatography column C2 operate as a single unit. In this process, the outlet valve V of the first basic chromatography device P1 is controlledcSwitching to waste liquid discharge and switching outlet valve V according to elution parameters at proper timecTo the sample collection port S, thereby completing the elution of the first chromatographic column C1 and obtaining the eluted product; the second basic chromatographic equipment P2 is controlled to directly load, and the preparation flows through the second chromatographic column C2 and then is discharged through an outlet valve V of P2cAnd (4) discharging waste liquid.
(iii) Continuous flow loading controlled by a second basic chromatography device P2
Referring to fig. 4, the second three-way valve V is2Is/are as follows
Mouth and
port communication, i.e., a column position valve V for the upper part of the second basic chromatography device P2aAnd a firstThe inlets of the two chromatographic columns C2 are communicated; a first three-way valve V1Is/are as follows
Mouth and
the ports communicate, i.e., the outlet valve V of the second basic chromatography device P2cCommunicating with the inlet of the first chromatographic column C1. Thus, both columns (C1, C2) are loaded by the second basic chromatographic apparatus P2, and the two basic chromatographic apparatuses (P1, P2) operate simultaneously; wherein the second basic chromatography device P2 is loaded, the first basic chromatography device P1 opens the flow path, and the preparation passes through the upper column position valve V of the second basic chromatography device P2aEnters a second chromatographic column C2 and then passes through a lower column position valve V of a second basic chromatographic device P2bOutlet valve VcEnters a first chromatographic column C1 and then passes through a lower column position valve V of a first basic chromatographic device P1bAnd an outlet valve VcAnd (4) discharging waste liquid. In this way, the preparation was continuously loaded via the second C2 and first C1 chromatography columns.
(iv) Washing and eluting the second chromatographic column C2, and loading the first chromatographic column C1
Referring to fig. 5, a first three-way valve V1The inlet of the first chromatographic column C1 was connected to a first basic chromatographic apparatus P1, a second three-way valve V2The inlet of the second chromatography column C2 was connected to a second basic chromatography device P2. In this way, the first basic chromatography device P1 and the first chromatography column C1 operate as a single unit, and the second basic chromatography device P2 and the second chromatography column C2 operate as a single unit. In this process, the outlet valve V of the second basic chromatography device P2 is controlledcSwitching to waste liquid discharge and switching outlet valve V according to elution parameters at proper timecTo the sample collection port S, thereby completing the elution of the second chromatography column C2 and obtaining the eluted product; the first basic chromatography device P1 is controlled to directly load, and the preparation flows through the first chromatographic column C1 and then passes through the outlet valve V of the first basic chromatography device P1cAnd (4) discharging waste liquid.
The chromatography device is provided withTwo-way flow continuous flow chromatography function is provided by two chromatography columns, by controlling two three-way valves (V)1、V2) Can selectively realize continuous flow loading of two chromatographic columns (C1, C2), or realize flushing elution of one chromatographic column and loading of the other chromatographic column. Of course, it is expected by those skilled in the art that the four operation modes (i), (ii), (iii) and (iv) described above can be combined and the order can be changed according to the actual experiment or production requirement.
Next, experimental data for purifying bevacizumab using the combined chromatography device of the present embodiment will be described.
Purification experiments of continuous flow chromatography of bevacizumab were performed using a conventional single column chromatography device and a two-flow path two-chromatography column combined chromatography device according to the present embodiment, respectively, and the experimental data are shown in table 1.
TABLE 1 comparison of purification Experimental quality data for bevacizumab
The data in table 1 show that the samples prepared using the apparatus and method of the present invention do not differ significantly from the samples prepared using conventional single column chromatography apparatus in terms of both quality (e.g., purity and impurities) and process performance (e.g., yield).
The combined chromatography apparatus according to the present embodiment was used to perform affinity chromatography experiments on bevacizumab, wherein the filler was Mabselect SURE from GE corporation. FIG. 8 is an experimentally obtained continuous flow chromatography test chromatogram, wherein FIG. 8(a) is the chromatogram of the first column C1 and FIG. 8(b) is the chromatogram of the second column C2. In the figure, the solid line indicates the ultraviolet UV280 collection curve and the dashed line indicates the start of loading. According to the chromatography spectrum, the combined chromatography device can achieve the chromatography quality of the traditional single-column chromatography device; in addition, the combined chromatographic device can support a plurality of chromatographic columns to run simultaneously, so that the production efficiency and the utilization rate of equipment and materials are improved, and meanwhile, the solvent consumption and the preparation cost are reduced.
Second embodiment
Referring to fig. 6, the second embodiment is a modification of the first embodiment, and a single chromatography column is added to the first embodiment, thereby constituting a two-channel three-column combined chromatography apparatus. The device comprises two basic chromatographic devices (P1, P2) and three chromatographic columns (C1, C2, C3). Further, due to the increase in the number of chromatography columns, a four-way valve is used in the present embodiment instead of the three-way valve in the first embodiment to connect the chromatography columns with the basic chromatography apparatus.
The inlet and outlet of each chromatography column and the respective column position valve of the base chromatography device are connected with a four-way valve, defined as the first four-way valve V'1And a second four-way valve V'2And a third four-way valve V'3And a fourth four-way valve V'4And a fifth four-way valve V'5And a sixth four-way valve V'6And a seventh four-way valve V'7And an eighth four-way valve V'8And a ninth four-way valve V'9And fourteenth through valve V'10. First four-way valve V'1Four outlets are respectively defined as
Second four-way valve V'2Four outlets are respectively defined as
N-th four-way valve V'nFour outlets are respectively defined as
The connection mode of each four-way valve is shown in fig. 6 and is expressed as follows:
first four-way valve V'1Is/are as follows
A column valve V connected to the upper part of the first basic chromatography device P1aThe connection is carried out by connecting the two parts,
port and second four-way valve V'2Is/are as follows
The mouth is connected with the water inlet pipe,
port and third four-way valve V'3Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourth four-way valve V'4Is/are as follows
A port connection;
second four-way valve V'2Is/are as follows
The port was connected to the inlet of the first chromatographic column C1,
port and first four-way valve V'1Is/are as follows
The mouth is connected with the water inlet pipe,
port and fifth four-way valve V'5Is/are as follows
The mouth is connected with the water inlet pipe,
port and ninth four-way valve V'9Is/are as follows
A port connection;
third four-way valve V'3Is/are as follows
The port is connected to the inlet of a second chromatography column C2,
port and first four-way valve V'1Is/are as follows
The mouth is connected with the water inlet pipe,
port and fifth four-way valve V'5Is/are as follows
The mouth is connected with the water inlet pipe,
port and seventh four-way valve V'7Is/are as follows
A port connection;
fourth four-way valve V'4Is/are as follows
The port is connected to the inlet of a third chromatographic column C3,
port and first four-way valve V'1Is/are as follows
Oral connection，
Port and fifth four-way valve V'5Is/are as follows
The mouth is connected with the water inlet pipe,
port and eighth four-way valve V'8Is/are as follows
A port connection;
fifth and fourth valve V'5Is/are as follows
A column position valve V connected with the upper part of the second basic chromatographic equipment P2aThe connection is carried out by connecting the two parts,
port and second four-way valve V'2Is/are as follows
The mouth is connected with the water inlet pipe,
port and third four-way valve V'3Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourth four-way valve V'4Is/are as follows
A port connection;
sixth four-way valve V'6Is/are as follows
A column valve V with a port communicating with the lower part of the first basic chromatography device P1bThe connection is carried out by connecting the two parts,
port and seventh four-way valve V'7Is/are as follows
The mouth is connected with the water inlet pipe,
port and eighth four-way valve V'8Is/are as follows
The mouth is connected with the water inlet pipe,
port and ninth four-way valve V'9Is/are as follows
A port connection;
seventh four-way valve V'7Is/are as follows
The port is connected to the outlet of the first chromatographic column C1,
port and sixth four-way valve V'6Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourteenth through valve V'10Is/are as follows
The mouth is connected with the water inlet pipe,
port and third four-way valve V'3Is/are as follows
A port connection;
eighth four-way valve V'8Is/are as follows
The port is connected with the outlet of the second chromatographic column C2,
port and sixth four-way valve V'6Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourteenth through valve V'10Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourth four-way valve V'4Is/are as follows
A port connection;
ninth four-way valve V'9Is/are as follows
The port is connected to the outlet of the third chromatographic column C3,
port and sixth four-way valve V'6Is/are as follows
The mouth is connected with the water inlet pipe,
port and fourteenth through valveV’10Is/are as follows
The mouth is connected with the water inlet pipe,
port and second four-way valve V'2Is/are as follows
A port connection;
fourteenth valve V'10Is/are as follows
A column valve V communicating with the lower part of the second basic chromatography device P2bThe connection is carried out by connecting the two parts,
port and seventh four-way valve V'7Is/are as follows
The mouth is connected with the water inlet pipe,
port and eighth four-way valve V'8Is/are as follows
The mouth is connected with the water inlet pipe,
port and ninth four-way valve V'9Is/are as follows
The ports are connected.
As can be seen from the connection relationship among the four-way valves, the inlet and the outlet of each chromatographic column are respectively connected with a column position valve V at the upper part of a basic chromatographic device through one four-way valveaAnd a column valve V at the lower part of the basic chromatographic equipmentbConnecting; and the remaining branch road of the four-way valve can be combed and arranged as follows: outlet of the first chromatographic column C1The cross four-way valve was connected to the inlet of the second chromatography column C2, the outlet of the second chromatography column C2 was connected to the inlet of the third chromatography column C3 through the four-way valve, and the outlet of the third chromatography column C3 was connected to the inlet of the chromatography column C1 through the four-way valve, so that the three chromatography columns were connected in series.
The continuous flow chromatography method of the two-channel three-column chromatography apparatus is described below.
(i) Continuous flow sample loading
Either of the basic chromatographic apparatuses is selected for the controlled loading, for example the first basic chromatographic apparatus P1 is selected for the controlled continuous flow loading, and two or three columns may be selected for the number of columns loaded. For example,
(1) the three chromatographic columns are all loaded with the sample,
the first basic chromatography device P1 controls the continuous flow loading, and the preparation passes through the column position valve V at the upper part of the first basic chromatography device P1aSequentially flows through a first four-way valve V'1And a second four-way valve V'2A first chromatographic column C1 and a seventh four-way valve V'7And a third four-way valve V'3A second chromatographic column C2 and an eighth four-way valve V'8And a fourth four-way valve V'4A third chromatographic column C3 and a ninth four-way valve V'9Fourteenth through valve V'10And a lower column position valve V of the second basic chromatography device P2bThen via the outlet valve V of the second basic chromatography device P2cDischarging waste liquid;
alternatively, the first and second electrodes may be,
(2) two chromatography columns (e.g. C1 and C2) were loaded,
the first basic chromatography device P1 controls the continuous flow loading, and the preparation passes through the column position valve V at the upper part of the first basic chromatography device P1aSequentially flows through a first four-way valve V'1And a second four-way valve V'2A first chromatographic column C1 and a seventh four-way valve V'7And a third four-way valve V'3A second chromatographic column C2 and an eighth four-way valve V'8Fourteenth through valve V'10And a lower column position valve V of the second basic chromatography device P2bThen via the outlet valve V of the second basic chromatography device P2cAnd (4) discharging waste liquid. It should be understood thatSo as to control the loading of the second chromatography column C2 and the third chromatography column C3, or the third chromatography column C3 and the first chromatography column C1 by changing the conduction path of the four-way valve as required.
It will be appreciated that the apparatus may also be adapted to control continuous flow loading using a second basic chromatographic apparatus P2, or to control loading simultaneously using two basic chromatographic apparatuses (P1, P2) (e.g.P 1 for columns C1, C2 and P2 for column C3). By using the chromatography device according to the present embodiment, the basic chromatography equipment and the chromatography column can be flexibly configured according to actual sample loading requirements, thereby meeting different requirements in experimental stages or stages such as pilot plant test and production.
(ii) Washing and eluting one chromatographic column, and loading the rest chromatographic columns
The inlets of the three chromatographic columns are respectively connected to the column position valves V on the upper parts of the two basic chromatographic devicesaAnd the outlets of the three chromatographic columns are respectively connected to the column position valves V at the lower parts of the two basic chromatographic devicesbWhile the outlet of one column is connected to the inlet of the next column, when the elution operation of the columns is performed, any one of the basic chromatographic apparatuses P1/P2 can be used to elute any one of the three columns C1/C2/C3, while the other basic chromatographic apparatus is used to control the loading of the other two columns, for example,
(1) p2 control the column C1 to wash and elute, P1 control the column C2, C3 to load,
adjusting the switching flow path of each four-way valve to ensure that the flow paths of elution are as follows in sequence: column position valve V of the upper part of the second basic chromatography device P2aAnd a fifth four-way valve V'5And a second four-way valve V'2A first chromatographic column C1 and a seventh four-way valve V'7Fourteenth through valve V'10And a lower column position valve V of the second basic chromatography device P2bThen through the outlet valve V of the second basic chromatography device P2cDischarging waste liquid, and timely switching outlet valve V according to elution parameterscTo the sample collection port S, thereby completing the elution of the first chromatographic column C1 by washing and obtaining the eluted product. In this procedure, the first basic chromatography device P1 was loadedUpper column valve V of P1aSequentially conducting first four-way valve V'1And a third four-way valve V'3A second chromatographic column C2 and an eighth four-way valve V'8And a fourth four-way valve V'4A third chromatographic column C3 and a ninth four-way valve V'9And a sixth four-way valve V'6And a lower column position valve V of the first basic chromatography device P1bThen passes through the outlet valve V of the first basic chromatography device P1cDischarging waste liquid;
alternatively, the first and second electrodes may be,
(2) p2 control the column C2 to wash and elute, P1 control the column C3, C1 to load,
adjusting the switching flow path of each four-way valve to ensure that the flow paths of elution are as follows in sequence: column position valve V of the upper part of the second basic chromatography device P2aAnd a fifth four-way valve V'5And a third four-way valve V'3A second chromatographic column C2 and an eighth four-way valve V'8Fourteenth through valve V'10A lower column valve V of the second basic chromatography device P2bThen through the outlet valve V of the second basic chromatography device P2cDischarging waste liquid, and timely switching outlet valve V according to elution parameterscTo the sample collection port S, thereby completing the elution of the second chromatography column C2 and obtaining the eluted product. In this process, the upper column position valve V of the first basic chromatography device P1aSequentially conducting first four-way valve V'1And a fourth four-way valve V'4A third chromatographic column C3 and a ninth four-way valve V'9And a second four-way valve V'2A first chromatographic column C1 and a seventh four-way valve V'7And a sixth four-way valve V'6And a lower column position valve V of the first basic chromatography device P1bThen passes through the outlet valve V of the first basic chromatography device P1cDischarging waste liquid;
in the alternative to this, either,
(3) p2 control the column C3 to wash and elute, P1 control the column C1, C2 to load,
adjusting the switching flow path of each four-way valve to ensure that the flow paths of elution are as follows in sequence: column position valve V of the upper part of the second basic chromatography device P2aAnd a fifth four-way valve V'5And a fourth four-way valve V'3The third layerChromatographic column C3 and ninth four-way valve V'9Fourteenth through valve V'10A lower column valve V of the second basic chromatography device P2bThen through the outlet valve V of the second basic chromatography device P2cDischarging waste liquid, and timely switching outlet valve V according to elution parameterscTo the sample collection port S, thereby completing the elution of the third chromatographic column C3 by washing and obtaining the eluted product. In this process, the upper column position valve V of the first basic chromatography device P1aSequentially conducting first four-way valve V'1And a second four-way valve V'2A first chromatographic column C1 and a seventh four-way valve V'7And a third four-way valve V'3A second chromatographic column C2 and an eighth four-way valve V'8And a sixth four-way valve V'6And a lower column position valve V of the first basic chromatography device P1bThen passes through the outlet valve V of the first basic chromatography device P1cAnd (4) discharging waste liquid.
It should be understood that the various modes of operation described above may be combined and permuted as desired.
It should be understood that when a large number of columns are not required in practical use, for example, the purified substance is relatively single, the two-channel three-column combined chromatography apparatus of the present embodiment can switch the flow channels of the apparatus by controlling the communication paths of the respective four-way valves, so that the apparatus is changed into various combinations, such as one basic chromatography device connected to one column, two basic chromatography devices connected to two columns, and the like, thereby flexibly adopting various combinations according to various requirements.
Third embodiment
Referring to FIG. 7, the third embodiment is a modification of the first embodiment, to which a third basic chromatography device P3, a third chromatography column C3 and a third three-way valve V are added3The three-flow-path three-chromatography-column combined chromatography device is formed. The device can adopt three flow paths to carry out continuous flow chromatography due to the adoption of three basic chromatography devices.
First three-way valve V1Are respectively defined as
And
second three-way valve V2Are respectively defined as
And
third three-way valve V3Are respectively defined as
And
as shown in FIG. 7, the first three-way valve V1Is/are as follows
A column valve V connected to the upper part of the first basic chromatography device P1aThe connection is carried out by connecting the two parts,
the port was connected to the inlet of the first chromatographic column C1,
outlet valve V of port and third basic chromatography device P3cConnecting; second three-way valve V2Is/are as follows
A column position valve V connected with the upper part of the second basic chromatographic equipment P2aThe connection is carried out by connecting the two parts,
the port is connected to the inlet of a second chromatography column C2,
oral and primary basis chromatographyOutlet valve V of plant P1cConnecting; third three-way valve V3Is/are as follows
The port and the upper column valve V of the third basic chromatography device P3aThe connection is carried out by connecting the two parts,
the port is connected to the inlet of a third chromatographic column C3,
outlet valve V of port and base chromatographic device P2cAnd (4) connecting. And the outlet of the first chromatographic column C1 is connected to the lower column position valve V of the first basic chromatographic apparatus P1bConnected, the outlet of the second chromatography column C2 is connected with the lower column position valve V of the second basic chromatography device P2bConnected to the outlet of the third column C3 and a lower column valve V of a third basic chromatography device P3bAnd (4) connecting.
In the present embodiment, three-way valves (V) are provided1、V2、V3) Three basic chromatographic devices (P1, P2 and P3) and three chromatographic columns (C1, C2 and C3) are connected to form a three-flow-path three-chromatographic-column combined chromatographic device. The device has a combined continuous flow chromatography running mode of continuous flow sample loading, sample loading of one chromatographic column and sample loading pretreatment and sample loading post-treatment of other chromatographic columns, and can realize an independent single-column chromatography running mode.
The operation of the three-flow-path three-column chromatography apparatus will be described.
(i) Continuous flow sample loading
For the combined chromatography apparatus having three basic chromatography devices (P1, P2, P3), one of the basic chromatography devices may optionally control one chromatography column or a plurality of chromatography columns to load.
For example,
(1) the first basic chromatographic equipment P1 controls the loading of chromatographic columns C1, C2 and C3,
the first basic chromatographic device P1 controls the sample feeding, and the second basic chromatographic device P2 controls the sample feedingAnd the third basic chromatography device P3 opens the flow path. The preparation passes through the upper column valve V of the first basic chromatographic device P1aFlows through the first three-way valve V in sequence1A first chromatographic column C1, a column position valve V at the lower part of the first basic chromatographic apparatus P1bAnd an outlet valve VcA second three-way valve V2A second chromatographic column C2, a column position valve V at the lower part of the second basic chromatographic device P2bAnd an outlet valve VcAnd a third three-way valve V3A third chromatographic column C3 and a lower column position valve V of a third basic chromatographic apparatus P3bAnd an outlet valve VcAnd discharging waste liquid.
It should be understood that the loading of three columns (C1, C2, C3) can also be controlled using the second basic chromatography device P2 or the third basic chromatography device P3.
As another example of the present invention,
(2) loading two chromatography columns with either basic chromatography apparatus, for example C1 and C2 with the first basic chromatography apparatus P1,
the first basic chromatography device P1 controls the sample loading, the second basic chromatography device P2 opens the flow path, and the preparation is passed through the upper column valve V of the first basic chromatography device P1aFlows through the first three-way valve V in sequence1A first chromatographic column C1, a column position valve V at the lower part of the first basic chromatographic apparatus P1bAnd an outlet valve VcA second three-way valve V2A second chromatography column C2 and a lower column position valve V of a second basic chromatography device P2bAnd an outlet valve VcAnd discharging waste liquid.
Similarly, the second basic chromatography device P2 can be used for controlling the loading of the chromatographic columns C2 and C3, or the third basic chromatography device P3 can be used for controlling the loading of the chromatographic columns C3 and C1.
As another example of the present invention,
(3) when applied to simpler single component purification operations, a single column is controlled to load using a single basic chromatographic apparatus, for example,
the second basic chromatography device P2 is used to control the loading of the second chromatography column C2, the second basic chromatography device P2 is used to control the loading, and the preparation is fed from the upper column position valve V of the second basic chromatography device P2aFlows through the second three-way valve V in sequence2A second chromatography column C2 and a lower column position valve V of a second basic chromatography device P2bAnd an outlet valve VcAnd discharging waste liquid.
(ii) Washing and eluting one chromatographic column, and loading the other two chromatographic columns
Because each three-way valve is connected with two basic chromatographic devices and one chromatographic column, any basic chromatographic device can be selected to control one chromatographic column to be washed and eluted, and the other basic chromatographic device can be selected to control the rest two chromatographic columns to be loaded.
For example,
(1) eluting with C1, loading onto C2 and C3,
the outlet valve V of the first basic chromatography device P1 is openedcSwitching the flow path to the waste liquid and switching the outlet valve V according to the elution parameters at the right timecTo the sample collection port, thereby completing the elution of the first chromatographic column C1 and obtaining the eluted product. The second basic chromatography device P2 is used to control the loading of the chromatographic columns C2 and C3, the third basic chromatography device P3 opens the flow path, and the preparation passes through the upper column position valve V of the second basic chromatography device P2aFlows through the second three-way valve V in sequence2A second chromatographic column C2, a column position valve V at the lower part of the second basic chromatographic device P2bAnd an outlet valve VcAnd a third three-way valve V3A third chromatographic column C3 and a lower column valve V of a basic chromatographic apparatus P3bAnd an outlet valve VcAnd discharging waste liquid.
As another example of the present invention,
(2) eluting with C2, loading onto C3 and C1,
the outlet valve V of the second basic chromatography device P2 is openedcSwitching the flow path to the waste liquid and switching the outlet valve V according to the elution parameters at the right timecTo the sample collection port, thereby completing the elution of the second chromatography column C2 and obtaining the eluted product. The third basic chromatography device P3 is used to control the loading of the chromatographic columns C3 and C1, the first basic chromatography device P1 opens the flow path, and the preparation passes through the upper column position valve V of the third basic chromatography device P3aFlows through the third three-way valve V in sequence3Third column C3A lower column valve V of the third basic chromatography device P3bAnd an outlet valve VcA first three-way valve V1A first chromatographic column C1 and a lower column position valve V of a first basic chromatographic apparatus P1bAnd an outlet valve VcAnd discharging waste liquid.
As another example of the present invention,
(3) eluting with C3, loading onto C1 and C2,
the outlet valve V of the third basic chromatography device P3cSwitching the flow path to the waste liquid and switching the outlet valve V according to the elution parameters at the right timecTo the sample collection port, thereby completing the elution of the third chromatographic column C3 and obtaining the eluted product. The first basic chromatography device P1 is used to control the loading of the chromatographic columns C1 and C2, the second basic chromatography device P2 is used to open the flow path, and the preparation passes through the upper column position valve V of the first basic chromatography device P1aFlows through the first three-way valve V in sequence1A first chromatographic column C1, a column position valve V at the lower part of the first basic chromatographic apparatus P1bAnd an outlet valve VcA second three-way valve V2A second chromatography column C2 and a lower column position valve V of a second basic chromatography device P2bAnd an outlet valve VcAnd discharging waste liquid.
It should be understood that the various modes of operation described above may be combined and permuted as desired. It should be understood that, according to the combined chromatography apparatus with three flow paths and three chromatography columns of the present embodiment, according to the actual requirements of washing elution and loading, the collection can be obtained by washing elution on two chromatography columns respectively after loading the continuous flow, and the loading on the other chromatography column can be continued. The technicians in this field can flexibly configure different combination modes according to the conditions and requirements of the actual purification operation of the biological preparation to meet the requirements of various conditions.
Fourth embodiment
Based on the concept of flexibly combining a plurality of basic chromatography apparatuses and a plurality of chromatography columns of the present invention, the basic chromatography apparatuses or chromatography columns can be expanded to a larger number. For example, two basic chromatography devices and five chromatography columns are selected, and the inlet and the outlet of each chromatography column are connected to the two basic chromatography devices, so that the condition that any one basic chromatography device can be used for washing and eluting any one chromatography column is met; meanwhile, in order to realize continuous flow sample loading of five chromatographic columns, the connection mode between the inlet and the outlet of three chromatographic columns in the second embodiment can be referred, the outlet of the previous chromatographic column is connected with the inlet of the next chromatographic column, and the chromatographic columns are connected in series in sequence, so that the sample loading of a plurality of chromatographic columns can be completed through the control of one basic chromatographic device.
The inlets of the five chromatographic columns are respectively connected to the column position valves at the upper parts of the two basic chromatographic devices, and the outlets of the five chromatographic columns are respectively connected to the column position valves at the lower parts of the two basic chromatographic devices; and the inlet and the outlet of each chromatographic column respectively need one four-way valve, namely ten four-way valves are needed. Thereby forming the combined chromatographic device of the two-flow path five-chromatographic column. For the operation of the device, reference is made to the operation of the two-flow three-column chromatography device of the second embodiment.
The above-described embodiments of the invention may achieve one or more of the following advantages:
(i) the invention keeps the function of the original chromatography equipment, and the continuous flow chromatography operation can be realized by using a single chromatography column or a plurality of chromatography columns.
(ii) The device can realize independent washing and elution of a plurality of chromatographic columns and sample loading of other chromatographic columns simultaneously by controlling the communication mode of the multi-way valve, thereby improving the utilization rate of equipment.
(iii) Experiments have shown that samples prepared using the apparatus and method of the present invention (e.g. purified preparations) do not differ significantly from samples prepared using conventional single column apparatus in terms of quality (e.g. purity and impurities) and process performance (e.g. yield).
(iv) Since the apparatus is a combination of existing equipment, it is not limited to laboratory scale, but is also suitable for pilot and production scale.
It should be understood that the above embodiments are only exemplary and are not intended to limit the present invention. Various modifications and alterations of the above-described embodiments may be made by those skilled in the art in light of the teachings of the present invention without departing from the scope thereof.
For example,
(i) in order to prevent the return and diffusion of the preparation in each flow path of the apparatus, a check valve may be added to the flow path of any of the multi-way valves of the present invention.
(ii) The present invention is applicable to, but not limited to, the purification of biological agents, including, inter alia, antibodies, fusion proteins, and cytokines, and unless otherwise specified, the term "antibody" includes reference to glycosylated and non-glycosylated immunoglobulins of any isotype or subclass or antigen binding regions thereof that compete for specific binding with intact antibodies, including human antibodies, humanized antibodies, chimeric antibodies, multispecific antibodies, monoclonal antibodies, polyclonal antibodies, and oligomers or antigen binding fragments thereof. Also included are proteins having antigen-binding fragments or regions, such as Fab, Fab ', F (ab')2, Fv, diabodies, Fd, dAb, macroantibodies (maxibody), single chain antibody molecules, Complementarity Determining Region (CDR) fragments, scFv, diabodies, triabodies, tetrabodies, and polypeptides that comprise at least a portion of an immunoglobulin sufficient to confer specific antigen-binding to a polypeptide of interest. Specific examples of known antibodies produced using the methods of the invention include, but are not limited to, certolizumab (sintilmab), adalimumab (adalimumab), bevacizumab (bevacizumab), Aflibercept (Aflibercept), infliximab (infliximab), nimotuzumab (nimotuzumab), pertuzumab (pertuzumab), ranibizumab (ranibizumab), rituximab (rituximab), trastuzumab (trastuzumab).
Claims (15)
1. A combined chromatography apparatus comprising a plurality of basic chromatography devices for controlling the flow of fluid into or out of a chromatography column, a plurality of chromatography columns and a plurality of multi-way valves, wherein the basic chromatography devices comprise a column-level valve and an outlet valve,
the inlet of each chromatographic column is connected with at least two of the basic chromatographic devices through the multi-way valve, the outlet of each chromatographic column is connected with at least one basic chromatographic device, and the combined chromatographic device can realize continuous flow chromatographic operation of multiple flow paths.
2. The modular chromatography apparatus of claim 1, wherein the outlet of each chromatography column is connected to the inlet of another chromatography column via the multi-way valve.
3. The combined chromatography device of claim 1, wherein the number of the plurality of basic chromatography apparatuses is two or three, the number of the plurality of chromatography columns is two, three or more, and the multi-way valve is a three-way valve, a four-way valve, a five-way valve or a six-way valve.
4. The combined chromatographic device according to claim 1, wherein the basic chromatographic apparatus comprises two column position valves, the inlet of the chromatographic column is connected with one column position valve or the outlet valve of the basic chromatographic apparatus through the multi-way valve, and the outlet of the chromatographic column is connected with the other column position valve of the basic chromatographic apparatus.
5. The combined chromatographic device according to claim 4, wherein the number of the basic chromatographic apparatuses is the same as that of the chromatographic columns, and is N, N is a positive integer greater than or equal to 2, the inlet of the first chromatographic column is connected with one of the column position valves of the first basic chromatographic apparatus and the outlet valve of the Nth basic chromatographic apparatus through a multi-way valve, the outlet of the first chromatographic column is connected with the other of the column position valves of the first basic chromatographic apparatus, the inlet of the second chromatographic column is connected with one of the column position valves of the second basic chromatographic apparatus and the outlet valve of the first basic chromatographic apparatus through a multi-way valve, the outlet of the second chromatographic column is connected with the other of the column position valves of the second basic chromatographic apparatus, and so on, the inlet of the Nth chromatographic column is connected with one of the column position valves of the Nth basic chromatographic apparatus and the outlet valve of the N-1 th basic chromatographic apparatus through a multi-way valve, the outlet of the Nth chromatographic column is connected with the other column position valve of the Nth basic chromatographic equipment.
6. The combined chromatographic device according to claim 4, wherein the number of the chromatographic columns is larger than that of the basic chromatographic equipment, the inlet and the outlet of each chromatographic column are connected with the column position valves of at least two basic chromatographic devices through the multi-way valves, and the inlet of each chromatographic column is also connected with the outlet of another chromatographic column through the multi-way valves.
7. The combined chromatographic apparatus according to claim 4, wherein the number of the basic chromatographic devices and the number of the chromatographic columns are both two, the inlet of the first chromatographic column is connected with one of the column-level valves of the first basic chromatographic device and the outlet valve of the second basic chromatographic device through a first three-way valve, and the outlet of the first chromatographic column is connected with the other column-level valve of the first basic chromatographic device; the inlet of the second chromatographic column is connected with one column position valve of the second basic chromatographic device and the outlet valve of the first basic chromatographic device through a second three-way valve, and the outlet of the second chromatographic column is connected with the other column position valve of the second basic chromatographic device.
8. The combined chromatographic apparatus according to claim 4, wherein the number of the basic chromatographic devices and the number of the chromatographic columns are three, the inlet of the first chromatographic column is connected with one of the column-level valves of the first basic chromatographic device and the outlet valve of the third basic chromatographic device by a first three-way valve, and the outlet of the first chromatographic column is connected with the other column-level valve of the first basic chromatographic device; the inlet of the second chromatographic column is connected with one column position valve of the second basic chromatographic equipment and the outlet valve of the first basic chromatographic equipment through a second three-way valve, and the outlet of the second chromatographic column is connected with the other column position valve of the second basic chromatographic equipment; the inlet of the third chromatographic column is connected with one column position valve of the third basic chromatographic device and the outlet valve of the second basic chromatographic device through a third three-way valve, and the outlet of the third chromatographic column is connected with the other column position valve of the third basic chromatographic device.
9. The combined chromatographic apparatus according to claim 4, wherein the number of the chromatographic columns is three, the number of the basic chromatographic devices is two, two four-way valves are connected to the two column position valves of each of the two basic chromatographic devices, one four-way valve is connected to the inlet and outlet of each of the three chromatographic columns, the inlet of each chromatographic column is connected to one of the column position valves of the two basic chromatographic devices and the outlet of the other chromatographic column through one four-way valve, and the outlet of each chromatographic column is connected to the other of the column position valves of the two basic chromatographic devices and the inlet of the other chromatographic column through one four-way valve.
10. The combined chromatography device according to any one of claims 1 to 9, wherein any one of the branch flow paths of the multi-way valve is provided with a one-way valve.
11. A method of continuous flow chromatography of a fluid using a combined chromatography device according to any of claims 1 to 10, comprising switching the passages of the multi-port valve, optionally performing single or multi-layer chromatography column continuous flow chromatography.
12. The continuous flow chromatography method of claim 11, wherein the method comprises,
and switching the multi-way valve connected with the inlet of one of the plurality of chromatographic columns to a mode connected with the column position valve, and switching the multi-way valve connected with the outlet of the chromatographic column to a mode connected with the multi-way valve connected with the inlet of the other chromatographic column, thereby continuously loading at least two chromatographic columns.
13. The continuous flow chromatography method of claim 11, wherein the method comprises,
and switching the multi-way valve connected with the inlet of one of the plurality of chromatographic columns to a mode connected with the column position valve and switching the multi-way valve connected with the outlet of the chromatographic column to a mode connected with the column position valve, thereby washing and eluting the chromatographic column.
14. A method of purifying a biological agent, the method comprising: use of a combined chromatography device according to any of claims 1 to 10 for continuous flow chromatography of a biological agent to obtain a purified product.
15. The purification method of claim 14, wherein the biological agent is an antibody preparation; further, the antibody is any one of the group consisting of certolizumab, adalimumab, bevacizumab, apraxizumab, infliximab, nimotuzumab, pertuzumab, ranibizumab, rituximab, and trastuzumab.
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| CN201811228757 | 2018-10-22 | ||
| CN2018112287570 | 2018-10-22 |
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Cited By (2)
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| CN112451996A (en) * | 2020-11-10 | 2021-03-09 | 浙江大学 | Optimization method for capturing protein by multi-column continuous flow chromatography |
| CN113341014A (en) * | 2021-06-01 | 2021-09-03 | 上海药明生物技术有限公司 | Continuous flow chromatography apparatus and method |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112451996A (en) * | 2020-11-10 | 2021-03-09 | 浙江大学 | Optimization method for capturing protein by multi-column continuous flow chromatography |
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| WO2022100325A1 (en) * | 2020-11-10 | 2022-05-19 | 浙江大学 | Optimization method for capturing protein by multi-column continuous flow chromatography |
| US11958881B2 (en) | 2020-11-10 | 2024-04-16 | Zhejiang University | Optimization method for capturing proteins by multi-column continuous chromatography (MCC) |
| CN113341014A (en) * | 2021-06-01 | 2021-09-03 | 上海药明生物技术有限公司 | Continuous flow chromatography apparatus and method |
| CN113341014B (en) * | 2021-06-01 | 2024-01-09 | 上海药明生物技术有限公司 | Continuous flow chromatography apparatus and method |
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