阅读说明：本技术 色谱系统 (Chromatography system ) 是由 J·威德哈马尔 K·海肯伯格 O·格雷尔松 L·马特松 K·卡尔松 于 2020-04-16 设计创作，主要内容包括：所公开的是一种色谱系统,其布置成执行对于特定柱的自动色谱柱效率测试,以确定诸如h和As之类的效率参数,并且可选地进一步向用户提供关于柱效率的指南,所述系统具有控制器,所述控制器布置成自动访问数据,以实现柱压缩板高度的计算,由此以便计算柱的效率,所述数据从特定柱以及将被用于那个柱中的树脂的至少存储参数来得出。(Disclosed is a chromatography system arranged to perform automatic chromatography column efficiency tests for a particular column to determine efficiency parameters such As h and As, and optionally further to provide guidance to a user regarding column efficiency, the system having a controller arranged to automatically access data derived from at least stored parameters of the particular column and the resin to be used in that column to effect calculation of column compression plate height, thereby to calculate the efficiency of the column.)

1. A chromatography system arranged to perform automatic chromatography column efficiency tests for a particular column to determine efficiency parameters such As h and As, and optionally further to provide a user with guidelines on column efficiency, the system having a controller arranged to automatically access data derived from the particular column and at least stored parameters of the resin to be used in that column to effect calculation of column compression plate height, thereby to calculate the efficiency of the column.

2. The chromatography system of claim 1, wherein said system determines whether said particular column is suitable for use or unsuitable for use.

3. The chromatography system of claim 2, wherein if the column is determined to be unsuitable, a suitable column repacking procedure is initiated.

4. The chromatography system of any preceding claim, further comprising a user interface, and wherein the chromatography system is arranged to automatically determine one or more efficiency parameters and to present a relevant action on the user interface based on the result, or alternatively the system may be configured to automatically initiate some or all of a relevant action based on the efficiency parameter.

5. The chromatography system of claim 1, wherein the system is configured to automatically or semi-automatically initiate one or more of the following actions in order to improve column efficiency:

standard column cleaning protocol (e.g. NaOH 0.5M) when 3 < h < 3.5 (grade 1)

Strong column cleaning protocol (e.g. NaOH 1M, acid and/or isopropanol) when 3.5 < h < 5 (grade 2)

Column repacking protocol when 5 < h < 7 (stage 3)

H < 7 (4 th stage) cancels the process and instructs the user to replace the resin.

6. The chromatography system of claim 5, wherein, once one or more of said actions are completed, said system is configured to further automatically determine one or more efficiency parameters and to test for any improved efficiency.

7. The chromatography system of claim 5 or 6, wherein the system is configured to repeat one or more of the above actions two or more times when an efficiency parameter is not reached, or at a higher level after a predetermined attempt at a lower level.

8. The chromatography system of any preceding claim, wherein said automated column check further comprises determining said column delta pressure to check said column packing performance.

9. The chromatography system of claim 8, wherein if the determined delta pressure is determined to be too low, a report is made to a user that there is an increased risk for gap build, and if the delta pressure is determined to be too high, a report is made to a user that the pressure differential may be outside the system pressure range, wherein a relevant range for delta pressure is optionally retrievable from the column database and/or the system settings.

10. A method for use in a chromatography system arranged to perform automatic chromatography column efficiency tests for a specific column to determine efficiency parameters such As h and As and optionally further provide guidance to a user regarding column efficiency, the method comprising the steps of:

a) providing a system controller

b) Configuring the controller to automatically access data derived from the particular column and at least stored parameters for resin in that column to enable the controller to calculate a column compression plate height, thereby to calculate the efficiency of the column, and optionally to perform any one or more of the additional steps of:

c) configuring the controller to determine whether the particular column is suitable for use or unsuitable for use;

d) performing a repacking procedure if the column is determined to be unsuitable;

e) further providing a user interface and configuring the system to automatically determine one or more efficiency parameters and present a recommendation-related action at the user interface based on the determination;

f) configuring the system to automatically or semi-automatically initiate one or more of the following actions in order to improve column efficiency:

conventional column cleaning protocol (e.g. NaOH 0.5M) when 3 < h < 3.5 (stage 1);

a strong column cleaning protocol (e.g. NaOH 1M, acid and/or isopropanol) when 3.5 < h < 5 (grade 2);

column repacking procedure when 5 < h < 7 (stage 3);

cancel the process when h < 7 (stage 4) and instruct to replace the column resin.

Technical Field

The present invention relates to a liquid chromatography system configured to operate with at least one column when feeding a sample to the liquid chromatography system for purifying the sample comprising a target product. In particular, the present invention relates to a method for preparing and identifying packed columns and column efficiency tests.

Background

An important factor in process chromatography is the binding capacity of the chromatography column for the solute (solute). The binding capacity directly affects the productivity and cost of the chromatography step. Binding capacity is defined in terms of dynamic/through-flow capacity or as maximum binding capacity. The dynamic capacity depends on the conditions of the solution flowing through the column filled with chromatography medium and can be expressed as the ratio between the column volume and the feed flow rate, the so-called residence time. If the residence time is infinitely long, the maximum binding capacity represents the through-flow capacity of the column.

When validating a process with a single or several chromatography columns used in a chromatography system, it is necessary that the results from the process are predictable and can be repeated without deviating from the process specifications. For single column systems, when changing columns, it is necessary that the properties of the new column are of the same type and operate in the same way, i.e. columns with the same or almost the same properties are required to achieve the desired result in the validation process.

In continuous chromatography, several columns are connected in an arrangement that allows the columns to be operated in series and/or in parallel, depending on the process requirements. Thus, all columns can be run substantially simultaneously but with slightly modified process steps. This procedure can be repeated so that each column is loaded, eluted and regenerated several times in the process. In contrast to 'conventional' chromatography, where a single chromatography cycle is based on several sequential steps, e.g. sample loading, washing, elution, stripping, Cleaning In Place (CIP) and re-equilibration, in continuous chromatography based on multiple columns, all these steps are performed simultaneously, but each on a different column.

For best results, the columns in a continuous chromatograph must be identical or nearly identical. The same applies when the column is replaced with a single chromatographic column used in the validation process. If the column performance variation is too large, the process will operate outside the verified performance range.

Continuous chromatography is an example of a periodic counter-current process, as all of the chromatography columns making up the system periodically move simultaneously in the opposite direction to the sample flow. The apparent movement of the column is achieved by appropriate reorientation of the inlet stream (let stream) and the outlet stream (outlet stream) to and from the column.

Historically, the basic factors for a reliable continuous process were:

1) the quality of the columns used, and more particularly the similarity or even identity (identity) between the columns,

2) a constant feed composition (feed composition), and

3) hardware dependence, e.g. constant flow rate delivered by pump、Valve functionality, etc.

If the columns are not identical, the theoretical calculations typically used to design a continuous chromatographic process will not be correct and it will become difficult to design an efficient and robust continuous chromatographic process. The same argument applies if the feed concentration and flow rate change over time in an unexpected manner.

Therefore, for scale-up considerations, a reliable pump with the same column in the system is necessary. However, packing the column with chromatographic media is complicated in order to obtain reproducible results. Even small differences in the number of plates or other filling properties can have a large impact on the end result. Furthermore, since the capacity of chromatography resins typically changes during the life/use of the resin, the process conditions selected for a new resin/media may not be applicable to a resin that has been used several times. An efficient continuous chromatography process that will always operate at its optimum will be even more complicated to design if the feed solution concentration will also change.

Disclosure of Invention

It is an object of the present disclosure to provide methods and apparatus configured to perform methods and computer programs that seek to mitigate, alleviate or eliminate one or more of the above-identified deficiencies in the art and disadvantages singly or in any combination.

This object is achieved by a liquid chromatography system configured to operate with at least one column and configured for purifying a sample comprising a target product using a predefined process. The liquid chromatography system includes a controller configured to:

controlling the operation of the chromatography system to run a predefined process,

-retrieving column data accessible from a data storage, the column data being specific to each column, an

Adapting at least one process parameter of a predefined process for each column based on the column data,

whereby a predefined process is adapted to each column to obtain the target product and maintain the performance of the liquid chromatography system.

An advantage is that more stable and reproducible results are obtained from the liquid chromatography process, since the process parameters can be adapted to compensate for production variations when producing each column.

Additional objects and advantages will be apparent to those skilled in the art from the detailed description.

Drawings

Figure 1 illustrates an overview of a bioprocess purification system designed to purify a product of interest using liquid chromatography.

Figure 2 illustrates a continuous chromatography with any number of columns based on simulated moving bed technology.

Figures 3a-3c illustrate the principle of three-column chromatography.

Figure 4 illustrates a single column liquid chromatography system.

FIG. 5 illustrates a simplified flow diagram for the liquid chromatography system of FIG. 4 having a pressure sensor.

FIG. 6 illustrates a simplified flow diagram for the liquid chromatography system of FIG. 4 having only a system pressure sensor.

FIG. 7 illustrates a simplified flow diagram for the liquid chromatography system of FIG. 4 having a virtual pressure sensor.

Fig. 8 illustrates the concept of assigning columns to types.

FIG. 9 illustrates data flow in a system suitable for fabricating columns and liquid chromatography systems.

FIG. 10 is a flow chart illustrating a method for manufacturing a column for a liquid chromatography system.

FIG. 11 is a flow chart illustrating a method for controlling a liquid chromatography system configured to operate with at least one column.

Detailed Description

The chromatography system is designed to purify a product of interest (e.g. protein, biomolecule from cell culture/fermentation, natural extract) using at least one packed column of resin to create a purification step. Each column is switched between loading and non-loading steps (e.g., washing and elution).

In fig. 1, an overview of a biological process purification system 10 configured to purify a target product using a separation process is shown. The bioprocess purification system comprises a number of steps related to cell culture 11, maintenance 12, capture 13, virus inactivation 14, purification 15 and delivery 16.

The cell culture step 11 may be a perfusion type culture comprising a continuous addition of nutrients for cell growth in the perfusion culture and a continuous removal of products and waste by drainage and filtration, for example using an Alternating Tangential Filtration (ATF) filter arrangement. This step may include process control for Viable Cell Density (VCD), and the next step in the process is initiated when the VCD reaches a predetermined value. VCD can be controlled by adapting the composition of the cell culture medium fed to the culture or by adding certain components directly to the culture. Alternatively, the cell culture is of a batch type.

The sample containing the target product is utilized in a cell-free extraction process, for example, by filtration, centrifugation, or another technique.

The holding step 12 is an optional step depending on the process requirements, for example if the filter is online before the capturing step 13. This step may comprise a process control of the weight and the next step in the process is started when a predetermined volume value is reached or alternatively after a certain time period or when a predetermined mass is reached. The holding step may be used to collect a volume of filtered feed from a perfusion cell culture or from a batch culture.

The capturing step 13 comprises at least one chromatography column which may bring the filter online prior to the capturing step. The capturing step 13 may comprise a continuous chromatography setup, which may for example be run as a periodic counter current chromatography, as illustrated in fig. 2, with a continuous feed of the sample comprising the target product from the cell culturing step 11, either directly or via the holding step 12. The capture step includes one or more batch elutions, and process control using an inline UV sensor manipulates changes in feed concentration and resin volume. The next step starts when a predetermined amount (e.g. volume, mass or time) is reached.

In the virus inactivation step 14, different options for virus inactivation are available depending on the process requirements. One option is to use a batch mode with a low pH in the hold up tank (hold up tank) for 30-60 minutes. This step may include process control of volume, time, temperature and pH. The next step starts when a predetermined time is reached.

The refining step 15 may be a straight-through process (STP) with a connected batch step or a continuous chromatography with a continuous loading step or a combination thereof. The flow rate is adjusted to the perfusion rate required by the producer cells, which means that the flow rate is determined by the previous step. This step may include process control for UV, flow and volume and the next step starts when a predetermined volume and volume is reached, alternatively when a time-out is reached.

The delivery step 16 may include a virus removal step, such as a virus filter, prior to the ultrafiltration step. The delivery step may be used as a concentration step for batch addition of sample from the refining step. The step of delivering may include continuous or batch delivery of the product, and may include continuous or batch removal of waste. This step may include process control of pH, conductivity, absorbance, volume and pressure, and delivery is achieved when a predetermined product concentration in a predefined environment is reached.

The automation layer 17 is used to handle decision points for the next step in the process. Different types of sensors (not shown), both online and offline, are integrated into the process flow to monitor different parameters that may be used to provide the automation layer 17 with data that may be used to manipulate decision points. Sensors include, but are not limited to, measuring only flow, VCD, weight, pressure, UV, volume, pH, conductivity, absorbance, and the like.

It should be noted that UV absorption is an example of a parameter that can be monitored to detect the composition of the purified sample. However, other parameters operating in other frequency ranges (e.g., IR, fluorescence, x-ray, etc.) may be used.

The capturing step 13 may comprise a continuous chromatography setup 20 (as illustrated in fig. 2) or a single column 240 (as illustrated in fig. 4). Continuous chromatography supports process intensification by reducing the footprint and improving productivity. In addition, continuous chromatography is particularly suitable for the purification of labile molecules, as short process times help ensure stability of the target product.

In fig. 2, a sample containing a target product is fed into the continuous chromatography 20 via an inlet 21, and eluting the target product is available at an outlet 22. The continuous chromatography 20 comprises a plurality of columns A, B, N, and each column is provided with a column inlet 23 and a column outlet 24. The column inlet 23 and column outlet 24 of each column are connected to a valve system 25, which valve system 25 is configured to cyclically connect the column to the inlet 21 and outlet 22 to achieve continuous purification of the target product. An example of a system configuration with three columns is described in connection with fig. 3a-3 c.

The continuous chromatography 20 is further provided with a buffer inlet 26 and a waste outlet 27 in order to be able to perform the required operations. The inline sensor 28 may be provided after the column outlet 24 of each column or assigned to the process flow and integrated into the valve system 25. Important parameters (e.g., UV) are measured to control the process, as described below. Another in-line sensor 28' may be provided before the column inlet 23 of each column to enable direct assessment of the performance of each column. An inline inlet sensor 26 may also be provided to monitor the composition of the sample fed into the continuous chromatograph 20.

The continuous chromatography may also include an offline sensor 29, which offline sensor 29 is designed to extract material from the process and thereafter evaluate selected parameters before the material is disposed of as waste.

The continuous chromatography comprises at least two (e.g. at least three) columns and the operating principle in a three column (3C) setup is described in connection with fig. 3 a-3C. The 3C setup features two parallel streams (feature): one for loading of two columns in the loading zone and one for non-loading steps, such as elution and regeneration of the third column.

In fig. 3a, which illustrates step 1, columns a and B are in the loading zone. Column a can be overloaded without sample loss because column B captures the flow through from column a. In this way, the utilization of the resin binding capacity is maximized.

In fig. 3B, which illustrates step 2, the overload column a is switched and column B becomes the first column and column C becomes the second column in the loading zone. The overloaded column a will now be subjected to non-loading steps, such as elution and regeneration in a parallel workflow.

In fig. 3c, which illustrates step 3, the overload column B in the loading zone is switched. At this time, column C becomes the first column, and column a becomes the second column in the loading zone, while column B is subjected to elution and regeneration in a parallel workflow. These three steps are repeated in a cyclic fashion until the desired target product volume, mass or amount is reached (or until the resin life is reached and the column needs to be refilled or exchanged).

The continuous chromatography setup illustrated in fig. 2 may utilize more than three columns, and in a four-column (4C) setup, the same principles apply. However, the non-loading step may become limited in the 3C setup, and the non-loading step can be split over two columns and run in parallel with the third flow path in the 4C setup. The 4C setup allows for balanced loading and unloading steps. More columns will lead to a more flexible system, while the complexity of the valve system 25 becomes more and more complex. However, some continuous chromatographs have sixteen or more columns.

Fig. 4 schematically illustrates one embodiment of a chromatography system 190, the chromatography system 190 comprising two 3-way input valves 160 and 161 arranged to select input fluids from the fluid sources a1, a2, B1, B2 for the two system pumps 150 and 151. The chromatography system 190 may further comprise:

a pressure sensor 200 for recording the system pressure in the flow path after the system pump, an

Mixer 210, which is used to ensure proper mixing of the fluid supplied by the pump.

These correspond to cell culture block 11 illustrated in fig. 1, as indicated by dashed line 110.

The system further comprises:

an injection valve 220 for injecting a sample into the fluid path,

a column connection valve 230 for selectively connecting/disconnecting a column 240 in the fluid path.

Pre-column (pre-column) pressure sensor 235 and post-column pressure sensor 236

An Ultraviolet (UV) monitor 250 for detecting output from the column.

A conductivity monitor 260, and

a pH monitor 265.

These correspond to the capture blocks 13 illustrated in fig. 1, as indicated by dashed lines 130.

The system further comprises:

an output selector valve 270 having two or more output positions, e.g., connected to a fraction collector 280, a waste receptacle, etc., corresponding to the delivery block 16 in fig. 1, as indicated by dashed line 160, and

a system controller 300 connected to the pump and valves for controlling the flow of liquid through the system and to the sensors and monitors for monitoring the flow, the connection being illustrated by dotted line 310, which corresponds to automation block 17 in fig. 1, as indicated by dashed line 170.

The chromatography system of fig. 4 represents a general example of how a single column chromatography system may be designed, as well as other embodiments may have a different design of two or more components including some components, with some components potentially lacking. For example, components corresponding to retention 12, virus inactivation 14, and purification 15 as illustrated in fig. 1.

Fig. 5 is a simplified flow diagram for a liquid chromatography system 190 according to fig. 4. In fig. 5, the flow path has been straightened out and some components have been removed to achieve a simpler view. In fig. 5, the system controller is shown connected only to pump 150, pressure sensor 200, pre-column pressure sensor 235, and post-column pressure sensor 236, but it may be connected to other components as described above. In fig. 5, the system includes both a pre-column pressure sensor 235 and a post-column pressure sensor 236, whereby the column pressure is measured directly by the pre-column sensor 235 and the incremental column pressure is measured by subtracting the pressure recorded by the post-column sensor 236 from the column pressure.

As outlined above, some systems do not have a pressure sensor other than system pressure sensor 200. Fig. 6 is a simplified flow diagram of such a liquid chromatography system 190 having a single pressure sensor 200 for recording the system pressure. As mentioned above, pressure control in such systems relies solely on the recorded system pressure from sensor 200. Fig. 7 is a simplified flow diagram of a liquid chromatography system according to an embodiment of the invention, wherein the controller 300 is arranged to estimate the pre-column pressure based on the recorded system pressure, the characteristics of the flow path, and the viscosity and flow rate of the liquid in the system. The estimated pre-column pressure may be referred to as a "virtual pressure sensor" schematically illustrated in FIG. 7 by a light dashed line.

According to one embodiment, the calculation of the virtual pressure signal may be based on the bernoulli equation for the pressure drop in the flow channel.

Flow passageWherein

L = length [ mm ]

D = diameter [ mm ]

Q = flow rate [ ml/min ]

V = viscosity [ cP ]

By providing the system controller with the length and diameter of the flow path and the viscosity of the liquid in the system, it can be arranged to calculate the pressure drop caused by the flow path up to the column at the current flow rate. In some systems, the length and size of the flow path between the system pressure sensor 200 and the column 240 may be normalized so that predefined parameters may be used for the calculation. In other systems, which is the most common situation, the flow paths between components in a chromatography system are user-defined, whereby a user of the system has to enter parameters through a user interface.

According to one embodiment, the main part of the flow path between the system pressure sensor 200 and the column 240 may be composed of capillary tubing of the same diameter, and then the flow path characteristics may be estimated as the total length of the tubing, thus excluding contributions from other components (e.g. valves, etc.) from the calculation. In other embodiments, contributions from valves, etc. in the flow path are considered and may be system defined, while the piping, etc. is user defined. It should be noted that where the flow path includes sections of different sizes (e.g., different inner diameter pipes), the pressure drop over each section must be calculated separately and eventually summed together to provide the total pressure drop.

When the pressure drop in the flow path up to the column 240 is estimated by the above calculation, the virtual pre-column pressure is calculated by subtracting the pressure drop from the system pressure recorded by the system pressure sensor 200.

Example (c):

if the system pressure is 5 bar and the calculated pressure drop over the flow path is 2 bar, the calculated virtual pre-column pressure is estimated to be 3 bar.

All pressure contributions after the virtual pressure sensor will be automatically compensated, since these will directly affect the measurement system pressure. Thus, for example, if a flow restrictor is added or removed, the measured system pressure will vary as well as the calculated pre-column pressure. Changes in the flow path between the system pressure sensor and the column must be noted in the estimation.

According to one embodiment, where the viscosity is not known, the controller may assume the use of water, thereby being able to estimate the viscosity for different temperatures using a known expression such as:

，

wherein, T = temperature [ K]；；；。

In real-world situations, there may be some factors that may affect the accuracy of the virtual pressure estimate. If the viscosity of the liquid is unknown and it is assumed to be water, but it has a higher viscosity, the estimated value Δ P for the flow path becomes too low. The calculated value for the virtual pressure signal then becomes higher than the actual value, whereby a pressure alarm will be triggered before the actual pressure becomes too high for the column. This is also the case if other components in the flow path (e.g. mixers, valves, etc.) generate a certain back pressure. Thus, for liquids where the viscosity is lower than water, the estimate will give a lower virtual pre-column pressure than the actual pressure. However, such liquids are mainly used for high pressure columns, where high accuracy of the pressure signal is not required, since most such columns withstand higher pressures than they are normally used with. According to one embodiment, the system is arranged to estimate the incremental column pressure by applying the same principles to the flow path after the column, and the virtual post column pressure can be estimated and used to calculate the virtual incremental column pressure.

As noted, the virtual pre-column pressure and the incremental column pressure may be used to control operation of the chromatography system, for example, by monitoring the pressure against a predefined or user-defined pressure limit, or by operating the chromatography system at a predefined column pressure, or the like.

The present disclosure illustrates a chromatographic process configured to operate with at least one column and configured for purifying a sample comprising a target product using a predefined process. The predefined process may be a general process, a verification process, or a special process, and can be predefined by the manufacturer or generated by the end user.

FIG. 9 illustrates data flow in a system suitable for fabricating columns and liquid chromatography systems. As described in more detail below, the column data is generated during the manufacture of the column and stored in a database dB accessible to the chromatography system 90. The data flow is indicated by dotted lines and the control signals are indicated by dashed lines.

The liquid chromatography system comprises a controller 91 configured to control operation of the chromatography system to run a predefined process, retrieve column data dB accessible from a data storage (the column data being specific to each column), and adapt at least one process parameter of the predefined process for each column based on the column data, whereby the predefined process is adapted to each column to obtain a target product and maintain performance of the liquid chromatography system.

The data storage may be a database (which is integrated in the chromatography system or accessible from a source external to the chromatography system (e.g., a cloud-based implementation)). Another alternative is to store the column data on individual columns, e.g. as memory chips, and to communicate with the chromatography system via RFID.

The process parameters include: pressure above each column; the flow rate of the sample into the column; the flow of residue (residue) out of the column; and/or the processed volume of the sample/time period (column volume/hour).

According to some embodiments, the liquid chromatography system is configured to operate with a single column for purifying the sample, and according to some embodiments, the liquid chromatography system is configured to operate with at least three columns for continuous purification of the sample.

Fig. 8 illustrates the concept of assigning columns to types 400 (also referred to as series). Each individual column 401-1, 401-2, 401-n in this example belongs to a specific type 400 having predetermined production parameters associated with the column. In this example, the production parameters define the range of different physical properties of the column assembly used and the different process properties used to manufacture the column.

The column includes a vessel (vessel) for holding the resin and the filter, and the column assembly may include hardware specific properties, physical dimensions of the vessel (e.g., height of the vessel), material properties of the resin (e.g., particle size and distribution), physical properties of the filter, and the like.

The process properties relate to the manufacturing process of the column, such as the height of the resin bed, the pressure boundary, the flow specification (specification), etc. The height of the resin bed may be the actual height or the height of the vessel.

The production parameters may further include the actual volume of resin in each column.

In some example embodiments, the liquid chromatography system further comprises a sensor 92a, 92b, said sensor 92a, 92b being adapted to read a sensor parameter, wherein the adaptation of the at least one process parameter is further based on the sensor reading. In some embodiments, the sensor readings include any of the following: UV, flow rate and pressure.

In some embodiments, the production parameters further include the actual volume of resin in each column.

The database accessible by the system may include historical data for each individual column and/or columns that belong to the same particular type (i.e., the same series) that was previously used in the liquid chromatography system to purify the sample. Historical data information from the controller is stored in a database for this purpose.

FIG. 10 is a flow chart illustrating a method for manufacturing a column for a liquid chromatography system. The present disclosure also includes a method for manufacturing a column for a liquid chromatography system, the column having an inlet and an outlet and comprising a vessel for holding a resin. The method comprises the following steps:

1) the type of column configured to be used in the process to purify the sample including the target product is selected (step S2).

2) A resin (which is a medium) is selected (step S3) based on the type of the column, the resin having a medium property. The media properties include measured and calculated properties of the media intended to be used in the column. Media parameters are affected by the media manufacturing process and the tolerances of the different media components used in manufacturing the media.

3) A vessel (i.e., hardware) for holding the resin is selected (step S4), the vessel having hardware properties. The hardware properties include measurement and calculation properties of the different components used to manufacture the column, which affect the functionality of the column, such as physical dimensions (with tolerances) and filter properties (if included).

4) The vessel is filled (step S5) with resin to form a resin bed using a column type-based pressure to establish a height of the resin bed.

5) Production parameters are determined based on the media properties and the hardware properties (step S6) to define column data for the column, and the column data is stored in a data storage device accessible to the liquid chromatography system. The data storage device may be integrated in the column or be a database accessible to the chromatography system, as described above.

In some examples, the column further comprises hardware in the form of a top filter disposed between the inlet and the resin bed, and the method further comprises: selecting a top filter based on the type of column, the top filter having top filter properties; and determining a production parameter further based on the top filter property to define the column data.

In some examples, the column further comprises hardware in the form of a bottom filter disposed between the outlet and the resin bed, and the method further comprises: selecting a bottom filter based on the type of column, the bottom filter having bottom filter properties; and determining a production parameter further based on the bottom filter property to define the column data.

The present disclosure also includes a column for a liquid chromatography system, the column having an inlet and an outlet and comprising a vessel for holding a resin, wherein the column is manufactured according to the above method. A system for manufacturing columns is illustrated in fig. 9, where the output from the system is the column and column data, which is stored in a database dB accessible to the chromatography system.

In some examples, the column further includes hardware in the form of a data storage device (e.g., a data chip) configured to store column data and a communication device configured to communicate the column data to the liquid chromatography system.

In some embodiments, the communication device is configured to communicate with the liquid chromatography system using RFID.

FIG. 11 is a flow chart illustrating a method for controlling a liquid chromatography system configured to operate with at least one column. The present disclosure also includes a method for controlling a liquid chromatography system configured to operate with at least one column and configured for purifying a sample comprising a target product using a predefined process, wherein the method comprises:

A) the operation of the chromatography system is controlled (step S12) to run a predefined process.

B) Column data accessible from a data storage is retrieved (S13), the column data being specific to each column, and at least one process parameter of a predefined process for each column is adapted (step S15) based on the column data.

Whereby a predefined process is adapted to each column in order to obtain a target product from the sample and maintain the performance of the liquid chromatography system.

In some examples, each column is of a specific type, and the data storage comprises historical data for each column and/or columns belonging to the same specific type previously used in the liquid chromatography system for purifying the sample, wherein the method further comprises adapting at least one process parameter of the predefined process based on the historical data (step S13 a).

In some examples, the data storage device comprises column data relating to production parameters at the time of production of each column, the production parameters comprising height of the resin bed, pressure boundary, flow specification, material properties, hardware specific properties, filter properties and physical dimensions of the vessel, wherein the method further comprises adapting at least one process parameter of the predefined process based on the production parameters (step S13 b).

In some examples, the liquid chromatography system further comprises a sensor adapted to read a sensor parameter, wherein the method further comprises adapting the at least one process parameter further based on the sensor reading (step S14 a). The sensor readings may include any of the following: UV, flow rate and pressure.

The data storage device may be selected as a database, which may be integrated in a liquid chromatography system. An alternative is to integrate a data storage device in each column.

In addition to the above, the preparation and identification of packed columns is an important step to ensure the robustness and safety of both the purification process and the final product. The column efficiency test plays a central role in the identification and monitoring of packed bed performance. Even though it cannot be used as a single parameter to predict purity and recovery (recovery), it is a rapid way to test column and equipment performance before starting the purification process. This test can also be used between runs to check for changes in bed integrity.

This annotation provides an overview of the theory behind the column efficiency test and the experimental test practices used therein. Test conditions are recommended and key parameters that affect measurement efficiency are discussed in order to facilitate development of robust test protocols.

The efficiency test is an analysis of the residence time distribution of the tracer substance (tracer substance) passing through the column. Typical test signals applied to the column are pulse or step signals. In order to characterize the chromatography column without interference, the tracer substance and eluent conditions are selected such that chemical interaction with the medium and perturbation of the fluid flow are avoided. The most common type of test signal applied is a pulse function. A small volume of tracer substance is added to the liquid stream near the inlet of the column and this pulse broadening (broadening) is analyzed as measured as a chromatographic peak at the column outlet.

Column efficiency is generally defined in terms of two parameters:

peak broadening above the column is described by an equivalent number of theoretical plates (equalization stages)

Peak symmetry by peak asymmetry factor A_sTo describe

Peak broadening is generally described as the number of plates N or equivalent to the theoretical plate Height (HETP). This concept is equivalent to a serial tank model, which reflects the number of equalization stages represented by the columns.

A widely used method for evaluating pulse tests (to determine plate number) involves measuring the peak width at half the maximum peak height. When applying the moment method, this approach is an alternative to numerical curve integration, where the first moment is the mean and the second moment is the variance of the retention volume/time. As outlined in fig. 1, the impulse response is plotted against time or elution volume, and the peak width at half-peak height is measured and related to the elution time or preferably to the elution volume at the maximum peak height. The retention time or retention volume measured at the maximum peak height corresponds to the average residence time or volume found under a symmetrical (gaussian) peak shape. A dimensionless and therefore convenient parameter for efficiency characterization is to reduce the plate height h. This parameter facilitates comparison of column efficiencies regardless of column length and media particle size.

The optimum column efficiency generally corresponds to an experimentally determined reduced plate height of h.ltoreq.3 for the porous media employed in chromatography in bioprocesses. This efficiency is achieved when the fully packed bed is tested with an optimized set of columns and systems under optimal test conditions.

A detailed description of the column efficiency test is provided in the application note (application note) 28-9372-07 AA issued by GE Healthcare, which is incorporated herein by reference in its entirety.

In one embodiment of the invention, the chromatography system is arranged to perform an automated column efficiency test to determine, for example, h and A_sSuch as efficiency parameters, and further provide guidance to the user regarding column efficiency. The automated test provided herein gives all possible data for calculating the height of the reduced plate, as the software automatically applies specific parameters to a given column and resin to calculate the efficiency of the column.This gives the possibility to know directly whether a given column is good enough for the application. If not, the user will be notified so that a more appropriate column can be used. An appropriate cleaning protocol can be applied, the column can be refilled, the column can be replaced by a new column, or any other necessary action can be taken. This gives a safer and robust procedure for the liquid chromatography step, since the efficiency of the column can be guaranteed.

According to one embodiment, the chromatography system is arranged to automatically determine, for example, the reduced plate heights h and A_sSuch as one or more efficiency parameters, and based on the results, direct a user of the system to perform the relevant action, or alternatively, the system may be configured to automatically initiate some or all of the relevant actions based on the efficiency parameters.

In one example, the system may be configured to automatically or semi-automatically initiate one or more of the following actions in order to improve column efficiency:

standard column cleaning protocol (e.g. NaOH 0.5M) when 3 < h < 3.5 (grade 1)

Strong (intense) column cleaning protocol (e.g. NaOH 1M, acid and/or isopropanol) when 3.5 < h < 5 (grade 2)

Column refill protocol when 5 < h < 7 (3 rd stage)

Cancel procedure and instruct user to replace resin when h < 7 (4 th level)

Once the action is completed, the system is preferably configured to automatically determine one or more efficiency parameters and verify the improved efficiency.

In alternative embodiments, the system may be configured to repeat one or more of the above acts two or more times without reaching the desired result. The system may be further configured to upgrade to a higher level after a predetermined attempt at one level.

It should be noted that the above ranges for the different grades may be different depending on the resin type and the column type. The range may also be different depending on the chromatographic process step in which the column is to be used. For example, a column prepared for the capture step may have a higher h than a column prepared for the intermediate or polishing step.

The chromatography system may be arranged to retrieve particle size data from a predefined standard data file in a database or from measurement data of the particle size of the particular resin batch (lot) used for determining the efficiency parameter. The batch-specific particle size data may be determined by individual or interconnected measurement units, or alternatively retrieved from a database, e.g. a cloud-based database, comprising the batch-specific particle size data.

According to one embodiment, the automated column check further comprises determining a column delta pressure to check column packing performance. In case the determined delta pressure is determined to be too low, there is an increased risk for gap building, whereas in case the delta pressure is determined to be too high, the system pressure range may be exceeded. The relevant range for incremental pressure may be retrieved from a column database and/or system settings.

The present disclosure further includes a computer program for controlling a liquid chromatography system comprising instructions which, when executed on at least one processor, cause the at least one processor to carry out the method according to the above method.

The present disclosure further comprises a computer readable storage medium carrying a computer program for controlling a liquid chromatography system as defined above.