Patent Description:
Various sensors and techniques relating to chromatographic procedures are known. For example, <CIT> discloses one such in which chemical sensors for gas chromatography are used.

In contrast, liquid chromatography is a well-known procedure for separating mixtures of molecules, for example separating proteins in liquid samples (see, for example, <CIT>). The proteins may typically be suspended in a fluid, and driven through a chromatography separation medium along with a buffer solution. The various sample molecules of the mixture travel at different speeds through a chromatography medium, causing them to separate. This separation may be completed by a fractionation step where the mobile phase may be directed to different containers, e.g. by an outlet valve of the chromatography apparatus.

In some applications, e.g. in the biopharmaceutical field, recent advancements in genetic engineering and cell culture technology have driven expression levels higher than ever, putting a considerable burden on down-stream purification, especially the fractionation step. While the introduction of new chromatography media compositions significantly improves the efficiency of a process based on conventional fixed bed chromatography, additional gains can be achieved by operating in a continuous manner. The latter is especially appealing when continuous bioreactors, such as those operated in perfusion mode, are employed.

In continuous chromatography, two or more identical columns are connected in an arrangement that allows columns to be operated in series and/or in parallel, depending on the method requirements.

Thus, all columns can be run in principle simultaneously, but in different stages. The procedure may be repeated, such that each column is loaded/packed, eluted, and regenerated several times in the process. Compared to 'conventional' chromatography, wherein a single chromatography cycle is based on several consecutive steps, such as loading, wash, elution and regeneration, in continuous chromatography based on multiple identical columns all these steps occur simultaneously but on different columns. Continuous chromatography operation results in a better utilization of chromatography media compositions, reduced processing time and reduced buffer fluid requirements, all of which benefits process economy. Continuous chromatography is sometimes denoted simulated moving bed (SMB) chromatography.

As previously mentioned each column may be loaded/packed, eluted, cleaned, and regenerated several times in the process. An essential factor for a reliable continuous chromatography process is the quality of the columns used, and more specifically the similarity or even density of media in each column. If the columns are not identical, the theoretical calculations will not be correct, and it will become difficult to design an efficient and robust continuous chromatography process. However, the loading/packing of a column with chromatography media composition, is very complex in order to obtain repeatable results. Even small differences in the number of plates or other packing properties can have a huge effect on the end result.

A problem with conventional solutions is that performing continuous chromatography is a cumbersome, complex and time consuming operation. Often the process must be interrupted to perform reconnection of fluid couplings/tubes, to perform packing of columns or to load a pre-packed column, to perform cleaning operations etc..

Thus, there is a need for an improved chromatography apparatus for performing continuous chromatography.

Furthermore, even though chromatography apparatus which performs continuous chromatography of the type mentioned above are known, for example those sold under the tradename of ÄKT Apilot, and these known instruments function well, providing a versatile instrument which can be re-plumbed for different functionality if needed, these instruments lack some functionality, such that there are only certain procedures in which they can be employed, and an extension of that versatility would be commercially attractive, but is not easy to achieve with the competing restraints of instrument size, maintaining versatility, practical aspects such as ease of cleaning internally and externally, and cost.

Embodiments of the present invention address the problems mentioned above. Thereby, an object of the present invention is to provide a chromatography instrument with one or more of: an increased functionality, for example able to operate in conventional batch chromatography as well as continuous chromatography; is useable across a wider range of applications; does not have a substantially increased overall size or manufacturing cost; and is simple to operate.

Good Manufacturing Practice (GMP) sets out guidelines for bioprocessing procedures, which if followed require cleanliness standards. Advantageously, the standards are easier to achieve with the proposed apparatus, for example where fluid paths in the instrument have, in one configuration at least, a continuous flow path with no substantive stagnant portions, thereby providing complete cleaning without the need to break down the fluid conduits. Embodiments of the proposed instrument provide a sanitary small-scale chromatography instrument suitable for both GMP and non-GMP work. Functionally wide flow and pressure ranges of the instrument makes it fit for both production of technical batches and scale-up studies as well as small-scale production of GMP-grade material. The high accuracy and flow range of the pumps enables precise gradient formation, covering a large range of chromatography column sizes and more repeatable results.

In embodiments, a modular construction provides increased functionality, for different uses. Interactive control software allows changes to be made in real-time and unexpected deviations to be quickly identified. The small, bench-top size frees up lab space. The instrument allows in-situ column packaging, i.e. the ability to compress chromatography media in the column, or each column where two or more columns are used, whilst being connected to the instrument, and without having to then disconnect any fluid conduits prior to performing chromatographic procedures.

More specifically, the present invention, as defined by the appended claims, is provided.

More advantages and benefits of the present invention will become readily apparent to the person skilled in the art in view of the detailed description below.

The invention will now be described in more detail with reference to the appended drawings, wherein:.

<FIG> shows a chromatography apparatus <NUM> according to an aspect of the invention. The apparatus includes, but it not limited to, individual modules <NUM> to <NUM> as listed below, at least some of which are demountable from an apertured front panel <NUM> of a housing <NUM> of the apparatus <NUM> and mounted thereon in one generally vertical plane, such that the liquid connections required between modules can be made only at the front face <NUM>. In practice the demountable modules have no more than two standard sizes which can, if needed, be repositioned on the panel <NUM> to suit a different procedure. Each module has a serial bus communication connection and power connection so that its physical position is immaterial to a controller for example located in the housing <NUM>, or located remotely. Thereby, the modules can be regarded as modular and thereby repositionable and/or interchangeable.

The chromatography apparatus shown in <FIG> has the following module modules:.

Modules can be omitted or repositioned as explained above. It will be apparent that some modules can be replaced with other modules or the space left by an omitted module can be filled with a blanking plate (see e.g. <NUM> <FIG>). More than one of the same numbered modules can be used where necessary.

Fluid interconnections between the fluid manipulating modules of the apparatus i.e. all the modules listed above except modules <NUM>,<NUM> and <NUM>, and external modules for example sample input reservoirs, buffer fluid reservoirs, chromatograph column(s) and fraction collection equipment, all not shown in <FIG>, are made via fluid conduits in this case in the form of flexible plastics tubing, which can be readily coupled and uncoupled to corresponding ports of the fluid manipulating modules, in any desired configuration, for example using a coupling as disclosed in a related application <CIT>.

<FIG> shows one possible liquid interconnection configuration between the main modular modules of the chromatography apparatus, connected in this case to two chromatography columns <NUM> and <NUM>, although the apparatus allows any workable interconnection between modules and additional parts such as multiple columns, and liquid reservoirs. Reconfigurable liquid interconnections are denoted by short chain dotted lines <NUM>.

At the heart of the apparatus <NUM> is the column valve <NUM>, which in this case has a construction as disclosed in our co-pending patent application filed on the same day as this application, and having the title 'VALVE UNIT FOR A CHROMATOGRAPHY APPARATUS. ' The value unit <NUM> provides multiple switching of flow for allowing flow in one or both columns <NUM>/<NUM> in either direction (up or down in the drawing). The user can select upflow or downflow, or select to bypass one or both columns. The flow can be directed to waste or to the next component in the flow path. The columns can also be connected in series. The Column valve includes pre-column and post-column pressure sensors. In addition the valve has a port <NUM> which can be used to change the volume of hydraulic cylinders <NUM> and <NUM> which are part of the columns <NUM> and <NUM>, for example to provide compression of the columns' contents, also known as column packing. That packing procedure can be automated. With such a system column diameters of between about <NUM> and <NUM> have been found to be packable in this way. The columns can be pre-packed, but rinsed and re-consolidated with the aid of pressure sensors in the value unit <NUM> measuring back-pressure resulting from pressure within the columns and in accordance with to known protocols, for example as described in <CIT>.

The remaining system <NUM> comprises:
Inlet valve groups A and B, <NUM>,<NUM>,<NUM> and <NUM>, suitable for providing selectable liquids including sample containing liquids buffer solutions, and cleansing fluids.

The inlet valves supply two system pumps, here each having a pair of pistons and associated one-way valves, providing a variable flow rate of between <NUM>-<NUM> per minute each (<NUM>/min max), with a high volume and resolution of flow, enabling accurate flow rates to be maintained. Such accuracy enables good repeatability of results for a wide range of column diameters.

The pumps supply, in series a flow restrictor <NUM>, which includes a system pressure monitor, a mixer valve <NUM>, and a mixer module <NUM>, before pumped liquid is diverted to the column valve unit <NUM>.

Any entrained air can escape via an air trap valve <NUM>, and an air trap vent <NUM>, which vent also has an air escape from the columns <NUM> and <NUM>. The air trap may be constructed in accordance with pending application <CIT>.

Once liquids reach the column valve unit they can be routed in accordance with the arrangement described in said co-pending application with the title 'VALVE UNIT FOR A CHROMATOGRAPHY APPARATUS', and thereby numerous modes of chromatography can be performed, from simple batch work, where a straightforward chromatographic separation process is performed using just one column, to procedures which more closely copy larger scale commercial procedures where two or more columns can be employed, one being readied for use while the other is being used for separation. The valve port numbering used is the same as that used in said co-pending application and here has the same arrangement.

Output from the chromatography column(s) is passed out through port <NUM> to: a conductivity monitor <NUM>, a UV light absorption monitor <NUM>, and a pH monitor <NUM>, and is thence directed into an appropriate storage vessel in dependence on the signals from the three monitors, and thereby, separated fractions are collected in an appropriate vessel <NUM>. Column washings can be collected in a waste vessel <NUM>.

The long chain dotted lines in <FIG> represent a system bus <NUM> which carries signals and power to and from the modules mentioned above, to and from a controller <NUM>. It will be appreciated that controlling and monitoring signals may be transmitted wirelessly according to known protocols, doing away with the need for a communication bus. The chromatography system <NUM> includes also a display screen <NUM>. Software running on the controller will display plural icons on the screen <NUM> and allow user manipulation of the icon on the screen to drag and drop the icons to form a series of icons representative of a user defined chromatography control method, for ease of use.

<FIG>, <FIG> and <FIG> show the system connected with tubing for various configurations, where only some of the modules referenced in <FIG> remain in place in these figures, and the apertures left by removed modules are blanked off with blanking plates <NUM>, screwed into place over the aperture to prevent accidental liquid ingress into the housing <NUM>.

In <FIG> a system <NUM>' with a configuration of modules suitable for regulated environments where systems are custom-built in a factory. The system is delivered mounted, calibrated, and performance tested and suitable for work in GMP environments. <FIG> shows one system with some modules removed, and <FIG> shows a system <NUM>‴ with more modules in place, similar to <FIG>, and showing typical tubular interconnections <NUM>.

In use, modules are easily removed or added to the system and installation finalized through a one-click activation in software which can recognize each module. The software can provide comprehensive and customizable operational control as well as pre-emptive maintenance. In addition to the modules described above, input-output communication modules can be used to interface with analog and/or digital external sensors or other equipment such as automatic fraction collecting devices. The wide flow rate and pressure ranges enables more than <NUM>-fold scaling in the range <NUM> to <NUM> internal diameter columns. This wide range makes the apparatus suitable to bridge the transition into GMP environments.

The packing (and re-packing) of chromatography columns, using the system described above is controllable fully by the controller <NUM> initiated by the control panel <NUM>. The controller <NUM> is able to drive the a display screen <NUM> (<FIG>) to aid visualisation of the packing process and progress. The control software includes an accessible column packing record. Columns packing records can therefore be defined, created, and updated from the software for traceability and quality assurance purposes. In addition, the record can be used to monitor column performance and provide statistics for usage, separation performance, and packing intervals.

The display screen can provide a process visualization which quickly gives an operator an overview of the system's function, progress through operational steps and alarms, only providing the desired amount of information at each step. The active flow path is always displayed in the process visualisation to minimize user errors. Real time changes can be made by selecting the appropriate process on the visualization screen, e.g. selecting or dragging icons on the screen. Control, graphical interfaces are provided for specific sections, such as the column valve <NUM>.

Preprogrammed steps are employed but these can be modified and saved as user-defined steps for added customization.

Claim 1:
A chromatography system (<NUM>) for separating mixtures of molecules in liquid samples, comprising:
a plurality of modules (<NUM> ... <NUM>) including at least one pump (<NUM>, <NUM>) and a column valve unit (<NUM>) connectable to plural chromatography columns (<NUM>, <NUM>); and characterised by:
a controller (<NUM>), the controller (<NUM>) being operable to control the or each pump (<NUM>, <NUM>) and the column valve unit (<NUM>) to perform different chromatographic processes, including chromatography employing just one column (<NUM>, <NUM>), as well as chromatography employing two or more columns (<NUM>, <NUM>) by selective valve opening in said column valve unit (<NUM>); wherein:
at least one of said plurality of modules (<NUM> ... <NUM>) is demountable from an apertured front panel (<NUM>) of a housing (<NUM>) of the chromatography system (<NUM>), and mounted thereon in one generally vertical plane, such that liquid connections required between respective modules (<NUM>...<NUM>) can be made only at a front face (<NUM>) of the chromatography system (<NUM>), wherein the controller (<NUM>) is further operable to perform one or more of column packing, column packing testing, and cleaning of at least a part of the chromatography system (<NUM>).