Sample preparation system and method for preparing sample using same

Abstract

The invention relates to a sample preparation system and a method of preparing a sample using the same.

Description

Technical Field

[0001] The present invention relates to a sample preparation system and a method for preparing a sample using the sample preparation system. Background Art

[0002] In order to produce a target substance such as a protein by purification, it is usually necessary to perform a clarification step on the sample containing the target substance before purification to remove contaminants, which include host cell-derived contaminants (e.g., intact cells, cell debris, host cell nucleic acids, and host cell proteins), culture medium-derived contaminants (e.g., proteinaceous and non-proteinaceous culture medium components), and vector-derived contaminants, including vector nucleic acids and viral vectors.

[0003] Clarification and purification are usually performed in two separate devices, wherein a first device clarifies the sample containing the target substance, and a different second device uses the clarified output from the first device as input for purification, which is usually provided manually to the second device. Therefore, the combination of clarification and purification of the sample is usually a rather slow and inefficient process, which may lead to degradation of the sample. Summary of the invention

[0004] In view of the above, the technical problem of the present invention is to provide a sample preparation system with which two unit operations (eg, clarification and purification) can be quickly performed in one device to improve the efficiency of each process.

[0005] The solution to the above technical problem is achieved through the embodiments described in the claims.

[0006] In particular, in one aspect, the present invention provides a sample preparation system comprising:

[0007] a serial arrangement of a first mechanism configured to be connected to a first consumable and a second mechanism configured to be connected to a second consumable;

[0008] a first valve disposed between the first mechanism and the second mechanism;

[0009] a second valve disposed downstream of the second mechanism;

[0010] a first line providing fluid communication between the first mechanism and the first valve;

[0011] a second line providing fluid communication between the first valve and the second valve via the second mechanism;

[0012] a third line providing fluid communication directly between the first valve and the second valve, configured to bypass the second mechanism; and

[0013] A pump is configured to move fluid within the sample preparation system.

[0014] According to the present invention, the sample preparation system comprises a serial arrangement of a first mechanism and a second mechanism. In particular, the second mechanism is located downstream of the first mechanism in the sample preparation system, and the first mechanism and the second mechanism are integrally connected to each other in the sample preparation system. The term "integrally connected" as used herein refers to being arranged in one device, i.e., arranged in the sample preparation system, rather than being arranged in a separate device. Since the sample preparation system of the present invention allows the first unit operation (e.g., clarification) and the second unit operation (e.g., purification) to be combined in an automatic manner in one device, the corresponding method is fast and has significantly improved process efficiency compared to the method performed in two separate devices.

[0015] The terms "automatic" and "automatically" used herein mean that the corresponding steps can be performed without human intervention. On the contrary, the word "manually" used herein means that the corresponding steps are performed by a person.

[0016] The first mechanism of the sample preparation system is not particularly limited as long as it is configured to be connected to the first consumable. According to an embodiment, the first mechanism is configured to be connected to the first consumable and support the first consumable. For example, the first mechanism can be a first filter holder or the end of the first line, thereby providing fluid communication between the first mechanism (i.e., the end of the first line itself in this case) and the first valve, which supports the first consumable from the downstream side of the first consumable. In the case where the first consumable is supported by the end of the first line from its downstream side, the first consumable is preferably supported on its upstream side by the end of the line connected to the first mechanism upstream of the first mechanism.

[0017] Preferably, the first mechanism is a first filter holder. In an embodiment, the first filter holder includes a first clamp configured to clamp the first consumable on an upper surface (i.e., upstream) and a second clamp configured to clamp the first consumable on a lower surface (i.e., downstream). Therefore, during operation, the first consumable can be firmly supported to resist internal pressure. According to an embodiment, each of the first clamp and the second clamp includes an O-ring configured to connect the upstream and downstream flow paths of the first mechanism when the O-ring is in a compressed state (i.e., when the clamp is closed). Preferably, one of the first clamp and the second clamp is a static clamp, i.e., it does not move, and the other is a movable clamp. Therefore, by pressing the movable clamp against the static clamp, the first consumable can be clamped. Preferably, the first clamp is a static clamp, and the second clamp is a movable clamp.

[0018] According to a preferred embodiment, the sample preparation system further comprises an input stack configured to hold at least one first consumable. Preferably, the input stack is configured to hold from 1 to 150 first consumables, more preferably from 1 to 50 first consumables.

[0019] In addition, the second mechanism is not particularly limited as long as the second mechanism is configured to be connected to the second consumable. According to an embodiment, the second mechanism is configured to be connected to the second consumable and support the second consumable. For example, the second mechanism may be a second filter holder or a component that may correspond to the second line to provide fluid communication between the first valve and the second valve.

[0020] When the second mechanism corresponds to a component of the second line, the second line is interrupted between the first valve and the second valve, and the second consumable is located at the interruption and supported by the first end component of the interrupted second line on the upstream side and the second end component of the interrupted second line on the downstream side.

[0021] According to another embodiment, wherein the second mechanism is a second filter holder, the second line is interrupted between the first valve and the second valve, and the second filter holder is located at the interruption and is connected to the first end member of the interrupted second line on the upstream side and the second end member of the interrupted second line on the downstream side. In an embodiment, the second filter holder includes a first clamp configured to clamp the second consumable on the upper surface (i.e., upstream) and a second clamp configured to clamp the second consumable on the lower surface (i.e., downstream). Therefore, during operation, the second consumable can be safely supported to resist internal pressure. According to an embodiment, each of the first clamp and the second clamp includes an O-ring configured to connect the upstream and downstream flow paths of the second mechanism when the O-ring is in a compressed state (i.e., when the clamp is closed). Preferably, one of the first clamp and the second clamp is a static clamp, i.e., it does not move, and the other is a movable clamp. Therefore, the second consumable can be clamped by pressing the movable clamp against the static clamp. Preferably, the first clamp is a static clamp and the second clamp is a movable clamp.

[0022] In the present invention, the first valve is arranged between the first mechanism and the second mechanism. The first valve is configured to guide the fluid to the second line or the third line of the sample preparation system. According to an embodiment, when the fluid is guided to the second line, the flow of the fluid to the third line is blocked. According to another embodiment, when the fluid is guided to the third line, the flow of the fluid to the second line is blocked. The first valve is not particularly limited. In an embodiment, the first valve can be, for example, a three-way diaphragm valve having three ports and can be in one of two states, directing the flow to one of its two outlets (i.e., the second line and the third line). In another embodiment, the first valve is a three-way ball valve.

[0023] In addition, the second valve is arranged downstream of the second mechanism. The second valve is configured to guide the fluid to the line downstream of the second valve. The second valve is not particularly limited. In an embodiment, the second valve can be, for example, a three-way diaphragm valve having three ports, two inlets of which are connected to the second line and the third line, and the three-way diaphragm valve can be in one of two states, guiding the flow from one of its two inlets (i.e., the second line and the third line) to the line downstream of the second valve. In another embodiment, the second valve is a three-way ball valve. Preferably, the line is the fourth line to be described further below.

[0024] The pump used in the sample preparation system of the present invention is not particularly limited. The pump is configured to move the fluid in the sample preparation system. For example, the pump can be a peristaltic pump, a diaphragm pump, or a piston pump. In an embodiment, the pump is arranged upstream of the first mechanism. In addition, the pump can be arranged downstream of the input sample valve to be described below. In addition, the pump can be arranged downstream of the process fluid input valve to be further described below. Therefore, the flow of the fluid in the sample preparation system can be easily controlled.

[0025] According to an embodiment of the present invention, the sample preparation system also includes a consumables processing device connected to the first mechanism, the consumables processing device is configured to provide the first consumable to the first mechanism, and receive the first consumable from the first mechanism. The consumables processing device operates in an automatic manner. Preferably, the consumables processing device is connected to the input stack, wherein the consumables processing device is configured to receive the first consumable from the first mechanism and provide the first consumable to a waste bin, and receive the first consumable from the input stack and automatically provide the first consumable to the first mechanism, and vice versa. By using the consumables processing device, new (unused) first consumables can be easily provided to the first mechanism, and the used first consumables can be easily and automatically removed.

[0026] The consumables handling device is not particularly limited. In an embodiment, the consumables handling device is an index wheel. Preferably, the index wheel has an opening in its in-plane direction, and the opening conforms to the shape of the first consumable at the first position of the index wheel. According to an embodiment, the index wheel can be rotated, thereby receiving the first consumable from the input stack and providing the first consumable to the first mechanism. In addition, the index wheel can be rotated to receive the first consumable from the first mechanism and provide it to the waste bin. In an embodiment, the consumables handling device is a filter handling device.

[0027] In an embodiment of the present invention, the sample preparation system further comprises an input line, which provides fluid communication between an input cup arranged upstream of the first mechanism and the first mechanism. According to an embodiment, the input sample valve is located downstream of the input cup and upstream of the first mechanism on the input line. Therefore, the input sample valve divides the input line into a portion upstream of the input sample valve and a portion downstream of the input sample valve. In this embodiment, the input sample valve is configured to guide the fluid to the input line or waste container downstream of the input sample valve. According to an embodiment, in the case where the fluid is guided to the input line downstream of the input sample valve, the flow of the fluid to the waste container is blocked. According to an alternative embodiment, in the case where the fluid is guided to the waste container, the flow of the fluid to the input line downstream of the input sample valve is blocked. The input sample valve is not particularly limited. In an embodiment, the input sample valve can be, for example, a three-way diaphragm valve having three ports and can be in one of two states, directing the flow to one of its two outlets (i.e., the input line and the waste container downstream of the input sample valve). In another embodiment, the input sample valve is a three-way ball valve. The input cup is configured to hold a fluid, preferably a sample to be prepared. According to an embodiment, the input cup has a volume from 10 mL to 150 mL, preferably from 20 mL to 80 mL.

[0028] In an embodiment, the sample preparation system further comprises a pipetting robot configured to transfer a fluid (such as a sample to be prepared) from at least one input container to an input cup. In an embodiment, one input container has a volume of 2 mL to 20 mL. The input container is not particularly limited and may be selected from a culture container, a rotary tube, and a multi-well plate.

[0029] There is no particular limitation on the pipetting robot that can be used in the present invention. According to an embodiment, the pipetting robot includes a pipette tip connected to a circulating pH electrode and a fluid input valve. The fluid input valve is configured to provide, for example, water, sodium hydroxide and / or pH neutralizing buffer to the pipette tip. Preferably, the pipetting robot also includes a pipette liquid sensor arranged on a pipette robot line, which provides fluid communication between the pipette tip, the pH electrode and the fluid input valve. The pipette liquid sensor is configured to detect whether there is liquid at its position. In addition, the pipetting robot may include a syringe connected to the circulating pH electrode.

[0030] In an embodiment, the sample preparation system also includes a pipette washing station, which is configured to receive the pipette tip of the pipette robot. In particular, the pipette washing station can have a wall, and when the pipette robot is arranged in the pipette washing station, the wall surrounds the pipette tip of the pipette robot. In addition, the pipette washing station can have an outlet at its bottom, and liquid can be discharged through the outlet. Specifically, the liquid can be provided by at least one of the fluid input valves of the pipette robot, and can be used to wash the pipette tip when the pipette robot is arranged in the pipette washing station. The outlet can be connected to a waste container, and the liquid can be pumped to the waste container by using a tip washing waste pump.

[0031] Since a pipetting robot may be used, the type of input container is not particularly limited. Thus, the pipetting robot provides flexibility in the selection of the input container, so that the selection of the input container depends only on the fluid to be processed (such as the sample to be prepared).

[0032] Preferably, the consumables handling device includes a connecting mechanism (means) configured to connect the input line and the first line, for providing fluid communication between the input line and the first line. Therefore, a flow path can be established between the input line and the first line. In the case where the consumables handling device is a dividing wheel, the connecting mechanism corresponds to a hole in the in-plane direction of the dividing wheel, which extends from the first surface of the dividing wheel to the second surface of the dividing wheel at a second position of the dividing wheel. The dividing wheel can be rotated so that the first opening of the hole on the first surface of the dividing wheel is connected to the input line, and the second opening of the hole on the second surface of the dividing wheel is connected to the first line.

[0033] According to an embodiment of the present invention, the sample preparation system further comprises a first liquid sensor arranged upstream of the first mechanism. The first liquid sensor is configured to detect whether liquid is present at the position where the first liquid sensor is arranged. Preferably, the first liquid sensor is arranged downstream of the input sample valve and upstream of the first mechanism. According to this configuration, the first liquid sensor can, for example, detect whether the input cup has been emptied.

[0034] In an embodiment, the sample preparation system according to the present invention further comprises a second liquid sensor, which is arranged downstream of the first mechanism and upstream of the second mechanism. The second liquid sensor is configured to detect whether there is liquid at the position where the second liquid sensor is arranged. Preferably, the second liquid sensor is arranged upstream of the first valve. According to this configuration, based on the detection result of the second liquid sensor, the first valve and the second valve can be switched to guide the fluid through the second line via the second mechanism, or through the third line to bypass the second mechanism.

[0035] The sample preparation system according to the present invention may further include a first pressure sensor, which is arranged upstream of the first mechanism. The first pressure sensor is configured to detect the pressure of the sample preparation system at a position where the first pressure sensor is arranged. Preferably, the first pressure sensor is arranged downstream of the pump and upstream of the first mechanism. In an embodiment, the first pressure sensor is arranged downstream of the pump and the first liquid sensor and upstream of the first mechanism. Therefore, the flow of the fluid flowing to the first mechanism can be easily controlled.

[0036] In an embodiment, the sample preparation system of the present invention further comprises a second pressure sensor, which is arranged downstream of the first mechanism and upstream of the second mechanism. The second pressure sensor is configured to detect the pressure of the sample preparation system at the position where the second pressure sensor is arranged. Preferably, the second pressure sensor is arranged upstream of the first valve. According to this configuration, based on the measurement result of the second pressure sensor, the first valve and the second valve can be switched to guide the fluid through the second line via the second mechanism or through the third line to bypass the second mechanism.

[0037] In an embodiment, the sample preparation system also includes a fourth line that provides fluid communication between the second valve and an output valve disposed downstream of the second valve. The output valve is configured to direct the fluid to an output line that provides fluid communication between the output valve and the output cup, or to a waste container line that provides fluid communication between the output valve and the waste container. According to an embodiment, in the case where the fluid is directed to the output line, the flow of the fluid to the waste container line is blocked. According to an alternative embodiment, in the case where the fluid is directed to the waste container line, the flow of the fluid to the output line is blocked. The output valve is not particularly limited. In an embodiment, the output valve can be, for example, a three-way diaphragm valve having three ports and can be in one of two states, directing the flow to one of its two outlets (i.e., the output line and the waste container line). In another embodiment, the output valve is a three-way ball valve.

[0038] Preferably, the sample preparation system also includes an output line that provides fluid communication between the output valve and the output cup. The output cup is configured to hold the fluid that has been output by the sample preparation system. In an embodiment, the volume of the output cup is 10mL to 150mL, preferably from 20mL to 80mL. According to an embodiment, the sample preparation system includes a liquid transfer robot that is configured to transfer fluid from the output cup to at least one output container. The liquid transfer robot can be the same as the liquid transfer robot configured to transfer fluid from at least one input container to the input cup as described above. In an embodiment, the volume of an output container is 2mL to 80mL. The output container is not particularly limited, and for example can be selected from a 15mL centrifuge tube, a 50mL centrifuge tube, and a multi-well plate.

[0039] Since it is possible to use a pipetting robot, the type of output container is not particularly limited. Therefore, the pipetting robot provides flexibility in the selection of the output container.

[0040] The sample preparation system according to the present invention can also include at least one process fluid input valve, which is arranged upstream of the first mechanism and is configured to provide a process fluid to the sample preparation system. The process fluid is not particularly limited, and for example, can be selected from water, phosphate buffered saline (PBS), elution buffer, low pH stripping buffer and sodium hydroxide. The elution buffer is not particularly limited, and its selection depends on the material to be eluted. For example, the elution buffer can be selected from citrate buffer, glycine / HCl buffer and acetate buffer. Each of the process fluids can be, for example, stored in a bottle and connected to the process fluid input valve by a line.

[0041] In an embodiment, the sample preparation system according to the present invention may further include a target substance detection sensor, which is arranged downstream of the second mechanism and configured to detect the target substance. According to this configuration, the target substance emerging from the first mechanism and / or the second mechanism can be easily detected. In an embodiment, the target substance detection sensor is also arranged downstream of the second valve. Preferably, the target substance detection sensor is also arranged upstream of the output valve. Therefore, according to the detection result of the target substance detection sensor, the fluid can be directed to the output cup or the waste container. Specifically, in the case where the target substance detection sensor can detect the target substance, the output valve can be switched to direct the target substance or the fluid containing the target substance to the output cup. In the case where the target substance detection sensor does not detect the target substance, the output valve can be switched to direct the fluid to the waste container.

[0042] The target substance is not particularly limited. For example, the target substance can be any biomolecule of interest, such as protein (such as antibody), hormone, vaccine, nucleic acid, exosome and virus and virus-like particle. In a preferred embodiment, the target substance is an antibody, more preferably a monoclonal antibody (mAb) or its fragment or derivative or nanobody. The example of a monoclonal antibody is adalimumab, cetuximab, rituximab, infliximab, omalizumab and denosumab. The target substance can be obtained from, for example, mammalian cells, bacterial cells or insect cells, culture medium and cell lines (such as "Chinese hamster ovary cells" (CHO cells), HeLa or human umbilical vein endothelial cells (HUVEC)).

[0043] In a preferred embodiment, the target substance detection sensor is a UV sensor. Specifically, by using a UV sensor, target substances such as proteins can be easily detected.

[0044] The first consumable that can be connected to the first mechanism is not particularly limited. Preferably, the first consumable is a first filter. More preferably, the first filter is a clarifying filter. According to an embodiment, the clarifying filter is a depth filter and / or a membrane, preferably a depth filter. The clarifying filter is configured to separate contaminants from a sample. Preferably, the clarifying filter is a cell culture clarifying filter. The cell culture clarifying filter is configured to separate cells and other contaminants from a sample. In an embodiment, the first consumable is a disposable consumable. The term "disposable consumable" as used herein means that the consumable cannot be cost-effectively recovered after use and is discarded due to, for example, filter clogging.

[0045] The sample to be prepared is not particularly limited as long as it contains multiple components, wherein at least one of the multiple components of the sample is one of the above-mentioned target substances. Other (impurity) components (contaminants) are not particularly limited and may depend on the preparation conditions of the target substance. Examples of other components are aggregates, host cell proteins, deoxyribonucleic acids and fragments thereof, and charge variants.

[0046] The source of the sample is not particularly limited. For example, the sample can be obtained by applying any biological, biochemical, chemical or pharmaceutical method. Therefore, the sample can be obtained by a purification method using different purification units previously performed. For example, the target substance can be produced by a suitable cell line (such as a CHO cell line), for example, by perfusion culture.

[0047] The second consumable that can be connected to the second mechanism is not particularly limited. Preferably, the second consumable is a second filter or a chromatographic column. More preferably, the second consumable is a purification filter or a purification chromatographic column. According to an embodiment, the purification filter is a membrane. In an embodiment, the purification chromatographic column contains functionalized beads. Preferably, the second consumable is configured to bind and elute the target substance. In an embodiment, the second consumable is a protein purification filter or a protein purification chromatographic column, more preferably a protein purification filter. Preferably, the protein purification filter or the protein purification chromatographic column is configured to bind and elute proteins, preferably antibodies. In an embodiment, the second consumable is a reusable consumable. The term "reusable consumable" used herein means that the consumable can be used for multiple runs of sample preparation before it needs to be discarded. For example, a reusable consumable needs to be replaced after performing from 50 to 150 operations. According to an embodiment, the purification filter or the purification chromatographic column includes a binding compound that allows the target substance in the purification filter or purification chromatographic column to be bound, respectively. For example, the binding compound is fixed on / in the filter matrix of the purification filter. The binding compound can be, for example, protein A, and protein A can bind to the target substance at a pH of 7 or higher. The target substance can then be eluted by using an elution buffer with a pH below 7.

[0048] In an embodiment, the sample preparation system further comprises a user interface touch screen. The user interface touch screen allows easy control and operation of the sample preparation system.

[0049] The sample preparation system of the present invention may also include a pH sensor, which is arranged downstream of the second mechanism. The pH sensor is configured to measure the pH of the liquid at the pH sensor position. Preferably, the pH sensor is further arranged downstream of the second valve and upstream of the output valve. According to an embodiment, the sample preparation system also includes a pH adjustment input line, which provides fluid communication between the pH adjusting agent reservoir and the second line upstream of the second mechanism. Therefore, the pH of the second consumable that can be connected to the second mechanism can be easily controlled. In addition, the sample preparation system may also include an elution fraction line, which provides fluid communication between the elution fraction reservoir and the second line downstream of the second mechanism.

[0050] The material of the production line of the present invention is not particularly limited. Preferably, the material can be selected from plastics such as polyvinyl chloride, polypropylene, polyethylene, polytetrafluoroethylene and fluorinated ethylene propylene. Therefore, these lines can provide flexibility and chemical resistance.

[0051] The sample preparation system according to the present invention may include more than one first mechanism and second mechanism arranged in series. In particular, each of these serial arrangements may have the same configuration as the serial arrangement described above. In the case where there is more than one serial arrangement in the sample preparation system, the serial arrangements are arranged in parallel to each other, and each of the serial arrangements has its own input cup and output cup to allow independent operation.

[0052] The sample preparation system of the present invention is capable of performing two unit operations (such as clarification and purification) in one device, so that the corresponding method is fast and has improved process efficiency compared with the method using a separate device. In particular, the target substance included in more than 20 input containers containing unfiltered samples can be clarified and purified, for example, in less than 2 hours. Therefore, the sample preparation system according to the present invention can provide a method for quickly clarifying and purifying samples, thereby sample degradation can be avoided. In addition, due to the automatic operation of the sample preparation system, the need for manual labor of each process can be reduced. In addition, the sample preparation system can produce a well-defined (i.e., representative) output sample containing the target substance, which can be directly further analyzed by various assays.

[0053] According to another aspect, the present invention provides a method for preparing a sample using a sample preparation system, wherein the method comprises at least one of the following steps:

[0054] (i) clarifying the sample by passing the sample through a first consumable connected to a first mechanism to provide a clarified sample, wherein the first consumable is a clarification filter (hereinafter referred to as "clarification process (i)"); and

[0055] (ii) purifying the sample by passing the sample through a second consumable connected to a second mechanism to provide a purified sample, wherein the second consumable is a purification filter or a purification chromatography column (hereinafter referred to as "purification process (ii)").

[0056] The present invention provides a method for preparing a sample using a sample preparation system, wherein the method comprises a clarification process (i), a purification process (ii), or both the clarification process (i) and the purification process (ii).

[0057] The following detailed description of the clarification process (i) relates to a method for preparing a sample using a sample preparation system, wherein the method includes the clarification process (i) but does not include the purification process (ii), and the following detailed description of the purification process (ii) relates to a method for preparing a sample using a sample preparation system, wherein the method includes the purification process (ii) but does not include the clarification process (i). Further details are given below of a method for preparing a sample using a sample preparation system, wherein the method includes both the clarification process (i) and the purification process (ii).

[0058] Unless explicitly stated otherwise, any definition of a feature in the context of a sample preparation system also applies to the corresponding feature of a method of preparing a sample using the sample preparation system.

[0059] According to a preferred embodiment of the method for preparing a sample of the present invention, the sample is a cell culture comprising a protein, the clarification filter is a cell culture clarification filter, and the purification filter is a protein purification filter or the purification chromatographic column is a protein purification chromatographic column. Compared with the method in which clarification and purification are performed in two separate devices, the corresponding method using the sample preparation system according to the present invention can clarify and purify the sample more effectively and faster. In addition, the method of the present invention can produce a well-defined (i.e., representative) output sample containing a target substance (such as a protein, preferably an antibody), which can be directly further analyzed by various assays.

[0060] Clarification process (i)

[0061] According to an embodiment, at the beginning of the clarification process (i), a clarification filter is provided to the first mechanism, wherein the clarification filter is new, i.e., unused (hereinafter referred to as "step (c1)"). In step (c1), preferably, the clarification filter is provided from an input stack by a consumables processing device.

[0062] In another embodiment, the clarification filter may already be connected to the first mechanism at the start of the clarification process (i). In both embodiments, it may be necessary to replace the clarification filter during the clarification process (i) to avoid clogging of the clarification filter and product losses. According to the embodiment, the clarification filter is automatically ejected from the first mechanism and automatically replaced by a new clarification filter. Preferably, based on the measured backpressure of the clarification filter measured by the first pressure sensor, or the pressure difference between the first pressure sensor and the second pressure sensor, and the pumping speed, it can be decided to automatically eject the clarification filter from the first mechanism and automatically replace it with a new clarification filter.

[0063] Then, the clarification filter is preferably clamped so that the upper and lower surfaces of the clarification filter are supported against the internal pressure during filtration (ie, "step (c2)").

[0064] In addition, an air pressure test (i.e., "step (c3)") can be performed on the sample preparation system and the clarification filter before the sample to be clarified is pumped to the clarification filter. In particular, in step (c3), the first valve and the second valve are set to close the flow path, and the pump pumps air into the sample preparation system to reach a preset pressure. According to an embodiment, the preset pressure is 0.1 bar to 4 bar, preferably 0.5 bar to 1 bar. Then, the pressure is monitored for a few seconds. At the end of the air pressure test, the pressure is released by setting the first valve, the second valve and the output valve so that the pressure is released to the waste container via the third line.

[0065] After the optional air pressure test, the sample to be clarified can be provided to the input cup of the sample preparation system (i.e., "step (c4) a"). Alternatively, the sample to be clarified can also be provided to the input cup of the sample preparation system before any one of steps (c1) to (c3) (i.e., "step (c4) b"). Preferably, the pipetting robot can provide the sample to be clarified to the input cup by transferring the sample to be clarified from at least one input container to the input cup of the sample preparation system.

[0066] Then, the sample to be clarified can be pumped from the input cup to the clarifying filter (i.e., "step (c5)"). According to the implementation of step (c5), the sample to be clarified passes through at least the input sample valve, the first liquid sensor, and the first pressure sensor on the way from the input cup to the clarifying filter.

[0067] Furthermore, the sample is clarified by passing the sample through a clarifying filter connected to the first mechanism (hereinafter referred to as "step (c6)").

[0068] The clarified sample can then be directed to an output cup (hereinafter referred to as "step (c7)"). According to an embodiment of step (c7), the clarified sample reaches the output cup from the downstream side of the clarifying filter through at least the second pressure sensor, the second liquid sensor, the third line, the target substance detection sensor, and the output valve.

[0069] When the input cup is empty, the first liquid sensor can detect that the sample to be clarified has been used up (ie, "step (c8)"). Specifically, the first liquid sensor can detect whether the sample to be clarified previously provided to the input cup has passed the position where the first liquid sensor is arranged.

[0070] After step (c8), a flushing step (i.e., "step (c9)") may be automatically performed. In particular, a flushing liquid (such as PBS) may be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Thus, the sample to be clarified still present upstream of the first mechanism may be directed to the first mechanism, and the sample in the clarifying filter may be flushed through the clarifying filter. Preferably, the flushing liquid that has passed through the clarifying filter containing the clarified sample is directed to the output cup.

[0071] After step (c8) and / or step (c9), a step of pumping air (i.e., "step (c10)") is preferably performed. Specifically, air can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. The air pushes the sample to be clarified or the flushing liquid still present in the upper flow field of the clarifying filter, as well as the partially clarified sample or flushing liquid still contained in the clarifying filter through the clarifying filter. The clarified sample thus produced can be directed to the output cup. By applying step (c10), the product recovery rate can be improved. The air pressure used is, for example, in the range of 0.5 bar to 2 bar. Therefore, the air pressure is lower than the bubble point of the clarifying filter, thereby avoiding the generation of foam. Typically, the bubble point of the clarifying filter is greater than 3 bar. After a waiting time of, for example, from 0 seconds to 5 seconds, the air pressure is released by reversing the pump. The measurement of the bubble point is well known to those skilled in the art. For example, the bubble point can be measured according to ISO 2942 or ASTM F316-03.

[0072] Then, the clarified filter can be ejected from the first mechanism (i.e., "step (c11)"). In the case where the first mechanism is the first filter holder and the clarified filter is clamped by the first clamp and the second clamp, the clarified filter is released before being ejected. The ejected clarified filter can be received by the consumables processing device and provided to the waste bin.

[0073] In addition, according to an embodiment, a connection mechanism included by the consumables handling device connects the input line and the first line to provide fluid communication between the input line and the first line so that the liquid path is closed (i.e., "step (c12)"). This can be achieved, for example, by using a dividing wheel as the consumables handling device and rotating the dividing wheel to a second position of the dividing wheel where a through fluid channel is located, which can in turn be connected to the input line and the first line.

[0074] Then, the clarified sample and / or the washing liquid containing the target substance present downstream of the first mechanism can be guided to the output cup by pumping air (ie, "step (c13)").

[0075] In an embodiment of the present invention, the output sample in the output cup may be transferred to at least one output container (ie, "step (c14)"). For example, this step may be performed by a pipetting robot.

[0076] Optionally, a cleaning liquid (such as a buffer, sodium hydroxide and / or water) is pumped through a process fluid input valve disposed upstream of the first mechanism to an output cup through the sample preparation system, and then pumped back from the output cup to the input cup, and then the sample preparation system can be cleaned (i.e., "step (c15)"). The resulting liquid is then directed to a waste container by using a waste pump.

[0077] Purification process (ii)

[0078] The purification filter or purification chromatographic column used in the purification process (ii) is preferably already present in the sample preparation system before the purification process (ii) starts. The purification filter or purification chromatographic column can be provided to the sample preparation system, for example, manually. In another embodiment, the purification filter or purification chromatographic column can be provided to the sample preparation system in an automatic manner.

[0079] According to a preferred embodiment, the purification filter or the purification chromatographic column is in a state of equilibrium. The term "equilibrium state" used herein means that the purification filter or the purification chromatographic column is in a state that allows the binding of target substances. This state can be realized, for example, by making the purification filter or the purification chromatographic column reach a certain pH value (such as 7 or higher) while containing a binding compound. When the purification filter or the purification chromatographic column is not in a state of equilibrium, a balance buffer (such as PBS) can be pumped through the purification filter or the purification chromatographic column. This step can be carried out at any point of the purification process (ii) before the step (p3) described below.

[0080] In addition, according to an embodiment, excessive air can be removed from the upstream side of the purge filter or the purge chromatographic column to avoid the air lock of the purge filter or the purge chromatographic column from being blocked. In particular, buffer (such as PBS) can be pumped to the output cup, and then reverse the pump, so that buffer is pulled to the entrance by the purge filter or the purge chromatographic column from the outlet of the purge filter or the purge chromatographic column, while the negative pressure at the entrance of the purge filter or the purge chromatographic column is monitored and controlled by the second pressure sensor. In an embodiment, pressure is controlled to 0.4 bar absolute pressure or higher. Then, after the balancing step and / or the step of removing excess air from the purge filter or the purge chromatographic column, the input cup, the output cup and the line except the second line of the sample preparation system can be emptied.

[0081] According to an embodiment, at the beginning of the purification process (i), a flow path is provided within the first mechanism, the flow path connecting the line upstream of the first mechanism and the first line of the sample preparation system (ie, "step (p1)"). Preferably, the line upstream of the first mechanism is an input line. In the case where a purification filter is present in the first mechanism, the purification filter is ejected before providing the flow path. In an embodiment of the present invention, the consumables handling device includes a connecting member connecting the input line and the first line, for providing fluid communication between the input line and the first line, thereby providing a flow path within the first mechanism. This can be achieved, for example, by using a dividing wheel as a consumables handling device and rotating the dividing wheel to a second position of the dividing wheel where a through fluid channel is positioned, and the through fluid channel can then be connected to the input line and the first line.

[0082] Optionally, in the case where the sample preparation system includes a target substance detection sensor, the sensor can be zeroed before the sample to be purified is provided to the purification filter or purification chromatographic column (i.e., "step (p2)"). For example, a buffer (such as PBS) can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Preferably, the buffer is pumped through the third line of the sample preparation system without passing through the second line of the sample preparation system.

[0083] Then, the sample to be purified can be provided to the input cup of the sample preparation system (ie, "step (p3)"). Preferably, the pipetting robot can provide the sample to be purified to the input cup by transferring the sample to be purified from at least one input container to the input cup of the sample preparation system.

[0084] Then, the sample to be purified can be pumped in the direction of the purification filter or the purification chromatographic column (i.e., "step (p4)"). According to an embodiment, when the sample to be purified is pumped in the direction of the purification filter or the purification chromatographic column, the first valve and the second valve are switched to bypass the second mechanism to avoid pumping excessive air into the purification filter or the purification chromatographic column, which may result in reduced performance and ultimately lead to air lock blockage of the purification filter or the purification chromatographic column. In an embodiment, when the sample to be purified is detected by the first liquid sensor, the first valve and the second valve are switched to guide the sample to be purified through the purification filter or the purification chromatographic column. Preferably, when the sample to be purified is detected by the second liquid sensor, the first valve and the second valve are switched to guide the sample to be purified through the purification filter or the purification chromatographic column. Therefore, even less air is pumped into the purification filter or the purification chromatographic column. Optionally, after the sample to be purified is detected by the first liquid sensor or the second liquid sensor, a predetermined volume of the sample to be purified can be pumped before switching the first valve and the second valve, and therefore even less air or no air is pumped into the purification filter or the purification chromatographic column.

[0085] When the input cup is empty, the first liquid sensor can detect that the sample to be purified has been used up (ie, "step (p5)"). Specifically, the first liquid sensor can detect whether the sample to be purified previously provided to the input cup has passed the position where the first liquid sensor is arranged.

[0086] Then, a flushing step (i.e., "step (p6)") can be automatically performed. In particular, a flushing liquid (such as PBS) can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Therefore, the sample to be purified still present upstream of the second mechanism can be guided to the second mechanism and purified by passing the sample to be purified through a purification filter or a purification chromatographic column.

[0087] Subsequently, according to one embodiment, the target substance detection sensor included in the sample preparation system can be reset to zero after the flushing step (i.e., "step (p7)"). For example, a buffer (such as an elution buffer) can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Preferably, the buffer is pumped through the third line of the sample preparation system without passing through the second line of the sample preparation system.

[0088] Optionally, after step (p6) and / or step (p7), at least any liquid still contained in the third line of the sample preparation system may be drained (i.e., "step (p8)"). In an embodiment of step (p8), any liquid still present in the input line, the first line, and / or the fourth line of the sample preparation system may be further drained. Thus, air may be introduced into the third line, which may be used to push the sample to the output cup in a subsequent step.

[0089] Then, according to an embodiment, an elution step (i.e., "step (p9)") is performed. Preferably, when a buffer (such as an elution buffer) is pumped in the direction of the purification filter or the purification chromatographic column, the first valve and the second valve are switched to bypass the second mechanism to avoid pumping excessive air into the purification filter or the purification chromatographic column, which may result in performance degradation and ultimately lead to air lock blockage of the purification filter or the purification chromatographic column. In an embodiment, when a buffer (such as an elution buffer) is detected at the first liquid sensor, the first valve and the second valve are switched to guide the buffer through the second line via the purification filter or the purification chromatographic column. Preferably, when a buffer (such as an elution buffer) is detected at the second liquid sensor, the first valve and the second valve are switched to guide the buffer through the second line via the purification filter or the purification chromatographic column. Therefore, even less air is pumped into the purification filter or the purification chromatographic column. Optionally, after a buffer (such as an elution buffer) is detected by the first liquid sensor or the second liquid sensor, a predetermined volume of buffer can be pumped before switching the first valve and the second valve, and therefore even less air or no air is pumped into the purification filter or the purification chromatographic column. The buffer can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. In an embodiment of step (p9), the elution buffer can be a mixture of a first elution buffer and a second elution buffer, wherein the first elution buffer can be provided to the sample preparation system through a first process fluid input valve arranged upstream of the first mechanism, and the second elution buffer can be provided to the sample preparation system through a second process fluid input valve arranged upstream of the first mechanism. Therefore, gradient elution can be performed by controlling the use of the first process fluid input valve and the second process fluid input valve.

[0090] Then, the purified sample can be collected in an output cup (i.e., "step (p10)"). In an embodiment, the purified sample corresponds to a fraction containing the target substance eluted from the purification filter or purification chromatographic column. Specifically, the fraction can be eluted by using an elution buffer in step (p9). According to an embodiment, the concentration of the target substance in the fraction is higher than 0.4 g / L, preferably higher than 0.5 g / L. Therefore, the fraction is suitable for direct analysis by various subsequent assays.

[0091] In the above-mentioned step (p10), the purified sample can be collected by different methods. According to an embodiment, the target substance detection sensor can be used to detect when the target substance passes through the sensor. In another embodiment, the sample can be purified by using a predetermined elution volume. In two embodiments, when a flushing step (step (p6)) has been performed before step (p10), the initial liquid flowing out from the purification filter or the purification chromatographic column is mainly the buffer used in the flushing step, and contains very little target substance, and is therefore preferably directed to a waste container. When the target substance detection sensor detects the target substance or has eluted a predetermined elution volume of an appropriate amount, the output valve is switched to direct the purified sample to the output cup. In a preferred embodiment, the target substance detection sensor is a UV sensor. Then, by using the output from the UV sensor, the volume of the elution buffer pumped through the system, the optical path length of the target substance (such as protein) and the extinction coefficient, the total eluted target substance can be calculated.

[0092] In an embodiment, after sufficient buffer (such as elution buffer) has been pumped in the direction of the purification filter or the purification chromatographic column, the first valve and the second valve can be optionally switched so that air that may be present in the third line of the sample preparation system provided in the previous step (i.e., step (p8)) can be used to direct the purified sample still present in the output line to the output cup (i.e., "step (p11)").

[0093] Preferably, the output sample present in the output cup can be transferred to at least one output container (ie, "step (p12)"). For example, this step can be performed by a pipetting robot.

[0094] Optionally, the sample preparation system may then be cleaned (i.e., "step (p13)") by pumping a cleaning liquid (such as a buffer, sodium hydroxide, and / or water) from a process fluid input valve disposed upstream of the first mechanism to an output cup through the sample preparation system, and then pumping it back from the output cup to the input cup. The resulting liquid is then directed to a waste container by using a waste pump.

[0095] According to an embodiment, if the cleanup filter or the cleanup chromatographic column reaches the end of its useful life, the cleanup filter or the cleanup chromatographic column is removed from the sample preparation system (ie, "step (p14)").

[0096] Combination of clarification process (i) and purification process (ii)

[0097] In a method for preparing a sample using a sample preparation system including a clarification process (i) and a purification process (ii) (hereinafter referred to as a "combined method"), the clarification process (i) (i.e., step (i)) is performed before the purification process (ii) (i.e., step (ii)).

[0098] The purification filter or purification chromatographic column used in the purification process (ii) of the combined method is preferably already present in the sample preparation system before the clarification process (i) begins. The purification filter or purification chromatographic column can be provided to the sample preparation system, for example, manually. In another embodiment, the purification filter or purification chromatographic column can be provided to the sample preparation system in an automatic manner.

[0099] According to a preferred embodiment, the purification filter or the purification chromatographic column is in a state of equilibrium. The term "equilibrium state" used herein means that the purification filter or the purification chromatographic column is in a state that allows the binding of target substances. This state can be achieved, for example, by making the purification filter or the purification chromatographic column reach a certain pH value (such as 7 or higher) while containing a binding compound. In the case where the purification filter or the purification chromatographic column is not in a state of equilibrium, an equilibrium buffer (such as PBS) can be pumped through the purification filter or the purification chromatographic column. This step can be performed at any point of the combined method before step (4) a or step (4) b described below.

[0100] In addition, according to an embodiment, excessive air can be removed from the upstream side of the purge filter or the purge chromatographic column to avoid the air lock of the purge filter or the purge chromatographic column from being blocked. In particular, buffer (such as PBS) can be pumped to the output cup, and then reverse the pump, so that buffer is pulled to the entrance by the purge filter or the purge chromatographic column from the outlet of the purge filter or the purge chromatographic column, while the negative pressure at the entrance of the purge filter or the purge chromatographic column is monitored and controlled by the second pressure sensor. In an embodiment, pressure is controlled to 0.4 bar absolute pressure or higher. Then, after the balancing step and / or the step of removing excess air from the purge filter or the purge chromatographic column, the input cup, the output cup and the line except the second line of the sample preparation system can be emptied.

[0101] According to an embodiment, at the beginning of the clarification process (i) of the combined method, a clarification filter is provided to the first mechanism, wherein the clarification filter is new, i.e., unused (hereinafter referred to as "step (1)"). In said step (1), preferably, the clarification filter is provided from an input stack by a consumables handling device.

[0102] In another embodiment, the clarification filter may already be connected to the first mechanism at the start of the clarification process (i). In both embodiments, it may be necessary to replace the clarification filter during the clarification process (i) to avoid clogging of the clarification filter and product losses. According to the embodiment, the clarification filter is automatically ejected from the first mechanism and automatically replaced by a new clarification filter. Preferably, based on the measured backpressure of the clarification filter measured by the first pressure sensor, or the pressure difference between the first pressure sensor and the second pressure sensor, and the pumping speed, it can be decided to automatically eject the clarification filter from the first mechanism and automatically replace it with a new clarification filter.

[0103] Then, the clarification filter is preferably clamped so that the upper and lower surfaces of the clarification filter are supported against the internal pressure during filtration (ie, "step (c2)").

[0104] In addition, an air pressure test (i.e., "step (3)") can be performed on the sample preparation system and the clarification filter before the sample to be clarified and purified is pumped to the clarification filter. In particular, in step (3), the first valve and the second valve are set to close the flow path, and the pump pumps air into the sample preparation system to reach a preset pressure. According to an embodiment, the preset pressure is 0.1 bar to 4 bar, preferably 0.5 bar to 1 bar. The pressure is then monitored for a few seconds. At the end of the air pressure test, the pressure is released by setting the first valve, the second valve, and the output valve so that the pressure is released to the waste container via the third line.

[0105] After the optional air pressure test, the sample to be clarified and purified can be provided to the input cup of the sample preparation system (i.e., "step (4) a"). Alternatively, the sample to be clarified and purified can also be provided to the input cup of the sample preparation system before any one of steps (1) to (3) (i.e., "step (4) b"). Preferably, the pipetting robot can provide the sample to be clarified and purified into the input cup by transferring the sample to be clarified and purified from at least one input container to the input cup of the sample preparation system.

[0106] Then, the sample to be clarified and purified can be pumped from the input cup to the clarification filter (i.e., "step (5)"). According to the implementation of step (5), the sample to be clarified and purified passes through at least the input sample valve, the first liquid sensor, and the first pressure sensor on the way from the input cup to the clarification filter.

[0107] Furthermore, the sample is clarified by passing the sample through a clarifying filter connected to the first mechanism (hereinafter referred to as "step (6)").

[0108] Then, the clarified sample immediately flows from the clarifying filter to the purification filter or purification chromatography column (i.e., "step (7)"). Therefore, as described in an embodiment of a method for preparing a sample using a sample preparation system, wherein the method includes a clarification process (i) instead of a purification process (ii), the clarified sample coming out of the clarifying filter is not directly directed to an output cup, but is immediately directed to a purification filter or purification chromatography column.

[0109] When the input cup is empty, the first liquid sensor can detect that the sample to be clarified and purified has been used up (ie, "step (8)"). Specifically, the first liquid sensor can detect whether the sample to be clarified and purified previously provided to the input cup has passed the position where the first liquid sensor is arranged.

[0110] After step (8), a flushing step (i.e., "step (9)") can be automatically performed. In particular, a flushing liquid (such as PBS) can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Therefore, the sample to be clarified and purified still existing upstream of the first mechanism can be directed to the first mechanism, and the sample in the clarifying filter can be flushed through the clarifying filter. Preferably, the flushing liquid passing through the clarifying filter containing the clarified sample is directed to the purification filter or the purification chromatographic column. Therefore, the clarified sample that needs to be purified still existing upstream of the second mechanism can be directed to the second mechanism and purified by passing the clarified sample through the purification filter or the purification chromatographic column.

[0111] After step (8) and / or step (9), it is preferred to perform a step of pumping air (i.e., "step (10)"). Specifically, air can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. The air pushes the sample or flushing liquid to be clarified and purified that is still present in the upper flow field of the clarifying filter, as well as the sample or flushing liquid that has been partially clarified but not yet purified that is still contained in the clarifying filter through the clarifying filter. The clarified sample thus produced can be directed to the purification filter or the purification chromatographic column. In addition, the clarified sample that needs to be purified can be pushed through the purification filter or the purification chromatographic column by the step of pumping air. By applying step (10), the product recovery rate can be improved. The air pressure used is, for example, in the range of 0.5 bar to 2 bar. Therefore, the air pressure is lower than the bubble point of the clarifying filter, thereby avoiding the generation of foam. After a waiting time, for example, from 0 seconds to 5 seconds, the air pressure is released by reversing the pump.

[0112] According to an embodiment of the combined method, the method further comprises the following steps in the following order between step (i) and step (ii), i.e. preferably at least after the above step (6):

[0113] (a) Pop-up clarification filter,

[0114] (b) providing a flow path within the first mechanism, the flow path connecting a line upstream of the first mechanism and a first line of the sample preparation system, and

[0115] (c) removing the sample from the line upstream of the first mechanism by directing the sample through a flow path of the sample preparation system, the first line, and the third line.

[0116] In particular, in the above step (a), the clarified filter is ejected from the first mechanism (i.e., "step (11)"). In the case where the first mechanism is the first filter holder and the clarified filter is clamped by the first clamping member and the second clamping member, the clarified filter is released before being ejected. The ejected clarified filter can be received by the consumables processing device and provided to the waste bin.

[0117] In addition, according to an embodiment, the flow path of the above step (b) can be provided by using a connection mechanism included in the consumables handling device, the connection mechanism connecting the input line and the first line, for providing fluid communication between the input line and the first line, so that the liquid path is closed (i.e., "step (12)"). This can be achieved, for example, by using a dividing wheel as the consumables handling device, and rotating the dividing wheel to a second position of the dividing wheel where a through fluid channel is positioned, and the through fluid channel can then be connected to the input line and the first line.

[0118] Then, in order to remove the sample from the line upstream of the first mechanism according to the above-mentioned step (c), a flushing step (i.e., "step (13)") can be performed. In particular, a flushing liquid (such as PBS) can be provided to the sample preparation system through a process fluid input valve arranged upstream of the first mechanism. Preferably, in this embodiment of step (c), the first valve and the second valve are switched to bypass the second mechanism so that the flushing liquid with the unclarified sample can be guided through the flow path provided in step (b), the first line and the third line of the sample preparation system. Therefore, any unclarified sample still present upstream of the first mechanism of the sample preparation system can be removed to avoid contamination and clogging of the purification filter or the purification chromatographic column. In addition, in an embodiment, step (13) can also be optionally used to zero the target substance detection sensor.

[0119] Then, the first valve and the second valve can be switched to guide the flushing liquid through the second line via the purification filter or the purification chromatographic column (i.e., "step (14)"). Therefore, the clarified sample that still needs to be purified, which is present downstream of the first valve and upstream of the second mechanism, can be guided to the second mechanism and purified by passing the clarified sample through the purification filter or the purification chromatographic column.

[0120] In an embodiment, after step (13) and / or step (14), at least any liquid still contained in the third line of the sample preparation system can be drained (i.e., "step (15)"). In an embodiment of step (15), any liquid still present in the input line and / or the first line of the sample preparation system can be further drained. Thus, air can be introduced into the third line, which can be used to push the sample to the output cup in a subsequent step.

[0121] Then, according to an embodiment, an elution step (i.e., "step (16)") is performed. Preferably, when a buffer (such as an elution buffer) is pumped in the direction of the purification filter or the purification chromatographic column, the first valve and the second valve are switched to bypass the second mechanism to avoid pumping excessive air into the purification filter or the purification chromatographic column, which may result in performance degradation and ultimately lead to air lock blockage of the purification filter or the purification chromatographic column. In an embodiment, when a buffer (such as an elution buffer) is detected at the first liquid sensor, the first valve and the second valve are switched to guide the buffer through the second line via the purification filter or the purification chromatographic column. Preferably, when a buffer (such as an elution buffer) is detected at the second liquid sensor, the first valve and the second valve are switched to guide the buffer through the second line via the purification filter or the purification chromatographic column. Therefore, even less air is pumped into the purification filter or the purification chromatographic column. Optionally, after a buffer (such as an elution buffer) is detected by the first liquid sensor or the second liquid sensor, a predetermined volume of buffer can be pumped before switching the first valve and the second valve, and thus even less air or no air is pumped into the purification filter or the purification chromatographic column. The buffer may be provided to the sample preparation system via a process fluid input valve disposed upstream of the first mechanism. In an embodiment of step (16), the elution buffer may be a mixture of a first elution buffer and a second elution buffer, wherein the first elution buffer may be provided to the sample preparation system via a first process fluid input valve disposed upstream of the first mechanism, and the second elution buffer may be provided to the sample preparation system via a second process fluid input valve disposed upstream of the first mechanism. Thus, gradient elution may be performed by controlling the use of the first process fluid input valve and the second process fluid input valve.

[0122] Then, the clarified and purified sample can be collected in an output cup (i.e., "step (17)"). In an embodiment, the clarified and purified sample corresponds to a fraction containing the target substance eluted from the purification filter or purification chromatographic column. Specifically, the fraction can be eluted by using an elution buffer in step (16). According to an embodiment, the concentration of the target substance in the fraction is higher than 0.4 g / L, preferably higher than 0.5 g / L. Therefore, the fraction is suitable for direct analysis by various subsequent assays.

[0123] In the above-mentioned step (17), the clarified and purified samples can be collected by different methods. According to an embodiment, the target substance detection sensor can be used to detect when the target substance passes through the sensor. In another embodiment, the collection of clarified and purified samples can be based on the step of using a predetermined elution volume. In two embodiments, when the rinsing step (step (13)) is performed before step (17), the initial liquid flowing out from the purification filter or the purification chromatographic column is mainly the buffer used in the rinsing step, and contains very little target substance, and is therefore preferably directed to a waste container. When the target substance detection sensor detects the target substance or has eluted an appropriate amount of predetermined elution volume, the output valve is switched to guide the clarified and purified sample to the output cup. In a preferred embodiment, the target substance detection sensor is a UV sensor. Then, by using the output from the UV sensor, the volume of the elution buffer pumped through the system, the optical path length of the target substance (such as protein) and the extinction coefficient, the total eluted target substance can be calculated.

[0124] In an embodiment, after sufficient buffer (such as elution buffer) has been pumped in the direction of the purification filter or purification chromatography column, the first valve and the second valve can be optionally switched so that air that may be present in the third line of the sample preparation system provided in the previous step (i.e., step (15)) can be used to direct the clarified and purified sample still present in the output line to the output cup (i.e., "step (18)").

[0125] Preferably, the output sample present in the output cup can be transferred to at least one output container (ie, "step (19)"). For example, this step can be performed by a pipetting robot.

[0126] Optionally, the sample preparation system may then be cleaned (i.e., "step (20)") by pumping a cleaning liquid (such as a buffer, sodium hydroxide, and / or water) from a process fluid input valve disposed upstream of the first mechanism to an output cup through the sample preparation system, and then pumping it back from the output cup to the input cup. The resulting liquid is then directed to a waste container by using a waste pump.

[0127] According to an embodiment, if the cleanup filter or the cleanup chromatographic column reaches the end of its useful life, the cleanup filter or the cleanup chromatographic column is removed from the sample preparation system (ie, "step (21)"). BRIEF DESCRIPTION OF THE DRAWINGS DETAILED DESCRIPTION

[0128] Figure 1An embodiment of the sample preparation system of the present invention is shown. In particular, the sample preparation system 1 includes: a serial arrangement of a first mechanism 2 and a second mechanism 3, the first mechanism 2 being configured to be connected to a first consumable 2a, and the second mechanism 3 being configured to be connected to a second consumable 3a; a first valve 4 being arranged between the first mechanism 2 and the second mechanism 3; a second valve 5 being arranged downstream of the second mechanism 3; a first line 6 providing fluid communication between the first mechanism 2 and the first valve 4; a second line 7 providing fluid communication between the first valve 4 and the second valve 5 via the second mechanism 3; a third line 8 providing fluid communication directly between the first mechanism 2 and the second valve 4, configured to bypass the second mechanism 3; and a pump 9 configured to move fluid within the sample preparation system 1. The first consumable 2a is connected to the first mechanism 2. The second consumable 3a is connected to the second mechanism 3. The first mechanism 2 is a first filter holder, which includes a first clamp 10a configured to clamp the first consumable 2a on an upper surface (i.e., upstream) and a second clamp 10b configured to clamp the first consumable 2a on a lower surface (i.e., downstream). The sample preparation system 1 also includes an input stack 11 that holds a first consumable 2a. The sample preparation system 1 also includes an input line 12 that provides fluid communication between an input cup 13 disposed upstream of the first mechanism 2 and the first mechanism 2. An input sample valve 14 is located downstream of the input cup 13 and upstream of the first mechanism 2 on the input line 12. In addition, a first liquid sensor 15 is disposed downstream of the input sample valve 14 and upstream of the first mechanism 2. A second liquid sensor 16 is disposed downstream of the first mechanism 2 and upstream of the second mechanism 3. A first pressure sensor 17 is disposed downstream of the first liquid sensor 15 and upstream of the first mechanism 2. A second pressure sensor 18 is disposed downstream of the first mechanism 2 and upstream of the second mechanism 3. The sample preparation system 1 also includes a fourth line 19 that provides fluid communication between the second valve 5 and an output valve 20 disposed downstream of the second valve 5. A target substance detection sensor 21 is disposed downstream of the second valve 5 and upstream of the output valve 20. The sample preparation system 1 also includes an output line 22 that provides fluid communication between the output valve 20 and an output cup 23. Furthermore, the sample preparation system 1 comprises a process fluid input valve 24 arranged upstream of the first mechanism 2. The sample preparation system 1 is connected to an external waste container (not shown) to which waste can be directed by using a waste pump 25.

[0129] Figure 2 (a) and Figure 2 (b) shows a side view of an embodiment of a portion of the sample preparation system of the present invention and an (external) waste bin 26. In particular, a consumables handling device 27 is shown, which is an indexing wheel and is connected to the first mechanism 2 and the input stack 11 holding the first consumables 2a.

[0130] Figure 3 An embodiment of a consumables handling device 27 in the form of an index wheel is shown. The index wheel has an opening 28 in its in-plane direction, which conforms to the shape of the first consumable at a first position of the index wheel. The index wheel has a hole 29 (corresponding to the connection mechanism) in the in-plane direction of the index wheel, and the hole 29 extends from the first surface of the index wheel to the second surface of the index wheel at a second position of the index wheel.

[0131] Figure 4 An embodiment of a pipetting robot (top) and an embodiment of a pipette washing station (bottom) that can be used in the present invention are shown. In particular, the pipetting robot 30 includes a pipette tip 31 connected to a pH electrode 32 and a fluid input valve 33. The pipetting robot 30 also includes a pipette liquid sensor 34 arranged on a pipetting robot line 35, which provides fluid communication between the pipette tip 31, the pH electrode 32 and the fluid input valve 33. In addition, the pipetting robot 30 includes a syringe 36 connected to the pH electrode 32. Figure 4 In the embodiment shown, the pipette washing station 37 has a wall 38. In addition, the pipette washing station 37 has an outlet 39 at its bottom. The outlet 39 is connected to a waste container (not shown) to which liquid can be pumped using a tip washing waste pump 40. Figure 4 Further shown are a storage container 41 for storing, for example, process liquids, an input container 42 and an output container 43 , which can be arranged next to the pipette washing station 37 .

[0132] Reference numerals list

[0133] 1 Sample preparation system

[0134] 2 First institution

[0135] 2a First Consumable

[0136] 3 Second institution

[0137] 3a Second Consumable

[0138] 4 First valve

[0139] 5 Second valve

[0140] 6 Frontline

[0141] 7 Second Line

[0142] 8 Third Line

[0143] 9 Pumps

[0144] 10a First clamping member

[0145] 10b Second clamping member

[0146] 11 Input Heap

[0147] 12 Input Line

[0148] 13 Input Cup

[0149] 14 Input sample valve

[0150] 15 First Liquid Sensor

[0151] 16 Second liquid sensor

[0152] 17 First pressure sensor

[0153] 18 Second pressure sensor

[0154] 19 Fourth Line

[0155] 20 Output valve

[0156] 21 Target substance detection sensor

[0157] 22 Output lines

[0158] 23 Output cup

[0159] 24 Process fluid inlet valve

[0160] 25 Waste Pump

[0161] 26 Waste Bin

[0162] 27 Consumables handling device

[0163] 28 Opening

[0164] 29 holes

[0165] 30 Liquid handling robot

[0166] 31 Pipette Tips

[0167] 32 pH electrode

[0168] 33 Fluid input valve

[0169] 34 Pipette Liquid Sensor

[0170] 35 Liquid Handling Robot Line

[0171] 36 Injection needle

[0172] 37 Pipette Washing Station

[0173] 38 wall

[0174] 39 Exit

[0175] 40 Tip Washing Waste Pump

[0176] 41 Storage Container

[0177] 42 Input Container

[0178] 43 Output Container

Claims

1. A sample preparation system, comprising: a serial arrangement of a first mechanism configured to be connected to a first consumable and a second mechanism configured to be connected to a second consumable; a first valve disposed between the first mechanism and the second mechanism; a second valve disposed downstream of the second mechanism; a first line providing fluid communication between the first mechanism and the first valve; a second line providing fluid communication between the first valve and the second valve via the second mechanism; a third line providing fluid communication directly between the first valve and the second valve, configured to bypass the second mechanism; as well as A pump is configured to move fluid within the sample preparation system.

2. The sample preparation system according to claim 1, in, The first mechanism is configured to be connected to the first consumable and to support the first consumable.

3. The sample preparation system according to claim 1 or 2, in, The second mechanism is configured to be connected to and support the second consumable.

4. The sample preparation system according to any one of claims 1 to 3, in, The first consumable is a first filter.

5. The sample preparation system according to any one of claims 1 to 4, in, The second consumable is a second filter or a chromatography column.

6. The sample preparation system according to any one of claims 1 to 5, in, The first mechanism is a first filter holder.

7. The sample preparation system according to any one of claims 1 to 6, further comprising: A consumables handling device is connected to the first mechanism, and is configured to provide the first consumables to the first mechanism and receive the first consumables from the first mechanism.

8. The sample preparation system according to claim 7, further comprising an input line providing fluid communication between an input cup disposed upstream of the first mechanism and the first mechanism, in, The consumables handling device includes a connection mechanism configured to connect the input line and the first line for providing fluid communication between the input line and the first line.

9. The sample preparation system according to claim 8, Also included is an input sample valve located on the input line downstream of the input cup and upstream of the first mechanism.

10. The sample preparation system according to any one of claims 1 to 9, It also includes a target substance detection sensor, which is arranged downstream of the second mechanism and is configured to detect a target substance.

11. A method for preparing a sample using the sample preparation system according to any one of claims 1 to 10, in, The method comprises at least one of the following steps: (i) clarifying the sample by passing the sample through a first consumable connected to a first mechanism to provide a clarified sample, wherein the first consumable is a clarification filter, and (ii) purifying the sample by passing the sample through a second consumable connected to a second mechanism to provide a purified sample, wherein the second consumable is a purification filter or a purification chromatography column.

12. The method for preparing a sample according to claim 11, in, The method comprises steps (i) and (ii), and step (i) is performed before step (ii), wherein the clarified sample immediately flows from the clarification filter into the purification filter or the purification chromatography column.

13. The method for preparing a sample according to claim 12, in, The method further comprises the following steps performed in the following order between step (i) and step (ii): (a) ejecting the clarification filter, (b) providing a flow path within the first mechanism, the flow path connecting a line upstream of the first mechanism and the first line of the sample preparation system, and (c) removing sample from a line upstream of the first mechanism by directing the sample through the flow path, the first line, and the third line of the sample preparation system.

14. A method for preparing a sample according to any one of claims 11 to 13, in, The sample is a cell culture comprising a protein, The clarification filter is a cell culture clarification filter, and The purification filter is a protein purification filter or a protein purification chromatography column.

15. A method for preparing a sample according to any one of claims 11 to 14, in, In step (i), the clarifying filter is automatically ejected from the first mechanism and automatically replaced by a new clarifying filter.

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