Methods and apparatus for concentrating samples and buffer exchange

Abstract

A microfluidic cartridge and associated apparatus for performing exchange of buffers or concentrating biologics in liquid are disclosed. The cartridge may include a liquid transport mechanism, a sample container, a series of concentration measurement chambers, and air pressure driven valves for flow control. The cartridge may have a port through which the sample is transported to viscosity, static light scattering, and / or dynamic light scattering measuring devices in sequence for measurement of interaction parameter, molecular weight, hydrodynamic radius, opalescence, viscosity, and / or osmolality.

Description

METHODS AND APPARATUS FOR CONCENTRATING SAMPLES AND BUFFER EXCHANGERelated Applications

[0001] This application claims priority to U.S. Provisional Patent Application 63 / 741,010, filed December 31, 2024 and U.S. Provisional Patent Application 63 / 750,624, filed January 28, 2025, both of which are incorporated by reference herein in their entireties.Technical Field

[0002] This application generally relates to methods and apparatus for concentrating or characterizing samples or buffer exchange, and more specifically to methods and apparatus for concentrating or characterizing protein samples or buffer exchange for protein samples.Background

[0003] Proteins, such as monoclonal antibodies (mAb), are produced in various biological or biotechnology processes. Such proteins are sometimes used for therapeutics. Proteins produced by such processes are often filtered and purified. However, proteins produced in such processes are typically at low concentration in a liquid media, which is not suitable for therapeutic uses or various testing purposes. Therefore, these proteins solutions are subjected to a process to exchange the media or buffer or concentrate to achieve a desired concentration. Centrifuge is typically used to exchange the buffer or concentrate protein solutions in a smaller scale. Such methods involve placing a sample in a special centrifuge tube with a filter at its base. The filter is designed in a way that large molecule like proteins remain in the tube while low molecular-weight media such as buffer permeates through the filter by the centrifugal force. After the media is removed, then a new media is added to the tube and the centrifuge step is repeated. After repeating a few cycles of the centrifugation, media removal, and media addition steps, the sample is taken out of the tube and the concentration of protein is measured. The concentration of the protein is typically measured with UV / VIS (ultraviolet-visible) absorption spectroscopy. This whole process takes a series of operator’s intervention. The rotation speed and duration of centrifugation need to be determined empirically. Also, reaching the target concentration of proteins requires numerous trial and error.

[0004] For protein samples having large volumes, tangential flow filtration (TFF) has been adopted since the filtration process with TFF is known to be more efficient in terms of protein063490-5011-WO 1recovery and filtration speed. This filtration process has been adopted in many protein manufacturing processes. While large scale filtration process is required for the commercial production, small scale TFF systems, such as a TFF system that includes an integrated diaphragm pump apparatus for small sample volume to 50 mL, has also been available. Such TFF systems can be used for the buffer exchange and for increasing of protein concentration. However, such systems do not have any means to directly measure the protein concentration except that it is inferred from the measured volume. Various cartridge style TFFs have also been developed and offered for medium size sample volumes in the range of 500 mL - 1,000 mL. Such systems can be also used for the buffer exchange and the concentrating process of protein solutions. Certain conventional microfluid filtration systems contain circuitry designed to concentrate a protein solution and measure concentration and viscosity within the circuitry. However, such systems are not used for buffer exchange and buffer addition if the concentration exceeds the target value. Measuring viscosity with mounted pressure sensors in the circuitry has a significant drawback - the pressure sensors need to measure pressure drop in the viscometer, which provides additional pressure drop caused by the flow of the viscous protein solution to the connected and subsequent fluidic paths. Therefore, the signal to noise ratio can be quite small due to the additional pressure drop. Also, mounting of the pressure sensors creates a roughness on the interior surface, which is known to cause an extra noise because of the flow disturbance. Therefore, the accuracy of the viscosity measurement is typically determined by the ratio of the extra pressure drop to the back pressure. Also, the microfiltration system needs to be cleaned after use as it is not expected to be disposable and to prevent contamination.

[0005] Therefore, there is a need for an apparatus for concentrating proteins and / or exchanging buffers that addresses the challenges and shortcomings of a conventional apparatus as described above.Summary

[0006] Disclosed herein are apparatus, disposable cartridges, and methods of preparing macromolecular samples in liquid phase at any desired concentration in any buffer and excipient media.

[0007] In accordance with some embodiments, an apparatus for concentrating and exchanging buffers of a solution includes a disposable cartridge. The cartridge has: a container holding a sample; optical chambers for concentration measurement of a protein solution; a tangential flow063490-5011-WO 2filtration filter with a valve for controlling the flow resistance of downstream flow paths returning with the container; ports through which air pressure can be supplied to control the valves and the container; and a buffer port to take in the buffer from an external pump. The apparatus also includes a disposable positive displacement syringe for drawing the sample from the container and dispensing the sample to the filtration filter cooperatively with two valves on the opposite sides of the syringe; a first external pump that transports a buffer to the cartridge cooperatively; and a second external pump that operates on the disposable syringe.

[0008] In accordance with some embodiments, an apparatus for concentrating and exchanging buffers of a solution includes a disposable cartridge that includes: two containers that include a first container holding a sample; optical chambers for concentration measurement of a protein solution; a tangential flow filtration filter with a valve to control the flow resistance of downstream flow paths; ports through which air pressure can be supplied to control one or more valves and the containers; a buffer port to take in a buffer from an external pump; and a supply port to exit a processed sample to an external instrument. The apparatus also includes a first external pump that transports a buffer to the cartridge cooperatively.

[0009] In accordance with some embodiments, a microfluidic device for processing a solution containing a sample material includes: one or more substrates defining one or more flow paths; a tangential flow filtration filter fluidically coupled with at least a subset of the one or more flow paths; one or more valves coupled with one or more sample flow paths located downstream from the tangential flow filtration filter for controlling a flow in the one or more sample flow paths located downstream from the tangential flow filtration filter; and one or more optical chambers fluidically coupled with the one or more flow paths for optical measurements of a liquid within the one or more optical chambers.

[0010] In accordance with some embodiments, an apparatus for processing a sample material includes a mount for holding any microfluidic device described herein; and a first pump fluidically coupled with a buffer path for transporting a buffer from the buffer path to the microfluidic device.

[0011] In accordance with some embodiments, a method for concentrating a sample material in a solution includes activating any apparatus described herein. The apparatus is coupled with the microfluidic device in fluidic coupling with the solution containing the sample material. The method also includes applying a pressure to the microfluidic device for transporting the solution063490-5011-WO 3to the tangential flow filtration filter for concentrating the sample material; and performing one or more optical measurements on the solution through the one or more optical chambers of the microfluidic device.

[0012] In accordance with some embodiments, a method for exchanging buffers for a sample material includes activating the apparatus of claim 35 for diafiltration, wherein the apparatus is coupled with the microfluidic device in fluidic coupling with a first solution that includes a first buffer and the sample material; and providing a second buffer distinct from the first buffer to the microfluidic device to provide a second solution that includes the second buffer and the sample material.

[0013] Such apparatus allows characterization of a candidate protein under various conditions with a single loading of sample having a small amount. Such apparatus can be used to facilitate various operations, including formulation optimization of the protein for therapeutic applications.Brief Description of Drawings

[0014] For a better understanding of the various described embodiments, reference should be made to the Description of Embodiments below, in conjunction with the following drawings in which like reference numerals refer to corresponding parts throughout the figures.

[0015] Figure 1 is a schematic drawing of a side view of an apparatus for buffer exchange and solution concentration in accordance with some embodiments.

[0016] Figure 2 is a schematic drawing illustrating top views of the bottom cartridge interface and mount and the high pressure air supply system in accordance with some embodiments.

[0017] Figure 3 is a schematic drawing illustrating a top view of cartridge elements and an apparatus for buffer exchange and concentrating a solution in accordance with some embodiments.

[0018] Figures 4Aand 4B are schematic drawings illustrating a top view and a cross-sectional view of a cartridge in accordance with some embodiments.

[0019] Figure 5A and 5B are schematic drawings illustrating a top view and a cross-sectional view of a transverse flow filter in accordance with some embodiments.

[0020] Figure 6 is a process flow diagram for ultrafiltration operation in accordance with some embodiments.

[0021] Figure 7 is a flow diagram for buffer exchange operation in accordance with some embodiments.063490-5011-WO 4

[0022] Figure 8 is a schematic diagram illustrating transportation of a sample in a cartridge to subsequent characterization devices in accordance with some embodiments.

[0023] Figure 9 is a schematic drawing illustrating a cartridge utilizing the pressure applied to each vial in accordance with some embodiments.Description of Embodiments

[0024] As described herein, conventional apparatus for concentrating proteins and / or exchanging buffers has several shortcomings and challenges. In addition, with such conventional apparatus, the concentration of the protein is typically not known until the end of the filtration process. If the concentration measured at the end of the filtration process is not at a target concentration, then the filtration process may need to be repeated especially when the measured concentration is below the target concentration. If the concentration is higher than the target concentration, then a buffer needs to be added to dilute. Such adjustment of concentrations may involve repeated filtration and concentration measurement operations. These are the drawbacks of conventional methods, which requires continuous attention by operators throughout the concentration process and is not suitable for automation.

[0025] Therefore, there is a need for an apparatus with precise filtration control and verification of the concentration during the filtration processes for fast and efficient operations for concentrating protein. Apparatus described herein provides a protein sample at a specific concentration in a particular buffer. Such “conditioned” sample is then transported to subsequent measurement instruments, such as a dynamic light scattering device, a static light scattering device, melting temperature spectroscopy, and a viscometer for characterization of the protein sample. After the protein sample is characterized, the protein sample is returned to the cartridge so that its concentration can be changed to different concentration. The protein sample can be then transported back to the measurement instruments for further characterization. Such operations can be repeated so that a protein is characterized at different concentrations and in different buffers. As the protein is characterized in multiple buffers (or excipients) and at various concentrations, such information can be used to optimize protein formulation for protein stability, injectability, and efficacy.

[0026] In some embodiments, one end of a microcapillary tube is inserted in a vial holding a sample container (sample vial) at or near the bottom of the liquid in the sample vial. The other end of the tube is connected to the opening end of the embedded microfluidic channel in a063490-5011-WO 5microfluidic cartridge. This input channel is connected to two embedded optical chambers holding the liquid with precision fluidic thickness, the chambers having a flat top and bottom walls that serve to define different precise liquid sample thickness and are made of UV and visibly transparent materials. These two different optical changes are used (and in some configurations located in series) for optical measurement of the concentration of the protein. These two chambers serve as flow chambers so liquid can continuously be sampled during the concentration process. The concentration can be measured based on the absorption of ultraviolet light. The depths of the chambers are different so that a wide dynamic range of concentration can be measured according to Beer’s Law. The exit of the chamber is connected to a positive displacement syringe, which is mounted directly to the cartridge. The plunger of the syringe is connected to a mechanical pumping mechanism so that the plunger can move upward for injection and downward for sample aspiration into the syringe. The aspiration and injection operations are in sync with activation of two fluidic valves located at near the tip of the syringe on opposite sides of a microfluidic circuit loop so that the sample flows properly in the controlled direction. First, the sample is aspirated out of sample vial into the syringe by opening the first valve on the vial side and closing the second valve towards the filter side. After aspiration, the sample is injected to the filter, by closing the first valve from the vial side and opening the second valve to the filter. The exiting flow of the filtered sample is returned to the sample vial through two flow paths connected in parallel. One of the flow paths is low in flow resistance whereas another path is high in resistance. A valve is placed between the outlet of the filter and the low resistance path. With the valve open, low resistance path dominates since a liquid flow readily through a low resistance path, concentration can be finely adjusted since low resistance would lower the filtration rate or flux rate. With the valve closed, the high resistance path is only available and thus the filtration rate is higher, and the concentration is changed faster. Additionally or alternatively, the vial can be pressurized so that the background fluidic pressure over the filter within the cartridge is increased. During the filtration process, the original vial sample and the returning filtered flow, known as the retentate, are mixed with a rotating magnetic stirring element inside the vial at the bottom of the liquid column. This stirring element may have a complex micromolded shape wherein it has paddles for easier rotation of the liquid sample especially at high concentrations. It can also be optionally threaded over the microcapillary input tube at the center of the vial and also is moveable up and down inside the063490-5011-WO 6vial. For simplicity, it can be depicted as a simple dipole magnetic sphere. In addition to the optical concentration measurements discussed above, the sample volume in the vial is also measured in real time to control the concentration to a high degree of precision so that continuous optical measurements are not necessary.

[0027] For the buffer exchange, a separate vial containing a new buffer (buffer vial) is connected to a separate syringe pump. The syringe pump is also equipped with a distribution valve so that the buffer is aspirated from different bottles and dispensed to different paths. Ports of the distribution valve are connected to the buffer bottles and another port of the distribution valve is connected to the cartridge. The pump delivers the buffer out of the bottles to the cartridge for buffer exchange. Numerous buffer bottles can be connected to the distribution valve.

[0028] Once the sample is verified to reach the target concentration, then the sample can be transported to measurement instruments such as dynamic light scattering and static light scattering, melting temperature spectroscopy, and or viscosity instrument. After the measurement is finished, the sample is retrieved back to the cartridge and to the sample vial to concentrate further or exchange the buffers. Repeat this process so that properties of a protein are measured as a function of concentration and in various buffers and excipients to obtain the valuable information such as protein molecule size, molecular weight, protein-protein interaction such as Kd (diffusion interaction parameter) and A2 (second virial coefficient), stability, and the viscosity.

[0029] Figure 1 is a schematic drawing of an apparatus (10) for exchanging a buffer (or media) and / or concentrating a solution (e.g., a solution containing various proteins, oligonucleotides, lipid nanoparticles, etc.) in accordance with some embodiments. In Figure 1, the apparatus (10) includes a first pump module (11), a disposable filter cartridge (24), a sample container (26), containers (16, 17) holding different buffers, a drainage collector (25) collecting and guiding the filtrate or permeate, a syringe (28) (e.g., a disposable zero dead volume syringe) attached to the cartridge (24) (e.g., via syringe adapter (36) shown in Figure 3), a second pump module (27) controlling movement of a plunger of the syringe (28), immersed magnetic ball or stirrer (29), and a cartridge mount (33). In some configurations, a sample is loaded into the sample container (26).

[0030] In some implementations, a buffer (18) (or media) identical to the buffer (or media) of the sample is loaded into the container (16) and another buffer (19) (or media) to exchange is loaded063490-5011-WO 7into the container (17). In some configurations, the buffer (18) is called a first buffer and the buffer (19) is called a second buffer. In some embodiments, the second buffer (19) is distinct from the first buffer (18). In some embodiments, a tube (21) connects the container (16) with a first port (12A) of a distribution valve (12) and a tube (20) connects the container (17) with a second port (12B) of the distribution valve (12). In some embodiments, a tube (23) connects a third port (12C) of the distribution valve (12) with the cartridge (24) and a buffer waste tube (22) connects a fourth port (12D) of the distribution valve (12) with the drainage collector (25). In some embodiments, the drainage collector (25) merges permeate waste from the TFF with buffer waste from syringe (14) into one waste stream. In some embodiments, the syringe (14) is a zerodead-volume syringe. This reduces or eliminates or greatly reduces the carry over of one buffer to another one.

[0031] In some embodiments, the first pump module (11) is coupled with a syringe (14) (and in some cases, the first pump module (11) is also called a syringe pump or a syringe pump module). In some embodiments, the first pump module (11) includes a pump actuator (13) for actuating the syringe (14). In some embodiments, a terminal end (34) of a plunger of the syringe (14) is locked in the pump actuator (13) with a knob (15).

[0032] In some embodiments, the second pump module (27) is coupled with a syringe (and in some cases, the second pump module is also called a second syringe pump or a second syringe pump module). In some embodiments, the second pump module (27) has a structure analogous to that of the first pump module (11).

[0033] While the first port (12A) is selected (e.g., by a computer system (190) or an electronic device with one or more processors (192) and memory (194), executing one or more programs, such as firmware, stored in the memory (194) without manual input or intervention) (or remains activated or open) for the distribution valve (12), aspiration action with the syringe (14) draws the buffer (18) from the container (16). Thereafter, the fourth port (12D) is selected (or activated) for the distribution valve (12), which, in some embodiments, causes the fourth port (12D) to open and the first port (12A) to close. While the fourth port (12D) is selected (or remains activated or open), dispensing operation with the syringe (14) dispenses the buffer (18) in the syringe (14) to the drainage collector (25). Repeating the aspiration and dispense operations (e.g., a few times) fills the distribution valve (12) and the syringe (14) with the buffer (18).063490-5011-WO 8

[0034] After the syringe (14) is filled with the buffer, the third port (12C) is selected (or activated) for the distribution valve (12), which, in some embodiments, causes the third port (12C) to open and other portions, such as the first port (12A) and the fourth port (12D) (and sometimes the second port (12B)) to close. Then the buffer (18) is transported to the cartridge (24) by injection motion of the pump actuator (13) (e.g., dispensing from the syringe (14)). In some implementations, the transported buffer (18) is used to prime the microfluidic channels in the cartridge (24), measure the baseline UV absorption reading with the buffer (18) (e.g., in the dual optical chambers), recover the protein molecules left in the microfluidic channels inside of the cartridge (24) after the filtration process is complete, and / or adjust the concentration of the protein if the concentration is higher than a target concentration.

[0035] If the buffer of the protein needs to be changed from one buffer to another buffer (e.g., from the first buffer (18) to the second buffer (19), for example), the buffer (18) in the syringe (14) and the distribution valve (12) needs to be replaced with buffer (19). This involves (i) dispensing all (or a substantial portion of) the buffer (18) remaining in the syringe (14) to the tube (22) (e.g., through the fourth port ( 12D)), (ii) aspirating the buffer (19) into the syringe (14), and (iii) dispensing the buffer (19) to the tube (22) (e g., through the fourth port (12D)). In some implementations, these operations are repeated (e.g., a few times) until the syringe (14) is filled only with the buffer (19). Thereafter, the buffer (19) is dispensed into the cartridge (24) (and to the sample container (26)) through the tube (23) (e.g., through the third port (12C)).

[0036] In some embodiments, the disposable filter cartridge (24) includes an adapter (35) (or a mount) for the sample container (26) for coupling the sample container (26) to the cartridge (24).

[0037] In some embodiments, the cartridge (24) also includes a syringe (28) (or is coupled with the syringe (28) via a syringe mount). In some embodiments, the syringe (28) is a disposable zero dead volume positive displacement syringe. In some embodiments, the cartridge (24) includes an adapter or a mount for mounting the syringe (28) (instead of including the syringe (28) itself as part of the cartridge (24)).

[0038] In some embodiment, two syringes (both syringe barrels and their associated syringe plungers) are mounted to the cartridge (24). The two syringes can be controlled to reciprocate out of phase such that one syringe is aspirating while another is simultaneously injecting fluid by way of associated valves. This arrangement acts as a continuous pump to increase the dispense rate of the sample to the TFF element and thereby roughly double the filtration speed and half the063490-5011-WO 9required time to reach a desired concentration by the end user. Also, either of these two syringes can be permanently attached to the cartridge (24) without the adapter (36) using laser bonding and be designed for zero dead volume performance with an elastic plunger more capable of working with high viscosity formulations. The syringes can be also bonded in parallel to the cartridge plane such that a user need only place the cartridge on top of the mount (33) without further need for syringe assembly thereby increasing the ease of use. In some embodiments, more than two syringes (e.g., three syringes) are mounted to the cartridge (24).

[0039] Although Figure 1 illustrates several components of the apparatus (10) and the cartridge (24), in some embodiments, the apparatus (10) includes only a subset, less than all, of the components described herein (e.g., the cartridge (24) is not part of the apparatus (10) in some embodiments). In some embodiments, the cartridge (24) includes only a subset, less than all, of the components described herein (e.g., the container (26) or the magnetic stirring element (29) is not part of the cartridge (24) in some embodiments). In some embodiments, the apparatus (10) includes a superset of the components described herein (e.g., the apparatus (10) includes one or more components not described herein). In some embodiments, the cartridge (24) includes a superset of the components described herein (e.g., the cartridge (24) includes one or more components not described herein).

[0040] Figure 2 is a schematic drawing illustrating top views of the cartridge interface / mount (33) and a high pressure air supply system (32) in accordance with some embodiments. The cartridge mount (33) has exits (e.g., 31 A, 3 IB, 31C, 3 ID, 3 IE, and 3 IF) of high-pressure air, which is controlled by the 3 / 2-way solenoid valves (e.g., 32A, 32B, 32C, 32D, 32E, and 32F). In some embodiments, respective exits (e.g., 31 A, 3 IB, 31C, 3 ID, 3 IE, or 3 IF) of the mount (33) are fluidically coupled with corresponding valves (e.g., 32A, 32B, 32C, 32D, 32E, or 32F) via respective pneumatic tubes. In some embodiments, as shown in Figure 2, the solenoid valves (32A) are mounted to an arrayed pneumatic manifold (232) which can then be connected to airtight ports on the cartridge mount cradle 33 via pneumatic tubing (not shown), in some configurations, via drilled passage ways in the mount. In such configurations, the cradle mount may have an underlying manifold structure to direct tubing to and solenoid valve actuators to their respective ports. In some embodiments, exits (31 A through 3 IF) are mated with corresponding ports (38, 39, 40, 41, 42, 43) of the cartridge (24) shown in Figure 3. In some embodiments, the supply system (32) is pneumatically connected with the air exits (31 A through063490-5011-WO 103 IF) that pneumatically control microfluidic valves on the cartridge (e g., valve 32A is fluidically coupled with exit 31 A, valve 32B is fluidically coupled with exit 3 IB, etc.).

[0041] In some embodiments, the top surface of the mount (33) is in direct contact with the bottom of the cartridge (24), as shown in Figure 1. In some embodiments, the cartridge mount cradle (33) also has elastomeric seals around various pneumatic ports (31A-31F) so as to improve airtight sealing of each pneumatic actuation channel. In some embodiments, the apparatus includes a clamp (124) to removably clamp down the top of the cartridge (24). This allows the clamp (124) to counter the lifting force applied on the top of the cartridge (24) when high pressure air is supplied to the channels in the cartridge. In some embodiments, the container (26) includes air-tight seal (which provides better seal than just as liquid tight seal) as the vial itself is pressurized to provide a bias back pressure to the transverse flow fdter and a lifting clamp is often used from below (not shown) to keep its interface (35) airtight and counteract any bias air pressure applied. This can be realized, for example, by a pneumatic lift that pushes on an o-ring seal as part of interface (35) that surrounds any liquid and air pressure ports in and out of the cartridge to the vial.

[0042] Figure 3 is a schematic drawing illustrating a top view of cartridge elements and the apparatus (10) for buffer exchange and concentrating a solution in accordance with some embodiments. In some embodiments, as shown in Figure 3, the cartridge (24) further includes an embedded filter (37), air pressure controlled valves (44, 45, 46, 47, 48), ports (38, 40, 41, 42, 43) through which external high pressure air is supplied to the valves, a port (39) through which air is supplied to the adapter (35) toward the sample container (26), embedded air paths (53, 54, 55, 57, 67) that connect the ports (38, 39, 40, 41, 42, 43) with the valves (44, 45, 46, 47, 48), an embedded air path (56) that connects the port (39) with the adapter (35) for the sample container (26), two optical chambers (49, 50) connected in series for the measurement of concentration of proteins, a port (52) for supply of a buffer, and a port (51) through which the sample is transferred to an external measurement equipment. The port (51) allows the sample to be transferred to the external measurement equipment so that the external measurement equipment may measure viscosity, molecular weight, molecular size, melting temperature, and / or proteinprotein interaction. The valves (44, 45, 46, 47, 48) can be embedded in the cartridge (24) or separately fabricated valves mounted (e.g., via laser bonding) to the cartridge (24).063490-5011-WO 11

[0043] In some configurations, the valves (44, 45, 46, 47, 48) are normally open and become closed only when an air pressure is applied to the connecting ports (38, 40, 41, 42, 43). In some other configurations, the valves (44, 45, 46, 47, 48) are normally closed and become open only when an air pressure is applied to the connecting ports (38, 40, 41, 42, 43). In some configurations, the supplied air pressure ranges from 2 to 6 bar to close the valves. In some embodiments, the regulated air supply is controlled by individual three-way solenoid valves (e.g., 32A through 32F) as shown in Figure 2.

[0044] A fluidic path (59) is designed to be a high flow resistance path so that a liquid flow induces the pressure drop (AP). In some embodiments, the pressure drop (AP) substantially follows the following relationship:AP = R x viscosity x flow ratewhere R is the resistance of the fluidic path (59). A high pressure drop in the path (59) increases the pressure inside of the filter (37), which results in high flux rate of media or buffer through the filter membrane of the filter (37) and thus a fast increase in protein concentration. In comparison, a fluidic path (58) has a lower resistance. When the valve (44) is open, a sample flows through both paths (58, 59). Since the fluidic path (58) is a low resistant path, and most of the sample flows through the path (58) and a smaller pressure drop across both paths is induced. The smaller pressure drop thus reduces the flux rate through the filter and concentration increase is smaller than that with the valve (44) closed. Therefore, by turning the valve (44) on and off, the change rate of concentration can be modulated for precise control.

[0045] In some embodiments, the cartridge (24) also includes filters (61 and 62) in the flow path. In some embodiments, the filter (61) is used to open or close the path into the filter (37). In some embodiments, the filter (62) is used to open or close the path out of the filter (37).

[0046] In some embodiments, the cartridge (24) also includes an embedded path (60). When the sample container (26) is coupled to the adapter (35), the embedded path (60), which is communicatively coupled to the tube (30) shown in Figure 1, is connected to the sample container (26) through the tube (30). This allows a sample to be delivered to the optical chambers (49, 50).

[0047] The two optical chambers (49, 50) are designed to have different depths. This provide different optical absorption path lengths so that wide optical densities corresponding to a wide range of concentration can be measured. In some configurations, the augmented measurable063490-5011-WO 12concentration ranges from 1 mg / mL to 300 mg / mL for proteins. In some embodiments, the depth of the chambers ranges from 0.1 mm to 0.8 mm. In some embodiments, the diameter of the optical chamber ranges from 1 mm to 5 mm. In some embodiments, the diameter is about 2 mm (e.g., between 1 mm and 3 mm). In some embodiments, the diameter is less than 3 mm. In some embodiments, when the cartridge (24) is mounted (e.g., with the mount (33) shown in Figure 2), all the air supply ports (38, 39, 40, 41, 42, 43) are mated leak free with corresponding exits (e.g., 31 A through 3 IF). In some embodiments, those air supply manifolds are connected to the 3 / 2 -way solenoid valves (e.g., 32A through 32F) for each port. In some embodiments, the pressure controller is connected to the external clean and dry air supply. In some embodiments, each solenoid valve is controlled electronically. In some embodiments, when the solenoid valve is on, then air is supplied to the corresponding port on the cartridge, and when the solenoid is off, then the port is open to ambient pressure. In some embodiments, when the solenoid valve is off, then air is supplied to the corresponding port on the cartridge, and when the solenoid is on, then the port is open to ambient pressure.

[0048] Figures 4A and 4B are schematic drawings illustrating a top view and a cross-sectional view of a cartridge in accordance with some embodiments. These figures illustrate how the buffer exchange and concentrating process work, in some embodiments, once the cartridge is mounted and clamped. In some configurations, the cartridge (24) is made of two or more layers of plastics. In some embodiments, the plastics include UV transparent cyclic olefin copolymer (COC) or cyclic olefin polymer (COP). In some embodiments, microfluidic embedded paths (71, 72, 73, 74) and the filter (37) are built at the interface of the two pieces of plastics, as shown in Figure 3B. In some embodiments, the two pieces are diffusion bonded, thermally bonded, or laser welded. A protein sample is loaded into the container (26), which is screwed in the cartridge or magnetically latched to the cartridge through an adapter (35) that is permanently bonded to cartridge (24). In some embodiments, separately fabricated adapter (35) is laser welded. In some embodiments, a disposable syringe (28) is connected to the Luer lock fitting (66) which is permanently laser welded to the cartridge (24) as well. In some configurations, valves (45, 46, 48) are normally open and become closed only when an air pressure is supplied ranging from 2 to 6 bars.063490-5011-WO 13Example 1 : How to concentrate the sample (ultrafiltration).

[0049] First, measure the sample volume in the container (26) after the sample is loaded.Machine vision with a camera (76) or detection of the capacitance change with the level of the sample is used to measure the volume. Open the valve (48) while the rest of valves (44, 45, 46, 47) are closed. The plunger (78) of the syringe (28) is moved all the way upward with the corresponding action of the pump (27). Move the plunger (78) downward to aspirate to withdraw the sample and fill the two optical chambers (49, 50). Measure the concentration of sample. Move the plunger (78) upward after the concentration measurement is finished. Open valves (44, 45, 46, 48) and dispense the buffer (18) in the container (16) to all the paths (e.g., paths 58 and 59 and other paths toward port 52, valves 45 and 48, adapter 36, chambers 49 and 50, and filter 37) to prime the fluidic channels of the cartridge (24) with the valve (47) closed. Measure the UV absorption of the buffer through the optical chambers (49, 50). Then the valves (45, 46, 47) are closed while the valve (48) is kept open. Move the plunger (78) downward to aspirate the sample to the syringe at a controlled speed by the predetermined volume of the sample. The sample flows through the tube (75), the shallower chamber (49), the deeper chamber (50), the microfluidic path (74), the valve (48), and the path (73) in sequence before entering the syringe (28). Close the valve (47, 48) and then open the valve (46). Then move the plunger (78) of the syringe (28) upward to dispense the sample in the syringe (28) through the valve (46), the path (72), the filter (37), both paths (58, 59) to the container (26). During this process, the sample flows through the chamber (64) of the filter and the retentate returns to the container (26) while some of the buffer crosses the filter membrane as a permeate. The permeate is collected in the drainage collector (25). The filtration process involving a transverse flow (79) is known as the tangential flow filtration (TFF). The filter membrane (80) is supported mechanically by a structure (81) with perforation as shown in FIG. 5 A and 5B. The filter retains molecules larger than the cut-off molecular weight and permeates smaller molecular weight. Such a process is known as ultrafiltration. Larger molecules like proteins are retained while small molecules like solvents, salts, sugars, or acids permeate. Polyethersulfone, hydrophilic polyvinylidene fluoride, or regenerated cellulose are used for the membrane (80). A thickness of the filter ranges from 100 to 300 pm. A small magnetic stirring element (29) is continuously rotating at the bottom of the container (26) by the rotating action of an external magnetic bar (77) remotely located underneath or to the side of the sample so that the vial liquid sample is well mixed. Repeat the063490-5011-WO 14above sequence of steps for the next filtration cycle. From the consecutive measurement of the concentration, the effective change of concentration is calculated. From the concentration change, the recovered volume in the sample container (26) is estimated and how much volume of the permeate is estimated. The measured volume with the camera (76) verifies the recovered volume. The difference in the estimated volume and the measured volume is the sample left inside of the cartridge (24). With one round of filtration, the flux rate of the permeate and the change in concentration are obtained under the settings. By varying the injection rate and turning on the valve (44) to adjust the resistance, the change rate of the concentration can be controlled precisely. Also, offset air pressure can be supplied to the container (26) through the air path (56) connected to the port (39). Applying higher pressure than the ambient pressure in the container helps aspirate the sample into the syringe (28) without creating air bubble or cavitation in the sample. Also, applied air pressure serves as an offset pressure, which adds to the flow induced pressure. The additional offset pressure increases the filtration rate of the filter. An additional benefit of the higher offset pressure is that shear sensitive proteins could be filtered at lower injection speed and thus shear rate while the same level of permeability can be achieved. At the end of the cycle, higher level of protein recovery is desired, a small volume of buffer can be transported from the buffer container (16) to the sample container (26) with the first pump module (11): the buffer is transferred to the sample container (26) through the tube (23), the port (52), the valves (45, 46), the filter (37), and the paths (58, 59). This process will recover some of the proteins remaining in these paths. The amount of the volume of the buffer is precisely metered with the first pump module (11). The additional volume of the buffer is accounted for to reach the final concentration. In some cases, proteins at high concentration for a gel-like structure, which prevents further filtration. The concentration at which the proteins form a gel is called a gel point. With the apparatus (10), the gel point can be determined when the protein concentration no longer changes with the filtration operation. Monitoring the flow of the permeate out of the drainage (25) provides additional evidence. This monitoring can be done with a camera by forming an in-line drip rate chamber and using machine vision to count the drip rate. Other flow rate sensors could also be used in some embodiments. Figure 6 is a flow diagram summarizing the process flow for the filtration operation in accordance with some embodiments.Example 2: How to exchange the buffers (diafiltration)063490-5011-WO 15

[0050] Buffer exchange begins with the ultrafiltration process. Once the protein concentration reaches the preset value, then the desired amount of the new buffer (19) in the container (17) is transferred to the cartridge (24) through the port (52), the valve (45), the filter (37), the valve (44), the paths (58, 59) to the container (26) in sequence. The protein solution mixed with the new buffer in the container is then processed for ultrafiltration again. The cycle of transferring the new buffer and ultrafiltration is repeated many times until the amount of original buffer reaches desired maximum fraction. Reaching the lower fraction of original buffer in the final solution requires more cycles. To replace the buffer, typically 10 to 20 times of the original volume of buffer is required. Once the concentration reaches the set value, stop and collect the sample from the sample container (26). Figure 7 is a flow diagram summarizing the process flow for the buffer exchange operation in accordance with some embodiments.

[0051] The cartridge (24) and the apparatus (10) can be used to change the concentration of the protein and change the buffer, and then change the protein concentration again in the new buffer. Therefore, any property measurement of the protein in various buffers and at various protein concentrations can be conducted with a single load of protein solution to the cartridge (24) if the conditioned sample is transported out of the cartridge (24) and transported back to the cartridge (24) after a measurement is finished. Numerous characterizations of proteins in solution require preparation of samples as a function of concentration in each buffer. For example, measurement of protein diffusion interaction parameter, Kd, requires the measurement of diffusivity of a protein at different concentrations in a buffer using the dynamic light scattering technique.Viscosity as a function of concentration is also an important relationship to be obtained for a candidate protein to determine the proper protein concentration for injectability, bottle fill ability, and efficacy. At the early stage of drug development, the available amount of candidate protein is limited and preparing many samples is not an option. Therefore, these measurements have not been performed due to limited sample amount. The cartridge (24) and the apparatus (10) described herein allow the measurement of all these important property measurements with smaller amount of sample. In particular, in some embodiments, the total volume stored in the cartridge with a single syringe port, including the volume inside of the retentate side of the filter, can be designed to be under 50 uL including the microcapillary feed from the container, all fluidic paths, and the fluid stored in the filter. In another embodiment only 100 uL or less of fluid is required to fill the total fluidic volume of the cartridge having two syringe zero dead space063490-5011-WO 16ports to act as a continuous syringe pump having reciprocating motion. As the syringe pump can be easily changed in regards to its total length of its linear motion, this storage can be made vanishingly small, or just a few stepper motor steps for very small samples.

[0052] Figure 8 illustrates the schematic of the arrangement of the cartridge (24) for characterization of samples, such as dynamic and static light scattering and viscosity measurements. When the sample reaches the set concentration, then the sample in the container (26) is transported to one or more testing devices. In some embodiments, first the swiveling arm (90) is positioned to engage with the port (51) of the cartridge (24). Then the valve (47) is open so that the instrument sample feed tube (91) is connected to the port (51) and ready to take in the sample. The syringe (28) aspirates and dispenses the sample to transport to the flow through type optical cell (94). During this process, the air bubble detector (92) detects the presence of air. If air is detected then, dispense the sample appropriately depending on when the air is detected so that the cell (94) is free of air bubble. Once the cell (94) fills in, dynamic and static light scattering measurements start by turning on the light source (95) and taking reading with the detector (96) at various angles. Transmittance at zero angle, scattering at 90 degrees, scattering at 173 degrees, at 30 degrees and other angles are conducted with appropriate type of detectors such as photodiode or avalanche photodiode. Another bubble detector is located between the optical cell (94) and the syringe (97). Using two bubble detectors, flow rate and volume of the sample can be measured simultaneously in addition to the bubble detection. Once the optical measurement is finished, then the sample is loaded into the test syringe (97). Then the viscosity is measured with the zero dead volume viscosity sensor (98). An example of a zero dead volume viscosity sensor (98) is described in US 12,078,582, which is incorporated by reference herein in its entirety. In some implementations, the syringe is 100 pL in volume. The viscosity sensor and the syringe are equipped with the sample retrieval feature: a positive pressure is supplied to the tube (100) with the switching valve (99) connecting the tube (100) with the viscosity sensor (98) while the plunger of the syringe (97) is retrieved. In some embodiments, the side port (104) of the syringe is closed during retrieval. With the retrieval, sample viscosity is measured repeatedly and at different flow rates or shear rates. To measure properties at different temperatures, both optical measurements and viscosity measurement modules are controlled separately with thermoelectric Peltier systems. Once the test is completed for the sample at one concentration in a buffer, the sample is recovered back to the sample container (26) of the cartridge (24). The063490-5011-WO 17switch valve is positioned so that the tube (100) is connected to the viscosity sensor (98). First move the plunger (105) of the syringe backward so that the side port (104) is open while the air is supplied to the tube (100) and the valve (47) is closed. Then the test sample is retrieved back to the syringe (97). Then open the valve (47) and aspirate the sample in the test syringe (97) into the syringe (28) for the predetermined volume and the valves. Once the sample is recovered, the swivel arm is lifted from the port (51) and rotated to a clean position (103) and the switch valve (99) is rotated so that the solvent supply tube (101) is connected to the viscosity sensor (98). The clean position (103) is connected to the waste collector which is not shown. Then the wet parts of the viscosity measurement and the optical measurement modules are cleaned and dried by supplying series of solvents and air to the port coupled with the solvent supply tube (101) if desired. The recovered sample is further conditioned by the ultrafiltration and diafiltration processes so that the concentration is changed, or the buffer is changed. Then the sample is transported back to the optical and viscosity modules for further measurements.

[0053] Alternative design of the cartridge (110) is disclosed in Figure 9. In this design, two sample vials (111, 112) are used. With this configuration, a sample flow from the sample vial (111) to the filter (113) and to the collector vial (112) is controlled mainly by the supplied air pressure difference to each vial. Initially sample is loaded into the vial (111). Increase pressure in the vial (111) so that sample flows from the vial (111) to vial (112) with all the valves in between open. During this time, the concentration of the sample is measured. The sample collected in vial (112) is transported back to the vial (111) by applying higher pressure to the vial (112). During this time, a valve (115) is closed so that fdter (113) is not subject to the same pressure applied to the vial (112) and is isolated. In some embodiments, the valve (115) is closed by pressure (e.g., air pressure) applied through a port (117). Repeat the ultrafdtration process again. The flow rate of the sample and offset pressure is determined by the difference in pressures and the average of the pressures in the vials. The higher the average pressure is, the higher filtration rate or flux rate results. The higher the pressure difference is, the higher the sample flow rate or shear rate results. By adjusting the pressure difference and the average pressure, the filtration flux rate and the shear rate. Transporting the buffer to the vial (111 ) is the same as those previously disclosed with the cartridge illustrated in Figure 2. While use of the zero dead volume positive displacement syringe is preferred, a diaphragm pump can be used instead.063490-5011-WO 18

[0054] In view of the principles and examples described herein, we now turn to certain embodiments.

[0055] In accordance with some embodiments, an apparatus (10) for concentrating and exchanging buffers of a solution includes a disposable cartridge (24).

[0056] The cartridge has: a container (26) holding a sample; optical chambers (49, 50) for concentration measurement of a protein solution; a tangential flow filtration filter (37) with a valve (44) for controlling the flow resistance of downstream flow paths (e.g., returning to the container); ports (e.g., ports 38, 39, 40, 41, 42, 43) through which air pressure can be supplied to control the valves and the container; and a buffer port (52) to take in the buffer from an external pump. The apparatus also includes a disposable positive displacement syringe (28) for drawing the sample from the container and dispensing the sample to the filtration filter cooperatively with two valves on the opposite sides of the syringe.

[0057] The apparatus further includes a first external pump (11) that transports a buffer to the cartridge cooperatively; and a second external pump (27) that operates on the disposable syringe.

[0058] In some embodiments, the valves are open or closed with air pressure.

[0059] In some embodiments, the container is supplied with a regulated air pressure.

[0060] In some embodiments, the cartridge has at least two optical chambers (49, 50) with different depths.

[0061] In some embodiments, the diameter of the optical chambers is under 3 mm. In some embodiments, the diameter of at least one optical chamber is under 3 mm. In some embodiments, the diameter of each optical chamber is under 3 mm. In some embodiments, the diameter of only one optical chamber is under 3 mm.

[0062] In some embodiments, the external pump has a selection valve (12) to select a single buffer from a plurality of buffers.

[0063] In some embodiments, the buffer port is quick connected to the cartridge.

[0064] In some embodiments, the valve (e.g., the valve coupled with the filter) is capable of changing the flow resistance by a factor of 50.

[0065] In some embodiments, the sample volume in the container is measured with machine vision (e.g., collecting an image with the camera (76) and processing the image with a computer device with image-based volume measurement programs).063490-5011-WO 19

[0066] In some embodiments, the apparatus further includes a supply port to transport the sample to an external property measurement device.

[0067] In some embodiments, the supply port is quick connected.

[0068] In some embodiments, the apparatus is interfaced with devices for optical and viscosity measurements through the supply port. For example, microfluidic devices for viscosity measurements described in the international patent application, PCT / US2023 / 069729, fded July 6, 2023, may be used.

[0069] In some embodiments, the optical measurements are dynamic and static light scatterings.

[0070] In some embodiments, the viscosity measurement is a flow through type viscosity measurement.

[0071] In some embodiments, the interface is made through a motorized swiveling arm (90) from one position to another.

[0072] In some embodiments, the measurements are performed under temperature control.

[0073] In some embodiments, the apparatus further includes one or more processors and memory storing one or more programs for execution by the one or more processors. The one or more programs include instructions for determining a melting temperature based on the optical and viscosity measurements.

[0074] In some embodiments, the cartridge is equipped with a diaphragm pump.

[0075] In some embodiments, the apparatus further includes one or more processor and memory storing instructions for determining a gel point for the sample concentration based on optical drip chamber measurement of concentration and flow of permeate in a drainage.

[0076] In some embodiments, the apparatus includes two disposable syringes mounted to increase filtration rate.

[0077] In accordance with some embodiments, an apparatus for concentrating and exchanging buffers of a solution includes a disposable cartridge that includes: two containers that include a first container holding a sample; optical chambers for concentration measurement of a protein solution; a tangential flow filtration filter with a valve to control the flow resistance of downstream flow paths; ports through which air pressure can be supplied to control one or more valves and the containers; a buffer port to take in a buffer from an external pump; and a supply port to exit a processed sample to an external instrument. The apparatus also includes a first external pump that transports a buffer to the cartridge cooperatively.063490-5011-WO 20

[0078] In some embodiments, transport of the sample from the first container to a second container (e.g., drainage collector (25) or a different container) is operated by the difference in pressure and average pressure applied to the two containers.

[0079] In accordance with some embodiments, a microfluidic device for processing a solution containing a sample material includes: one or more substrates (e.g., 69, 70) defining one or more flow paths; a tangential flow filtration filter (37) fluidically coupled with at least a subset of the one or more flow paths; one or more valves (e.g., 44) coupled with one or more sample flow paths located downstream from the tangential flow filtration filter for controlling a flow in the one or more sample flow paths located downstream from the tangential flow filtration filter; and one or more optical chambers (e.g., 49, 50) fluidically coupled with the one or more flow paths for optical measurements of a liquid within the one or more optical chambers.

[0080] In some embodiments, the one or more optical chambers include two or more optical chambers fluidically coupled with the one or more flow paths for optical measurements of the liquid within the one or more optical chambers.

[0081] In some embodiments, the two or more optical chambers include a first optical chamber having a first depth and a second optical chamber with a second depth distinct from the first depth. In some embodiments, the second depth is at least 1.5 times the first depth. In some embodiments, the second depth is at least two times the first depth. In some embodiments, the second depth is at least three times the first depth.

[0082] In some embodiments, the diameter of at respective optical chamber of the one or more optical chambers is under 3 mm.

[0083] In some embodiments, the one or more flow paths include two or more flow paths; and the one or more valves include two or more valves (e.g., 62, 44) coupled with the two or more flow paths (e.g., 58, 59) for controlling a flow resistance for the flow in two or more sample flow paths located downstream from the tangential flow filtration filter.

[0084] In some embodiments, the microfluidic device further includes one or more ports fluidically coupled with the one or more valves for providing air pressure to the one or more valves for activating the one or more valves.

[0085] In some embodiments, the microfluidic device further includes a buffer port for receiving a buffer from outside the microfluidic device.063490-5011-WO 21

[0086] In some embodiments, the tangential flow filtration filter has a thickness between about 100 pm and about 300 gm.

[0087] In some embodiments, the microfluidic device further includes a supply port fluidically coupled with the one or more flow paths for outputting at least a portion of the sample material from the microfluidic device.

[0088] In some embodiments, the microfluidic device further includes a container mount for removably coupling a sample container for storing the solution.

[0089] In some embodiments, the microfluidic device further includes a sample container for storing the solution.

[0090] In some embodiments, the sample material includes proteins.

[0091] In accordance with some embodiments, an apparatus for processing a sample material includes a mount for holding any microfluidic device described herein; and a first pump fluidically coupled with a buffer path for transporting a buffer from the buffer path to the microfluidic device.

[0092] In some embodiments, the first pump includes: a first mount for a positive displacement syringe; and a first actuator for moving a plunger of the first positive displacement syringe.

[0093] In some embodiments, the apparatus further includes a second pump fluidically coupled with a sample path for transporting a solution containing a sample material to the microfluidic device.

[0094] In some embodiments, the second pump includes a second mount for a positive displacement syringe; and a second actuator for moving a plunger of the second positive displacement syringe.

[0095] In some embodiments, the apparatus further includes a first buffer container mount for removably coupling with a first buffer container; and a second buffer container mount for removably coupling with a second buffer container distinct from the first buffer container.

[0096] In some embodiments, the apparatus further includes a buffer valve for fluidically coupling with a first buffer path for the first buffer container at a first time and fluidically coupling with a second buffer path for the second buffer container at a second time mutually exclusive to the first time.

[0097] In accordance with some embodiments, a method for concentrating a sample material in a solution includes activating any apparatus described herein. The apparatus is coupled with the063490-5011-WO 22microfluidic device in fluidic coupling with the solution containing the sample material. The method also includes applying a pressure to the microfluidic device for transporting the solution to the tangential flow filtration filter for concentrating the sample material; and performing one or more optical measurements on the solution through the one or more optical chambers of the microfluidic device.

[0098] In some embodiments, the further includes determining a concentration of the sample material in the microfluidic device based on the one or more optical measurements.

[0099] In some embodiments, the concentration of the sample material is determined while the concentration of the sample material remains in the microfluidic device.

[0100] In some embodiments, the method further includes, in accordance with a determination that the determined concentration is lower than a target concentration, continuing to apply the pressure to the microfluidic device for transporting the solution to the tangential flow filtration filter for further concentrating the sample material.

[0101] In some embodiments, the method further includes, in accordance with a determination that the determined concentration is higher than a target concentration, providing additional buffer the sample material to lower the concentration of the sample material.

[0102] In accordance with some embodiments, a method for exchanging buffers for a sample material includes activating the apparatus of claim 35 for diafiltration, wherein the apparatus is coupled with the microfluidic device in fluidic coupling with a first solution that includes a first buffer and the sample material; and providing a second buffer distinct from the first buffer to the microfluidic device to provide a second solution that includes the second buffer and the sample material.

[0103] In some embodiments, the second solution includes substantially the second buffer instead of the first buffer.

[0104] In some embodiments, the second solution is substantially free from the first solution.

[0105] In some embodiments, a total liquid volume on board inside the cartridge is less than 100 uL.

[0106] The foregoing description, for the purpose of explanation, has been described with reference to specific embodiments. However, the illustrative discussions above are not intended to be exhaustive or to limit the scope of the claims to the precise forms disclosed. Many modifications and variations are possible in view of the above teachings. The embodiments were063490-5011-WO 23chosen and described in order to best explain the principles of the various described embodiments and their practical applications, to thereby enable others skilled in the art to best utilize the principles and the various described embodiments with various modifications as are suited to the particular use contemplated.063490-5011-WO 24

Claims

What is claimed is:

1. An apparatus for concentrating and exchanging buffers of a solution, comprising:a disposable cartridge, the cartridge having:a container holding a sample;optical chambers for concentration measurement of a protein solution;a tangential flow fdtration filter with a valve for controlling the flow resistance of downstream flow paths returning with the container;ports through which air pressure can be supplied to control the valves and the container; anda buffer port to take in the buffer from an external pump;a disposable positive displacement syringe for drawing the sample from the container and dispensing the sample to the filtration filter cooperatively with two valves on the opposite sides of the syringe;a first external pump that transports a buffer to the cartridge cooperatively; and a second external pump that operates on the disposable syringe.

2. The apparatus of claim 1, wherein the valves are open or closed with air pressure.

3. The apparatus of claim 1, wherein the container is supplied with a regulated air pressure.

4. The apparatus of claim 1, wherein the cartridge has at least two optical chambers with different depths.

5. The apparatus of claim 1, wherein the diameter of the optical chambers is under 3 mm.

6. The apparatus of claim 1, wherein the external pump has a selection valve to select a single buffer from a plurality of buffers.

7. The apparatus of claim 1, wherein the buffer port is quick connected to the cartridge.

8. The apparatus of claim 1, wherein the valve is capable of changing the flow resistance by a factor of 50.

9. The apparatus of claim 1, wherein the sample volume in the container is measured with machine vision.

10. The apparatus of claim 1, further comprising a supply port to transport the sample to an external property measurement device.

11. The apparatus of claim 10, wherein the supply port is quick connected.063490-5011-WO 2512. The apparatus of claim 10, wherein the apparatus is interfaced with devices for optical and viscosity measurements through the supply port.

13. The apparatus of claim 14, wherein the optical measurements are dynamic and static light scatterings.

14. The apparatus of claim 14, wherein the viscosity measurement is a flow through type viscosity measurement.

15. The apparatus of claim 14, wherein the interface is made through a motorized swiveling arm from one position to another.

16. The apparatus of claim 14, wherein the measurements are performed under temperature control.

17. The apparatus of claim 14, further comprising one or more processors and memory storing one or more programs for execution by the one or more processors, the one or more programs including instructions for determining a melting temperature based on the optical and viscosity measurements.

18. The apparatus of claim 14, wherein the cartridge is equipped with a diaphragm pump.

19. The apparatus of claim 1, further comprising one or more processor and memory storing instructions for determining a gel point for the sample concentration based on optical drip chamber measurement of concentration and flow of permeate in a drainage.

20. The apparatus of claim 1, including two disposable syringes mounted to increase filtration rate.

21. An apparatus for concentrating and exchanging buffers of a solution, comprising:a disposable cartridge comprising:two containers that include a first container holding a sample;optical chambers for concentration measurement of a protein solution;a tangential flow filtration filter with a valve to control the flow resistance of downstream flow paths;ports through which air pressure can be supplied to control one or more valves and the containers;a buffer port to take in a buffer from an external pump; anda supply port to exit a processed sample to an external instrument;a first external pump that transports a buffer to the cartridge cooperatively.063490-5011-WO 2622. The apparatus of claim 21 , wherein transport of the sample from the first container to the second container is operated by the difference in pressure and average pressure applied to the two containers.

23. A microfluidic device for processing a solution containing a sample material, the microfluidic device comprising:one or more substrates defining one or more flow paths;a tangential flow filtration filter fluidically coupled with at least a subset of the one or more flow paths;one or more valves coupled with one or more sample flow paths located downstream from the tangential flow filtration filter for controlling a flow in the one or more sample flow paths located downstream from the tangential flow filtration filter; andone or more optical chambers fluidically coupled with the one or more flow paths for optical measurements of a liquid within the one or more optical chambers.

24. The microfluidic device of claim 23, wherein:the one or more optical chambers include two or more optical chambers fluidically coupled with the one or more flow paths for optical measurements of the liquid within the one or more optical chambers.

25. The microfluidic device of claim 24, wherein:the two or more optical chambers include a first optical chamber having a first depth and a second optical chamber with a second depth distinct from the first depth.

26. The microfluidic device of claim 23, wherein the diameter of at respective optical chamber of the one or more optical chambers is under 3 mm.

27. The microfluidic device of claim 23, wherein:the one or more flow paths include two or more flow paths; andthe one or more valves include two or more valves coupled with the two or more flow paths for controlling a flow resistance for the flow in two or more sample flow paths located downstream from the tangential flow filtration filter.

28. The microfluidic device of claim 23, further comprising:one or more ports fluidically coupled with the one or more valves for providing air pressure to the one or more valves for activating the one or more valves.

29. The microfluidic device of claim 23, further comprising:063490-5011-WO 27a buffer port for receiving a buffer from outside the microfluidic device.

30. The microfluidic device of claim 23, wherein:the tangential flow filtration filter has a thickness between about 100 pm and about 300 pm.

31. The microfluidic device of claim 23, further comprising:a supply port fluidically coupled with the one or more flow paths for outputting at least a portion of the sample material from the microfluidic device.

32. The microfluidic device of claim 23, further comprising:a container mount for removably coupling a sample container for storing the solution.

33. The microfluidic device of claim 23, further comprising:a sample container for storing the solution.

34. The microfluidic device of claim 23, wherein:the sample material includes proteins.

35. An apparatus for processing a sample material, the apparatus comprising:a mount for holding the microfluidic device of claim 23; anda first pump fluidically coupled with a buffer path for transporting a buffer from the buffer path to the microfluidic device.

36. The apparatus of claim 35, wherein:the first pump includes:a first mount for a positive displacement syringe; anda first actuator for moving a plunger of the first positive displacement syringe.

37. The apparatus of claim 35, further comprising:a second pump fluidically coupled with a sample path for transporting a solution containing a sample material to the microfluidic device.

38. The apparatus of claim 37, wherein:the second pump includes:a second mount for a positive displacement syringe; anda second actuator for moving a plunger of the second positive displacement syringe.

39. The apparatus of claim 35, further comprising:a first buffer container mount for removably coupling with a first buffer container; and063490-5011-WO 28a second buffer container mount for removably coupling with a second buffer container distinct from the first buffer container.

40. The apparatus of claim 39, further comprising:a buffer valve for fluidically coupling with a first buffer path for the first buffer container at a first time and fluidically coupling with a second buffer path for the second buffer container at a second time mutually exclusive to the first time.

41. A method for concentrating a sample material in a solution, the method comprising: activating the apparatus of claim 35, wherein the apparatus is coupled with the microfluidic device in fluidic coupling with the solution containing the sample material;applying a pressure to the microfluidic device for transporting the solution to the tangential flow filtration filter for concentrating the sample material; andperforming one or more optical measurements on the solution through the one or more optical chambers of the microfluidic device.

42. The method of claim 41, further comprising:determining a concentration of the sample material in the microfluidic device based on the one or more optical measurements.

43. The method of claim 42, wherein:the concentration of the sample material is determined while the concentration of the sample material remains in the microfluidic device.

44. The method of claim 42, further comprising:in accordance with a determination that the determined concentration is lower than a target concentration, continuing to apply the pressure to the microfluidic device for transporting the solution to the tangential flow filtration filter for further concentrating the sample material.

45. The method of claim 42, further comprising:in accordance with a determination that the determined concentration is higher than a target concentration, providing additional buffer the sample material to lower the concentration of the sample material.

46. A method for exchanging buffers for a sample material, the method comprising:activating the apparatus of claim 35 for diafiltration, wherein the apparatus is coupled with the microfluidic device in fluidic coupling with a first solution that includes a first buffer and the sample material; and063490-5011-WO 29providing a second buffer distinct from the first buffer to the microfluidic device to provide a second solution that includes the second buffer and the sample material.

47. The method of claim 46, wherein:the second solution includes substantially the second buffer instead of the first buffer.

48. The method of claim 46, wherein:the second solution is substantially free from the first solution.063490-5011-WO 30

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