Systems, devices, and methods for cell processing

Abstract

Cell processing systems, devices, and methods are disclosed herein. [Solution] A system for cell processing may comprise a plurality of instruments, each independently configured to perform one or more cell processing operations on a cartridge, and a robot capable of moving the cartridge between each of the plurality of instruments.

Description

[Technical Field]

[0001] (Cross-reference of related applications)

[0001] This application claims the interests of U.S. Provisional Patent Application No. 62 / 987,745, filed on 10 March 2020, and U.S. Provisional Patent Application No. 63 / 093,038, filed on 16 October 2020. The contents of each of these are included in this application by reference.

[0002]

[0002] The devices, systems, and methods described herein relate to the production of cell products for biomedical use using automated systems. [Background technology]

[0003]

[0003] Cell therapies based on hematopoietic stem cells (HSCs), chimeric antigen receptor (CAR) T cells, NK cells, tumor infiltrating lymphocytes (TILs), T-cell receptors (TCRs), regulatory T cells (Tregs), gamma delta (γδ) T cells, and others rely on the production of cell products. The production of such cell products typically requires multiple cell processing steps. Conventional solutions for producing cell products rely on cumbersome manual operations performed in expensive biosafety cabinets and / or cleanrooms. Skilled laboratory technicians, appropriate sterile enclosures such as cleanroom equipment, and associated protocols and procedures for GMP (Good Manufacturing Practice) production are costly. Many current manufacturing processes use numerous manual reagent preparation and instrument handling steps in the manufacturing protocol, and these processes can take days or even weeks. Even platforms described as automated cell processing in closed systems typically rely on pre-configured sets of instruments and tubing. This limits operational flexibility and does not guarantee the prevention of process failures caused by accidental operator errors / human errors.

[0004]

[0004] Most efforts to automate the production of cell products have focused on automating individual processing steps in the cell therapy manufacturing workflow. Even systems that automate several steps lack comprehensive process flexibility, process robustness, and process scalability. Various embodiments disclosed herein address these and other limitations of previous attempts at automating cell processing. [Overview of the project]

[0005]

[0005] This disclosure generally relates to methods and systems for processing cell products. By processing cell products in cartridges that are moved between instruments, several modifications can achieve one or more advantages over conventional cell manufacturing systems. These advantages include, for example, improved sterility, automation, reduced material costs, reduced labor costs, improved reproducibility, improved reliability, reduced risk of operator error, reduced risk of contamination, improved process flexibility, increased capacity, improved instrument throughput, improved process scalability, and reduced process time. Modifications of this disclosure can reduce the cost of providing a cleanroom environment by including a sterile enclosure and / or can utilize work cells with a smaller footprint than current manufacturing equipment. Furthermore, the modifications disclosed herein may be able to be implemented more quickly and with a reduced risk of cell product loss.

[0006]

[0006] In some variations, the present disclosure provides a system for cell processing. The system comprises a plurality of instruments, each independently configured to perform one or more cell processing operations on a cartridge, and a robot capable of moving the cartridge between each of the plurality of instruments.

[0007]

[0007] In some variations, the system may be housed within a work cell. In some variations, the work cell may be automated. In some variations, multiple instruments may be configured to interact with (interface) a cartridge to perform cell processing operations on the cartridge. In some variations, the system may include a processor. The processor may be configured to control the robot and the multiple instruments.

[0008]

[0008] In some variations, the system may be configured to accept two or more cartridges. In some variations, the system may include a cartridge. In some variations, the cartridge may include multiple modules. In some variations, the cartridge may include a bioreactor module. In some variations, the cartridge may include a cell selection module. In some variations, the cell selection module may include a magnetically activated cell selection module. In some variations, the cartridge may include a classification module. In some variations, the classification module may include a fluorescence-activated cell sorting (FACS) module. In some variations, the cartridge may include an electroporation module. In some variations, the cartridge may include a counterflow centrifugal elutriation (CCE) module.

[0009]

[0009] In some variations, the cartridge may include one or more sterile fluid transfer ports. In some variations, the cartridge may include a fluid transfer bus fluidly coupled to each module. In some variations, the cartridge may include a pump fluidly coupled to the fluid transfer bus.

[0010]

[0010] In some variations, the system may include a pump actuator configured to work in conjunction with the pump. In some variations, the system may include a bioreactor instrument. In some variations, the bioreactor instrument may include multiple slots for a cartridge. In some variations, the system may include a cell selection instrument. In some variations, the cell selection instrument may include a magnetically activated cell selection instrument.

[0011]

[0011] In some variations, the system may include a classification instrument. In some variations, the classification instrument may include a fluorescence-activated cell classification (FACS) instrument. In some variations, the system may include an electroporation instrument. In some variations, the system may include a countercurrent centrifugation (CCE) instrument. In some variations, the system may include a reagent storage container.

[0012]

[0012] In some variations, the cartridge may include a bioreactor module and a selection module. In some variations, the cartridge may include a bioreactor module and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and an electroporation module. In some variations, the cartridge may include a bioreactor module, a selection module, a CCE module, and an electroporation module. In some variations, the cartridge may include a second bioreactor module having an internal volume that is 2 times, 5 times, or 10 times larger than the internal volume of the first bioreactor.

[0013]

[0013] In some variations, the system may include an enclosure. In some variations, the enclosure may include an ISO 7 cleanroom. In some variations, the enclosure may include an ISO 6 cleanroom. In some variations, the enclosure may include an ISO 5 cleanroom. In some variations, the enclosure may include a feedthrough. In some variations, the system may perform automated production of cell products.

[0014]

[0014] In some variations, the present disclosure provides a cartridge for cell processing comprising a liquid transfer bus and a plurality of modules. Each module is fluidically coupled to the liquid transfer bus.

[0015]

[0015] In some variations, the cartridge may include one or more sterile fluid transfer ports. In some variations, the cartridge may include a bioreactor module. In some variations, the cartridge may include a cell selection module. In some variations, the cell selection module may include a magnetically activated cell selection module. In some variations, the cartridge may include a classification module. In some variations, the classification module may include a fluorescence-activated cell classification (FACS) module. In some variations, the cartridge may include an electroporation module. In some variations, the cartridge may include a countercurrent centrifugation (CCE) module.

[0016]

[0016] In some variations, the cartridge may include a mechanoporation module. In some variations, the cartridge may include a second bioreactor module having an internal volume that is 2 times, 5 times, or 10 times larger than the internal volume of the first bioreactor. In some variations, the cartridge may include a bioreactor module, a selection module, and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and an electroporation module. In some variations, the cartridge may include a bioreactor module, a selection module, a CCE module, and an electroporation module.

[0017]

[0017] In some variations, the present disclosure provides a method for processing cells. This method involves moving a cartridge containing cell products between a number of instruments inside a closed automated work cell. The instruments may work in conjunction with the cartridge to perform cell processing steps on the cell products.

[0018]

[0018] In some variations, a cell processing step may be performed on the cell product. In some variations, for each cell product, all cell processing steps in this method are performed in a single cartridge.

[0019]

[0019] In some variations, the cell product can be divided into multiple cell product parts. In some variations, the cell processing step can be performed in parallel on the multiple cell product parts. In some variations, at least two of the multiple cell product parts can be combined.

[0020]

[0020] In some variations, the work cell may include a robot configured to move the cartridge. In some variations, the work cell may include a processor. The processor may be configured to control the robot and the plurality of instruments. In some variations, the work cell may be configured to receive two or more cartridges.

[0021]

[0021] In some variations, the cartridge may include a plurality of modules. In some variations, the cartridge may include a bioreactor module. In some variations, the cartridge may include a cell selection module. In some variations, the cell selection module may include a magnetic-activated cell selection module.

[0022]

[0022] In some variations, the cartridge may include a sorting module. In some variations, the sorting module may include a fluorescence-activated cell sorting (FACS) module. In some variations, the cartridge may include an electroporation module. In some variations, the cartridge may include a countercurrent centrifugal elutriation (CCE) module. In some variations, the cartridge may include one or more sterile fluid transfer ports. In some variations, the cartridge may include a liquid transfer bath fluidly coupled to each module. In some variations, the cartridge may include a pump fluidly coupled to the liquid transfer bath.

[0023]

[0023] In some variations, the work cell may include a pump actuator configured to be interlocked with a pump. In some variations, the work cell may include a bioreactor instrument. In some variations, the bioreactor instrument may include a plurality of slots for the cartridge. In some variations, the method may include performing the cell processing steps in parallel for two or more cartridges.

[0024]

[0024] In some variations, the work cell may include a cell selection device. In some variations, the cell selection device may include a magnetically activated cell selection device.

[0025]

[0025] In some variations, the work cell may include a classification device. In some variations, the classification device may include a fluorescence-activated cell classification (FACS) device. In some variations, the work cell may include an electroporation device. In some variations, the work cell may include a countercurrent centrifugation (CCE) device. In some variations, the work cell may include a reagent storage container.

[0026]

[0026] In some variations, the cartridge may include a bioreactor module and a selection module. In some variations, the cartridge may include a bioreactor module and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and an electroporation module. In some variations, the cartridge may include a bioreactor module, a selection module, a CCE module, and an electroporation module.

[0027]

[0027] In some variations, the work cell may include an enclosure. In some variations, the enclosure may include an ISO 7 cleanroom. In some variations, the enclosure may include an ISO 6 cleanroom. In some variations, the enclosure may include an ISO 5 cleanroom. In some variations, the enclosure may include a feedthrough.

[0028]

[0028] In some variations, this method can carry out automated production of cell products. In some variations, the cell products may include chimeric antigen receptor (CAR) T cell products. In some variations, the cell products may include natural killer (NK) cell products. In some variations, the cell products may include hematopoietic stem cell (HSC) cell products. In some variations, the cell products may include tumor-infiltrating lymphocyte (TIL) cell products. In some variations, the cell products may include regulatory T (Treg) cell products.

[0029]

[0029] In some variations, the present disclosure provides a method for processing a solution containing cell products, which is performed in an automated system. The method comprises one or more cell processing steps, which are performed sequentially in any order, selected from an enrichment step, a concentration step, a buffer exchange step, a formulation step, a washing step, a selection step, a standing step, a growth step, a tissue digestion step, an activation step, a transduction step, a transfection step, and a harvesting step.

[0030]

[0030] In some variations, the concentration step may include concentrating a selected cell population by transporting the solution to the cartridge's CCE module via a liquid transfer bus, operating a robot to move the cartridge to the CCE instrument to engage the CCE module with the CCE instrument, and operating the CCE instrument to concentrate the selected cell population in the solution into the CCE module.

[0031]

[0031] In some variations, the washing step may include washing a selected cell population in the solution by transporting the solution to the cartridge's CCE module via a liquid transfer bus, operating a robot to move the cartridge to the CCE instrument and interlock the CCE module with the CCE instrument, operating the CCE instrument to cause the CCE module to remove the culture medium from the solution, to introduce the culture medium into the solution, and / or replace the culture medium in the solution.

[0032]

[0032] In some variations, the selection step may include transporting the solution to a selection module of the cartridge via a liquid transfer bus, operating a robot to move the cartridge to a selection device to interlock the selection module with the selection device, and operating the selection device to cause the selection module to select the selected cell population in the solution.

[0033]

[0033] In some variations, the classification step may include transporting the solution to the classification module of the cartridge via a liquid transfer bus, operating a robot to move the cartridge to a classification device to engage the classification module with the classification device, and operating the classification device to cause the classification module to classify the cell populations in the solution.

[0034]

[0034] In some variations, the standing step may include transporting the solution from the cartridge to the bioreactor module via a liquid transfer bus, operating a robot to move the cartridge to the bioreactor instrument to engage the bioreactor module with the bioreactor instrument, and operating the bioreactor instrument to maintain the cells in the bioreactor module.

[0035]

[0035] In some variations, the growth step may include growing cells by transporting the solution from the cartridge to the bioreactor module via a liquid transfer bus, operating a robot to move the cartridge to the bioreactor apparatus, linking the bioreactor module with the bioreactor apparatus, and operating the bioreactor apparatus to grow the cells in the solution into the bioreactor module by cell replication.

[0036]

[0036] In some variations, the tissue digestion step may include transporting an enzyme reagent via a liquid transfer bus to a module containing a solution containing tissue, and releasing a selected cell population into the solution by the enzyme reagent digesting the tissue.

[0037]

[0037] In some variations, the activation step may include activating a selected population of cells in a solution by transporting an activation reagent to a module containing a solution of cell products via a liquid transfer bus.

[0038]

[0038] In some variations, the electroporation step may include transporting the solution from the cartridge to the electroporation module via a liquid transfer bus, operating a robot to move the cartridge to the electroporation apparatus to engage the electroporation module with the electroporation apparatus, and operating the electroporation apparatus to have the electroporation module perform electroporation of the selected cell population in the presence of a vector.

[0039]

[0039] In some variations, the transduction step may include transduction of a selected cell population in solution by transporting an effective amount of vector to a module containing a solution containing cell products via a liquid transport bus. In some variations, the filling / completion step may include transporting a formulation solution to a module containing cell products via a liquid transport bus to produce the final cell product, and transporting the final cell product to one or more product collection bags.

[0040]

[0040] In some variations, the method may include manually or automatically sterilizing the cartridge at the feed-through port. In some variations, the method may include manually or automatically introducing one or more of the fluid and cell products into the cartridge via the sterile fluid transfer port. In some variations, the method may include manually or automatically removing the cell products from the cartridge. In some variations, the cell products may include immune cells. In some variations, the steps are, in order, a concentration step, a selection step, an activation step, a transduction step, a proliferation step, and a harvesting step.

[0041]

[0041] In some variations, the immune cells may include genetically modified chimeric antigen receptor T cells. In some variations, the immune cells may include genetically modified T cell receptor (TCR) cells. In some variations, the immune cells may include natural killer (NK) cells. In some variations, the cell product may include hematopoietic stem cells (HSCs). In some variations, the method may include, in order, a concentration step, a selection step, a standing step, a transduction step, and a collection step. In some variations, the cell product may include tumor-infusing lymphocytes (TILs). In some variations, the method may include, in order, a tissue digestion step, a washing step, an activation step, a proliferation step, and a collection step.

[0042]

[0042] A counterflow centrifugal eltriation (CCE) module is also described herein, which includes a conical element having an inner and outer surface fixedly attached to the distal end of a linear member having an inner and outer surface. The proximal end of the linear member is rotatably attached to a pivot point to allow the linear member to be extended, retracted, and rotated.

[0043]

[0043] A work cell is also described herein, comprising an enclosure, a plurality of instruments, each independently configured to perform one or more cell processing operations on a cartridge, and a robot capable of moving the cartridge between each of the plurality of instruments.

[0044]

[0044] In some variations, the enclosure may include an air filtration inlet configured to maintain ISO 7 or higher air quality within the internal zone of the work cell. In some variations, the work cell may be automated. In some variations, the instrument may work in conjunction with the cartridge to perform cell processing operations on the cartridge. In some variations, the work cell may be equipped with a processor. The processor may be configured to control a robot and multiple instruments.

[0045]

[0045] In some variations, the work cell may be configured to accept two or more cartridges. In some variations, the work cell may include a cartridge. In some variations, the cartridge may include multiple modules. In some variations, the cartridge may include a bioreactor module. In some variations, the cartridge may include a cell selection module. In some variations, the cell selection module may include a magnetically activated cell selection module. In some variations, the cartridge may include a classification module.

[0046]

[0046] In some variations, the classification module may include a fluorescence-activated cell classification (FACS) module. In some variations, the cartridge may include an electroporation module. In some variations, the cartridge may include a countercurrent centrifugation (CCE) module. In some variations, the cartridge may include one or more sterile fluid transfer ports. In some variations, the cartridge may include a fluid transfer bus fluidically coupled to each module. In some variations, the cartridge may include a pump fluidically coupled to the fluid transfer bus.

[0047]

[0047] In some variations, the work cell may include a pump actuator configured to work in conjunction with a pump. In some variations, the work cell may include a bioreactor instrument. In some variations, the bioreactor instrument may include multiple slots for a cartridge. In some variations, the work cell may include a cell selection instrument. In some variations, the cell selection instrument may include a magnetically activated cell selection instrument. In some variations, the work cell may include a classification instrument. In some variations, the classification instrument may include a fluorescence-activated cell classification (FACS) instrument. In some variations, the work cell may include an electroporation instrument.

[0048]

[0048] In some variations, the work cell may include a countercurrent centrifugation elutriation (CCE) apparatus. In some variations, the work cell may include a reagent storage container. In some variations, the cartridge may include a bioreactor module and a selection module. In some variations, the cartridge may include a bioreactor module and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and a CCE module. In some variations, the cartridge may include a bioreactor module, a selection module, and an electroporation module. In some variations, the cartridge may include a bioreactor module, a selection module, a CCE module, and an electroporation module. In some variations, the cartridge may include a second bioreactor module having an internal volume that is 2 times, 5 times, or 10 times larger than the internal volume of the first bioreactor. In some variations, the enclosure may include a feedthrough. In some variations, the work cell may perform automated production of cell products. In some variations, the system may include multiple bioreactor apparatuses. Each bioreactor apparatus may be configured to accept a single cartridge.

[0049]

[0049] A rotor is also described herein, comprising a first side including a first fluid tube, a second side opposite to the first side including a second fluid tube, and a cone coupled between the first fluid tube and the second fluid tube.

[0050]

[0050] In some variations, the cone may include a biconical shape. In some variations, the bicone may comprise a first cone including a first base and a second cone including a second base. The first base may face the second base. In some variations, the rotor may include a magnetic portion. In some variations, the rotor may define an axis of rotation. In some variations, at least a portion of the first fluid tube and at least a portion of the second fluid tube may extend parallel to the axis of rotation. In some variations, at least a portion of the first fluid tube and at least a portion of the second fluid tube may be coaxial.

[0051]

[0051] In some variations, the cone may contain a volume of about 10 mL to about 40 mL. In some variations, the cone may have a cone angle of about 30 degrees to about 60 degrees. In some variations, at least a portion of the rotor may be optically transparent. In some variations, the rotor may have an asymmetric shape. In some variations, the first part may be conical and the second part may have a paddle shape.

[0052]

[0052] In some variations, the cartridge for cell processing may comprise a liquid transfer bath and a plurality of modules. Each module may be fluidically coupled to the liquid transfer bath. The cartridge may comprise a countercurrent centrifugal ertriation (CCE) module including the rotor described herein.

[0053]

[0053] A rotor comprising a first fluid tube and a first cone coupled to the first fluid tube is also described herein. The first cone may contain a first volume. A second fluid tube may be coupled to the first cone. A second cone may be coupled to the second fluid tube. The second cone may contain a second volume greater than the first volume. A third fluid tube may be coupled to the second cone.

[0054]

[0054] In some variations, the first cone may include a first bicone, and the second cone may include a second bicone. In some variations, the first bicone may include a third cone including a first base and a fourth cone including a second base. The first base may face the second base. The second bicone may include a fifth cone including a third base and a sixth cone including a fourth base. The third base may face the fourth base.

[0055]

[0055] In some variations, the rotor may have a magnetic portion. In some variations, at least a portion of the rotor may be optically transparent. In some variations, the first fluid tube may include an inlet and the third fluid tube may include an outlet.

[0056]

[0056] The present invention also describes a system for cell processing comprising a cartridge having a housing that includes a rotor configured to separate cells from a fluid, and an instrument that includes a magnet configured to magnetically rotate the rotor in conjunction with the cartridge.

[0057]

[0057] In some variations, the cartridge may be configured to move between multiple devices. In some variations, an air gap may exist between the housing and the magnet. In some variations, the housing may house a rotor. In some variations, the housing may include consumable components and the magnet may include durable components.

[0058]

[0058] In some variations, the magnets may be releasably coupled to the housing. In some variations, the magnets may be configured to move relative to the housing. In some variations, the isolated cells may include a first size and a first density, and the unisolated cells in the fluid may include a second size and a second density different from the first size and first density. A cartridge for cell processing comprising a liquid transfer bus and a plurality of modules is also described herein. Each module may be fluidically coupled to the liquid transfer bus. The cartridge may comprise a countercurrent centrifugal ertriation (CCE) module including the rotor described herein.

[0059]

[0059] A countercurrent centrifugal ertriation (CCE) method is also described herein, which includes moving a rotor that defines the axis of rotation toward a magnet, flowing a fluid through the rotor, and magnetically rotating the rotor about the axis of rotation using a magnet while the fluid is flowing through the rotor.

[0060]

[0060] In some variations, an optical sensor can be used to generate image data of one or more of the fluid and particles in the rotor. Based at least partially on the image data, one or more of the rotor's rotational speed and the fluid flow rate can be selected.

[0061]

[0061] In some variations, one or more of the fluid and cells can be illuminated using the illumination source. In some variations, the method may include moving the rotor away from the magnet. In some variations, the method may include moving the rotor toward the illumination source and optical sensor, and moving the rotor away from the illumination source and optical sensor.

[0062]

[0062] In some variations, moving the rotor may involve using a robot to move the magnet forward and backward relative to the rotor. In some variations, rotating the rotor may involve a rotational speed of up to 6,000 RPM. In some variations, flowing the fluid may involve a flow rate of up to approximately 150 ml / min while the rotor is rotating.

[0063]

[0063] A method for magnetically activated cell selection is also described herein, which includes flowing a fluid containing input cells into a flow cell. The cell set may be labeled with a magnetically activated cell selection (MACS) reagent. The cell set can be magnetically attracted to a magnetic array for a dwell time. After the dwell time, the cell set can be flowed out of the flow cell.

[0064]

[0064] In some variations, the method may include culturing the MACS reagent with input cells in order to label the cell set with the MACS reagent. In some variations, culturing the MACS reagent may involve temperatures from about 1°C to about 10°C. In some variations, flowing the cell set out of the flow cell may include flowing gas through the flow cell. In some variations, after a residence time, the method may include flowing a fluid without the cell set out of the flow cell. In some variations, the residence time may be at least about 1 minute. In some variations, the magnet array may be located outside the flow cell. In some variations, the method may include moving the magnet array relative to the flow cell. In some variations, moving the magnet array may include moving the magnet array away from the flow cell to facilitate the outflow of the cell set from the flow cell. In some variations, the longitudinal axis of the flow cell may be perpendicular to the ground. In some variations, beads may not be present in the flow cell.

[0065]

[0065] A magnetically activated cell selection (MACS) module is also described, comprising a flow cell containing an elongated cavity having a cavity height. The magnet array may include multiple magnets. Each magnet may be spaced apart by a separation distance. The ratio of cavity height to separation distance may be approximately 20:1 to approximately 1:20.

[0066]

[0066] In some variations, the flow cell may include a set of linear channels. The linear channels include a first channel parallel to a second channel and a third channel in fluid communication with each of the first and second channels. In some variations, the first channel may include a first cavity height, and the second channel may include a second cavity height. The ratio of the first cavity height to the second cavity height may range from about 1:1 to about 3:7. In some variations, the ratio of the length of the third channel to the diameter of the third channel may range from about 2:1 to about 6:1.

[0067]

[0067] In some modifications, a first fluid tube can be connected to the inlet and outlet of the flow cell. The first fluid tube may be configured to receive a cell set from the flow cell. A second fluid tube can be connected to the inlet and outlet of the flow cell. The second fluid tube may be configured to receive a fluid from the flow cell that does not contain a cell set.

[0068]

[0068] In some variations, the cartridge for cell processing may comprise a liquid transfer bus and a plurality of modules. Each module may be fluidically coupled to the liquid transfer bus. The cartridge may comprise the magnetically activated cell selection (MACS) module described herein.

[0069]

[0069] A system for cell processing is also described herein, comprising a cartridge including a rotor configured for countercurrent centrifugation of cells in a fluid. A first magnet may be configured to magnetically rotate the rotor and to separate cells from the fluid in the rotor. The cartridge may further include a flow cell configured to communicate with the rotor and to receive cells from the rotor. A second magnet may be configured to magnetically separate cells in the flow cell.

[0070]

[0070] In some variations, the illumination source may be configured to illuminate the cells. The optical sensor may be configured to generate image data corresponding to the cells. In some variations, the system may include one or more of the following: an oxygen deficiency sensor, a leak sensor, an inertia sensor, a pressure sensor, and a bubble sensor. In some variations, the system may include one or more valves and pumps.

[0071]

[0071] In some variations, the separated cells may have a first size and a first density, and the unseparated cells of the fluid may have a second size and a second density that is different from the first size and first density.

[0072]

[0072] The electroporation module also described herein comprises a fluid tube configured to receive a first fluid containing cells and a second fluid, an electrode set coupled to the fluid tube, a pump coupled to the fluid tube, and a controller including a processor and memory. The controller may be configured to generate a first signal to introduce the first fluid into the fluid tube using the pump, to generate a second signal to introduce the second fluid into the fluid tube so that the second fluid separates the first fluid from the third fluid, and to generate an electroporation signal to perform electroporation of cells in the fluid tube using the electrode set.

[0073]

[0073] In some variations, the second fluid may include gas or oil. In some variations, the controller may be configured to generate a third signal to introduce a third fluid into the fluid tube. The third fluid is separated from the first fluid by the second fluid.

[0074]

[0074] In some variations, the cartridge for cell processing may comprise a liquid transfer bus and a plurality of modules. Each module may be fluidically coupled to the liquid transfer bus. The cartridge may comprise the electroporation module described herein.

[0075]

[0075] A method for performing cell electroporation is also described herein, which includes receiving a first fluid containing cells into a fluid tube, receiving a second fluid into the fluid tube so as to separate the first fluid from a third fluid, and performing cell electroporation by applying an electroporation signal to the first fluid.

[0076]

[0076] In some variations, the method may include receiving a third fluid, separated from the first fluid by a second fluid, into a fluid tube. In some variations, when an electroporation signal is applied, the first fluid may be substantially static.

[0077]

[0077] A method for performing electroporation of cells is also described herein, which includes receiving a first fluid containing cells into a fluid tube, applying a resistance measurement signal to the first fluid using an electrode set, measuring the resistance between the first fluid and the electrode set, and applying an electroporation signal to the first fluid based on the measured resistance.

[0078]

[0078] In some variations, the method may include receiving a second fluid containing a gas into a fluid tube before applying an electroporation signal to the fluid. The first fluid is separated from the third fluid by the second fluid.

[0079]

[0079] A bioreactor comprising an enclosure including a bottom, a top, and at least one side wall is also described herein. A gas permeable membrane can be bonded to the bottom and one or more of the side walls of the enclosure.

[0080]

[0080] In some variations, the enclosure may include one or more nesting surfaces curved about the longitudinal axis of the enclosure. In some variations, the one or more nesting surfaces may include a set of concentric toroids. In some variations, the enclosure may include an annular shape. In some variations, the enclosure may include a first chamber having a first volume and a second chamber having a second volume. The first chamber is separated from the second chamber, and the first volume is smaller than the second volume. In some variations, the enclosure may include columns extending along the longitudinal axis of the enclosure. In some variations, a cavity may exist between the enclosure and the gas permeable membrane. In some variations, the gas permeable membrane may extend along the bottom and side walls of the enclosure. In some variations, the outer surface of the gas permeable membrane may include one or more protrusions.

[0081]

[0081] In some variations, the bottom of the gas permeable membrane may include an angle of about 3 to about 10 degrees with respect to the bottom of the enclosure. In some variations, the gas permeable membrane may include a curved surface. In some variations, the gas permeable membrane may include a set of patterned curved surfaces. In some variations, the set of patterned curved surfaces may have a radius of curvature of about 50 mm to about 500 mm.

[0082]

[0082] In some variations, the cartridge for cell processing may comprise a liquid transfer bath and a plurality of modules. Each module may be fluidically coupled to the liquid transfer bath. The cartridge may comprise the bioreactor modules described herein. In some variations, the system for cell processing may comprise the cartridge described herein and further comprise a bioreactor instrument configured to work in conjunction with the cartridge. The bioreactor instrument may comprise a stirrer configured to be coupled to the bioreactor. The stirrer may be configured to agitate a cell medium containing cells. In some variations, a fluid connector may be configured to couple the bioreactor to the liquid transfer bath. The fluid connector may comprise a foldable sidewall. In some variations, the system may comprise a temperature controller coupled to the bioreactor. In some variations, the system may comprise a gas regulator coupled to the bioreactor.

[0083]

[0083] A fluid connector is also described herein, comprising a first connector including a first proximal end configured to couple with a first fluid device and a first distal end including a first port. The second connector may include a second proximal end configured to couple with a second fluid device and a second distal end including a second port configured to couple with the first port. The first distal end may include a first lumen, and the second distal end may include a second lumen. One of the first valve and the second valve may be configured to move in parallel within the first lumen and the second lumen.

[0084]

[0084] In some variations, the first valve and the second valve may be configured to transition from a closed configuration to an open configuration only when the first valve is coupled with the second valve. In some variations, the first port and the second port may be configured to transition between an open configuration and a closed configuration. In some variations, the first connector may include a first port actuator, and / or the second connector may include a second port actuator. In some variations, the second port may be coupled to the first port to define a chamber.

[0085]

[0085] In some variations, one or more of the first and second connectors may include a sterilizer port configured to connect to a sterilizer source. When the second port is connected to the first port, the sterilizer port may be configured to be in fluid communication with the first distal end and the second distal end.

[0086]

[0086] In some variations, the chamber may be configured to receive one or more of a fluid and a sterilizer from the sterilizer port. In some variations, the sterilizer port may be configured to receive a sterilizer so that the sterilizer sterilizes the first and second connectors.

[0087]

[0087] In some variations, the first connector may include a first valve, and the second connector may include a second valve configured to be coupled to the first valve. In some variations, the first seal may include a first port coupled to a second port, and the second seal may include a first valve coupled to a second valve. In some variations, the sterilizer may include one or more of vaporized hydrogen peroxide and ethylene oxide.

[0088]

[0088] In some variations, the fluid connector may include one or more robot engagement mechanisms. In some variations, the first connector may include a first alignment mechanism, and the second connector may include a second alignment mechanism configured to be coupled to the first alignment mechanism in a predetermined axial rotation configuration. In some variations, one or more of the first and second fluid devices may include a fixture.

[0089]

[0089] In some variations, the system may further include a robot configured to operate the fluid connector, and a controller including memory and a processor. The controller may be coupled to the robot. The controller may be configured to generate a first port signal for coupling a first port to a second port using a robot arm. In some variations, the controller may be configured to generate a first valve signal for translating a first valve relative to a second valve using a robot arm, and further to generate a second valve signal for moving the first and second valves to an open configuration. In some variations, the controller may be configured to generate a second port signal for uncoupling a first port from a second port. Sterility of the fluid connector may be maintained before coupling the first port to the second port and after uncoupling the first port from the second port.

[0090]

[0090] In some variations, a fluid pump can be coupled to the sterilizer source. The controller may be configured to generate a first fluid pump signal for circulating the fluid into the chamber via the sterilizer port. In some variations, the controller may be configured to generate a second fluid pump signal for circulating the sterilizer into the chamber via the sterilizer port, at least to sterilize the chamber.

[0091]

[0091] In some variations, the controller may be configured to generate a third fluid pump signal for removing the sterilizer from the chamber. In some variations, the controller may be configured to generate a thermal sterilization signal for thermally sterilizing the fluid connector. In some variations, the controller may be configured to generate a radiation sterilization signal for sterilizing the fluid connector using radiation. In some variations, the robot may be configured to connect the fluid connector between at least two of a plurality of instruments and cartridges.

[0092]

[0092] In some variations, the fluid connector may further include a controller coupled to a robot, which includes memory and a processor. The controller may be configured to generate a port signal for coupling a first port to a second port using a robot arm, to generate a first valve signal for translating a first valve relative to a second valve using a robot arm, and to generate a second valve signal for moving the first and second valves to an open configuration.

[0093]

[0093] A non-temporary computer-readable medium for converting user-defined cell processing operations into cell processing steps to be executed by an automated cell processing system is also described herein. The non-temporary computer-readable medium may include stored instructions. When executed on a processor, these instructions may include the steps of receiving an ordered input list of cell processing operations and executing a conversion model on the ordered input list to generate an ordered output list of cell processing steps that can be executed by the system.

[0094]

[0094] In some variations, an ordered output list can be performed by the system to control the robot to move one or more cartridges, each containing cell products, between instruments, and to control the instruments to perform cell processing steps for each cell product.

[0095]

[0095] In some variations, the method may include receiving one or more sets of cell processing parameters. Each set of cell processing parameters is associated with one of the cell processing operations, and each set of cell processing parameters defines the characteristics of the cell processing step performed by the instrument in the cell processing step. In some variations, the transformation model may include constraints on an ordered output list determined by the configuration of the automated cell processing system. In some variations, the constraints may include information about the configuration of the automated cell processing system. In some variations, the constraints may include one or more of the following: the type and / or number of instruments, the type and / or number of modules on the cartridge, the type and number of storage units on the cartridge, the type and / or number of sterile fluid transfer ports on the cartridge, and the number and location of fluid paths between the modules, storage units, and sterile fluid transfer ports on the cartridge.

[0096]

[0096] In some variations, the step may further include receiving a set of two or more ordered input lists of cell processing operations to be performed in two or more cartridges in an automated cell processing system, and running a conversion model on the set of ordered input lists to generate an ordered output list of cell processing steps. The ordered output list may be run by the system to control a robot to move two or more cartridges, each containing cell products, between instruments, and to control the instruments to perform cell processing steps for each cell product in each cartridge.

[0097]

[0097] In some variations, the automated cell processing system may include a non-temporary computer-readable medium as described in any of the preceding claims.

[0098]

[0098] In some variations, a computer-implemented method for translating user-defined cell processing operations into cell processing steps to be executed by the processor of an automated cell processing system may include receiving an ordered input list of cell processing operations and running a translation model on the ordered input list to generate an ordered output list of cell processing steps that can be executed by the system.

[0099]

[0099] In some variations, the method may include controlling a robot to move one or more cartridges, each containing cell products, between instruments, and controlling the instruments to perform a cell processing step for each cell product.

[0100]

[0100] In some variations, the method may include receiving one or more sets of cell processing parameters. Each set of cell processing parameters is associated with one of the cell processing operations, and each set of cell processing parameters defines the characteristics of the cell processing step performed by the instrument in the cell processing step. In some variations, the transformation model may include constraints on an ordered output list determined by the configuration of the automated cell processing system. In some variations, the constraints may include information about the configuration of the automated cell processing system.

[0101]

[0101] In some variations, the constraints may include one or more of the following: the type and / or number of instruments, the type and / or number of modules on the cartridge, the type and number of storage units on the cartridge, the type and / or number of sterile fluid transfer ports on the cartridge, and the number and location of fluid paths between the modules, storage units, and sterile fluid transfer ports on the cartridge.

[0102]

[0102] In some variations, the method may include receiving a set of two or more ordered input lists of cell processing operations to be performed in two or more cartridges in an automated cell processing system; running a transformation model on the set of ordered input lists to generate an ordered output list of cell processing steps; controlling a robot to move two or more cartridges, each containing cell products, between instruments; and controlling the instruments to perform cell processing steps for each cell product in each cartridge.

[0103]

[0103] Further modifications, features, and advantages of the present invention will become apparent from the following detailed description and through the implementation of the present invention. [Brief explanation of the drawing]

[0104] [Figure 1A]

[0104] This is a block diagram of an exemplary variation of the cell processing system. [Figure 1B]

[0105] This is a block diagram of an exemplary variation of a cartridge. [Figure 2A]

[0106] This is a block diagram of an exemplary variation of a cell processing system. [Figure 2B]

[0106] This is a perspective view of an exemplary variation of a work cell in a cell processing system. [Figure 2C]

[0106] This is a perspective view of an exemplary variation of the work cell and cartridge of the cell processing system. [Figure 2D]

[0106] This is a block diagram of an exemplary variation of the cell processing system. [Figure 2E]

[0106] This is a block diagram of another exemplary variation of the cell processing system. [Figure 3]

[0107] This is a block diagram of another exemplary variation of the cell processing system. [Figure 4A]

[0108] A perspective view of another exemplary variation of the cell processing system. [Figure 4B]

[0108] Another perspective view of another exemplary variation of the cell processing system. [Figure 5]

[0109] A perspective view of another exemplary variation of the cell processing system. [Figure 6]

[0110] This is a schematic diagram illustrating an example of a cartridge modification. [Figure 7]

[0111] This is a schematic diagram of another exemplary variation of the cartridge. [Figure 8A]

[0112] This is a side view of an exemplary variation of the cartridge. [Figure 8B]

[0112] This is a top view of an example of a modified cartridge. [Figure 8C]

[0112] This is a side view of an exemplary deformation of the cartridge. [Figure 8D]

[0112] This is a perspective view of an exemplary variation of the cartridge. [Figure 9]

[0113] This is a side cross-sectional view of an exemplary deformation of a cartridge. [Figure 10A]

[0114] Exemplary variations of rotary valves and actuators are shown. [Figure 10B]

[0114] An exemplary variation of a rotary valve docked to an actuator is shown. [Figure 11A]

[0115] This is a perspective view of an exemplary variation of a cartridge including an extended configuration CCE module. [Figure 11B]

[0115] This is a side cross-sectional view of an exemplary variation of a CCE module in a storage configuration. [Figure 11C]

[0115] Side cross-sectional view of an exemplary variation of the extended configuration CCE module. [Figure 12A]

[0116] This is a perspective view of an exemplary variation of a magnetically activated cell classification (MACS) instrument that includes a magnet in an ON configuration. [Figure 12B]

[0116] A perspective view of an exemplary variation of a MACS device including a magnet in an OFF configuration. [Figure 13A]

[0117] This is a perspective view of an exemplary variation of a cartridge and bioreactor device. [Figure 13B]

[0117] A perspective view of an exemplary variation of a cartridge coupled to a bioreactor device. [Figure 14]

[0118] This is a perspective view of an exemplary variation of a bioreactor device, including a set of cartridges and cavities configured to receive these cartridges. [Figure 15]

[0119] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 16A]

[0120] This is a schematic diagram illustrating an exemplary variation of a fluid connector. [Figure 16B]

[0120] This is a detailed schematic diagram of the fluid connector shown in Figure 16A. [Figure 16C]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which is the coupling configuration. [Figure 16D]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which has an open port configuration. [Figure 16E]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A that receives gas. [Figure 16F]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which receives the sterilizing agent. [Figure 16G]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which has an open valve configuration. [Figure 16H]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which transfers fluid between fluid devices coupled to the fluid connector. [Figure 16I]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which has a closed valve configuration. [Figure 16J]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which has a closed port configuration. [Figure 16K]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, which has a separate configuration. [Figure 16L]

[0120] This is a schematic diagram of the fluid connector shown in Figure 16A, separated from the sterilizer source. [Figure 17A]

[0121] This is a front perspective view of a fluid connector with a closed port configuration. [Figure 17B]

[0121] Figure 17A is a rear perspective view of a fluid connector with a closed port configuration. [Figure 17C]

[0121] Figure 17B is a rear view of the fluid connector with a closed port configuration. [Figure 17D]

[0121] This is a front perspective view of a fluid connector with an open port configuration. [Figure 17E]

[0121] Figure 17D is a rear perspective view of a fluid connector with an open port configuration. [Figure 17F]

[0121] Figure 17E is a rear view of the fluid connector with an open port configuration. [Figure 18A]

[0122] This is a side view of a fluid connector with a separate configuration. [Figure 18B]

[0122] This is a side cross-sectional view of a fluid connector with a separate configuration. [Figure 18C]

[0122] This is a side view of a fluid connector with a coupling configuration. [Figure 18D]

[0122] This is a side cross-sectional view of a fluid connector with a coupling configuration. [Figure 18E]

[0122] This is a side view of a fluid connector with an open port configuration. [Figure 18F]

[0122] This is a side cross-sectional view of a fluid connector with an open port configuration. [Figure 18G]

[0122] This is a side view of a fluid connector with an open valve configuration. [Figure 18H]

[0122] This is a side cross-sectional view of a fluid connector with an open valve configuration. [Figure 19]

[0123] This is a schematic diagram of an exemplary variation of a fluid connector system. [Figure 20A]

[0124] This is a schematic diagram of an exemplary variation of a fluid connector system. [Figure 20B]

[0124] This is a schematic diagram of an exemplary variation of the fluid connector connection process. [Figure 20C]

[0124] This is a schematic diagram of an exemplary variation of the fluid connector connection process. [Figure 21]

[0125] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 22]

[0126] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 23]

[0127] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 24A]

[0128] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 24B]

[0128] This is a schematic diagram of an exemplary variation of the fluid connector connection process. [Figure 24C]

[0128] This is a schematic diagram of an exemplary deformation of a valve. [Figure 25A]

[0129] This is a block diagram of an exemplary variation of a fluid connector system. [Figure 25B]

[0129] This is a schematic diagram of an exemplary variation of the fluid connector connection process. [Figure 25C]

[0129] This is a schematic diagram of an example of a valve deformation. [Figure 26A]

[0130] This is a side view of an exemplary variation of a pump actuator and a pump. [Figure 26B]

[0130] This is a side view of an exemplary deformation of a pump actuator coupled to a pump. [Figure 27]

[0131] This is a flowchart illustrating an exemplary variation of a method for transferring fluid using a fluid connector. [Figure 28]

[0132] This is a flowchart illustrating an exemplary variation of a cell processing method. [Figure 29]

[0133] This is a flowchart illustrating an exemplary variation of a cell processing method. [Figure 30A]

[0134] This is a flowchart illustrating an exemplary variation of a cell processing method for autologous CAR-T cells or genetically engineered TCR cells. [Figure 30B]

[0134] This is an illustrative flowchart of a modified method for processing allogeneic CAR-T cells or genetically modified TCR cells. [Figure 31]

[0135] This is a flowchart illustrating an exemplary variation of a cell processing method for HSC cells. [Figure 32]

[0136] This is a flowchart illustrating an exemplary variation of a cell processing method for TIL cells. [Figure 33]

[0137] This is a flowchart illustrating an exemplary variation of a cell processing method for NK-CAR cells. [Figure 34A]

[0138] This is a flowchart illustrating an exemplary variation of a cell processing method for Treg cells. [Figure 34B]

[0138] This is a flowchart illustrating an example of a modified method for processing Treg cells. [Figure 34C]

[0138] This is a flowchart illustrating an example of a modified method for processing Treg cells. [Figure 35]

[0139] This is a flowchart illustrating an exemplary variation of a cell processing method. [Figure 36]

[0140] This is an illustrative flowchart of a transformation that demonstrates how to perform the transformation model. [Figure 37]

[0141] This is an exemplary variation of a graphical user interface related to the initial process design interface. [Figure 38]

[0142] This is an exemplary variation of a graphical user interface related to process generation. [Figure 39]

[0143] This is an exemplary variation of a graphical user interface related to empty processes. [Figure 40]

[0144] This is an exemplary variation of a graphical user interface related to the addition of reagent and consumable containers. [Figure 41]

[0145] This is an exemplary variation of a graphical user interface related to process parameters. [Figure 42]

[0146] This is an exemplary variation of a graphical user interface related to patient weight process parameters. [Figure 43]

[0147] This is an exemplary variation of a graphical user interface related to preprocessing analysis. [Figure 44]

[0148] This is an exemplary variation of a graphical user interface related to white blood cell count preprocessing analysis. [Figure 45]

[0149] This is an exemplary variation of a graphical user interface related to process parameter calculation. [Figure 46]

[0150] This is an exemplary variation of a graphical user interface related to a completed process setup. [Figure 47]

[0151] This is an exemplary variation of a graphical user interface related to process activation settings. [Figure 48]

[0152] This is an exemplary variation of a graphical user interface related to the entered process activation settings. [Figure 49]

[0153] This is an exemplary variation of a graphical user interface related to the initial process operation. [Figure 50]

[0154] This is an exemplary variation of a graphical user interface related to dragging process behavior. [Figure 51]

[0155] This is another exemplary variation of a graphical user interface related to dragging process behavior. [Figure 52]

[0156] This is an exemplary variation of a graphical user interface related to the entered process behavior. [Figure 53]

[0157] This is an exemplary variation of a graphical user interface related to product monitoring. [Figure 54]

[0158] This is another exemplary variation of a graphical user interface related to product monitoring. [Figure 55]

[0159] This is a block diagram illustrating an exemplary variation of a manufacturing workflow. [Figure 56]

[0160] This is a block diagram of an exemplary variation of a cell separation system. [Figure 57]

[0161] This is a side cross-sectional view of an exemplary deformation of a counterflow centrifugal ertriation (CCE) module. [Figure 58]

[0162] This is a side cross-sectional view of an exemplary deformation of a magnetically activated cell selection (MACS) module. [Figure 59A]

[0163] This is a perspective view of an exemplary variation of the CCE system. [Figure 59B]

[0163] A perspective view of an exemplary variation of the CCE system. [Figure 59C]

[0163] A perspective view of an exemplary variation of the CCE system. [Figure 59D]

[0163] A side cross-sectional view of an exemplary variation of the CCE system. [Figure 59E]

[0163] A side cross-sectional view of an exemplary deformation of the rotor of the CCE module. [Figure 59F]

[0163] A side cross-sectional view of an exemplary deformation of the rotor of the CCE module. [Figure 59G]

[0163] A side cross-sectional view of an exemplary deformation of the rotor of the CCE module. [Figure 60A]

[0164] This is a plan view of an exemplary variation of the rotor of a CCE module. [Figure 60B]

[0164] A perspective view of an exemplary variation of the rotor of the CCE module. [Figure 60C]

[0164] A perspective view of an exemplary variation of the rotor of the CCE module. [Figure 60D]

[0164] This is a side view of an exemplary variation of the rotor of the CCE module. [Figure 60E]

[0164] This is a perspective view of an exemplary deformation of the rotor inside the housing. [Figure 60F]

[0164] This is a schematic plan view of an exemplary deformation of the rotor of the CCE module. [Figure 60G]

[0164] This is a schematic plan view of an exemplary deformation of the rotor of the CCE module. [Figure 60H]

[0164] This is a side view of an exemplary variation of the rotor of the CCE module. [Figure 60I]

[0164] A perspective view of another exemplary variation of the rotor of the CCE module. [Figure 60J]

[0164] A perspective view of yet another exemplary variation of the rotor of the CCE module. [Figure 60K]

[0164] A schematic plan view of another exemplary variation of the rotor dimensions of the CCE module. [Figure 60L]

[0164] An image of an exemplary variant set of rotors for the CCE module. [Figure 61A]

[0165] This is a schematic diagram illustrating an exemplary variation of the cell separation process. [Figure 61B]

[0165] This is a schematic diagram of an exemplary variation of the cell separation process. [Figure 61C]

[0165] This is a schematic diagram of an exemplary variation of the cell separation process. [Figure 62A]

[0166] This is a perspective view of an exemplary variation of the MACS system in the first configuration. [Figure 62B]

[0166] A perspective view of an exemplary variation of the MACS system in the second configuration. [Figure 62C]

[0166] This is a side cross-sectional view of an exemplary variation of the MACS system. [Figure 62D]

[0166] A perspective view of an exemplary variation of the MACS system in the second configuration. [Figure 62E]

[0166] This is a plan view of an exemplary variation of the flow cell and magnet array of the MACS system. [Figure 62F]

[0166] This is a plan view of an exemplary deformation of a flow cell in a MACS system. [Figure 62G]

[0166] This is a schematic diagram of exemplary variations of the flow cell and magnet array. [Figure 63A]

[0167] This is a perspective view of an exemplary variation of a magnet array. [Figure 63B]

[0167] Perspective view of an exemplary variation of a magnet array. [Figure 63C]

[0167] Perspective view of an exemplary variation of a magnet array. [Figure 63D]

[0167] Perspective view of an exemplary variation of a magnet array. [Figure 63E]

[0167] Perspective view of an exemplary variation of a magnet array. [Figure 64A]

[0168] This is a perspective view of an exemplary variation of a flow cell. [Figure 64B]

[0168] Side cross-sectional view of an exemplary deformation of a flow cell. [Figure 64C]

[0168] This is a schematic diagram of an exemplary variation of the MACS system. [Figure 65A]

[0169] This is a schematic diagram illustrating an exemplary variation of a flow cell. [Figure 65B]

[0169] This is a schematic diagram of an exemplary deformation of a flow cell. [Figure 65C]

[0169] This is a schematic diagram of an exemplary deformation of a flow cell. [Figure 66A]

[0170] This is a schematic diagram illustrating an exemplary variation of the cell separation process. [Figure 66B]

[0170] This is a schematic diagram of an exemplary variation of the cell separation process. [Figure 66C]

[0170] This is a schematic diagram of an exemplary variation of the cell separation process. [Figure 67A]

[0171] This is a schematic diagram of an exemplary variation of a cell processing system. [Figure 67B]

[0171] This is a schematic diagram of an exemplary variation of the cell processing system. [Figure 67C]

[0171] This is a schematic diagram of an exemplary variation of the cell processing system. [Figure 67D]

[0171] This is a schematic diagram of an exemplary variation of the cell processing system. [Figure 68A]

[0172] This is a cross-sectional perspective view of an exemplary deformation of a bioreactor. [Figure 68B]

[0172] This is a side cross-sectional view of an exemplary deformation of the bioreactor. [Figure 68C]

[0172] A perspective view of an exemplary variation of the bioreactor enclosure. [Figure 68D]

[0172] This is a plan view of an exemplary variation of the bioreactor enclosure. [Figure 68E]

[0173] This is a perspective view of an exemplary deformation of a bioreactor membrane. [Figure 68F]

[0173] This is a side view of an exemplary deformation of the bioreactor membrane. [Figure 68G]

[0173] A perspective view of an exemplary deformation of the bioreactor membrane. [Figure 68H]

[0173] This is a bottom view of an exemplary deformation of the bioreactor membrane. [Figure 69A]

[0174] This is a side cross-sectional view of an exemplary deformation of a bioreactor enclosure. [Figure 69B]

[0174] This is a cross-sectional perspective view of an exemplary deformation of the bioreactor enclosure. [Figure 70]

[0175] This is an exploded perspective view of an exemplary deformation of a bioreactor. [Figure 71A]

[0176] This is a plan view of an exemplary deformation of a bioreactor. [Figure 71B]

[0176] Side cross-sectional view of an exemplary deformation of the bioreactor. [Figure 72]

[0177] This is a schematic diagram of an exemplary variation of an electroporation system. [Figure 73]

[0178] This is an exploded perspective view of an exemplary deformation of an electroporation module. [Figure 74A]

[0179] This is a schematic diagram illustrating an exemplary variation of the electroporation process. [Figure 74B]

[0179] This is a schematic diagram of an exemplary variation of the electroporation process. [Figure 75]

[0180] This is a schematic diagram of an exemplary variation of the electroporation process. [Figure 76A]

[0181] This is a plot of exemplary variations of the electroporation process. [Figure 76B]

[0181] This is a plot of exemplary variations of the electroporation process. [Figure 76C]

[0181] This is a plot of exemplary variations of the electroporation process. [Figure 76D]

[0181] This is a plot of exemplary variations of the electroporation process. [Figure 77A]

[0182] This is a flowchart illustrating an exemplary variation of a method for separating cells. [Figure 77B]

[0182] This is a flowchart illustrating an example variation of a method for concentrating cells. [Figure 77C]

[0182] This is a flowchart illustrating a modified method of buffer exchange. [Figure 78]

[0183] This is a flowchart of another illustrative variation of a method for separating cells. [Figure 79A]

[0184] This is a flowchart illustrating an exemplary variation of the closed-loop method 7900 for separating cells. [Figure 79B]

[0184] This is a flowchart of an exemplary variation of the closed-loop method 7910 for performing cell elutriation. [Figure 79C]

[0184] This is a flowchart of an exemplary variation of the closed-loop method 7920 for collecting cells. [Figure 80A]

[0185] This is a flowchart illustrating an exemplary variation of a method for separating cells. [Figure 80B]

[0185] This is a flowchart illustrating an exemplary variation of a method for selecting cells. [Figure 81]

[0186] This is a flowchart of another illustrative variation of a method for separating cells. [Figure 82A]

[0187] This is a flowchart illustrating an exemplary variation of a method for preparing a bioreactor. [Figure 82B]

[0187] This is an illustrative flowchart of a modified method for installing a bioreactor. [Figure 82C]

[0187] This is a flowchart of an exemplary variation of a method for preparing a bioreactor. [Figure 82D]

[0187] This is a flowchart of an exemplary variation of a calibration method for a bioreactor. [Figure 82E]

[0187] This is a flowchart illustrating an exemplary variation of a method for mixing reagents. [Figure 82F]

[0187] This is a flowchart illustrating an exemplary variation of a method for mixing reagents. [Figure 82G]

[0187] This is a flowchart of an exemplary variation of a method for culturing cells. [Figure 82H]

[0187] This is a flowchart illustrating an exemplary variation of a method for refrigerating cells. [Figure 82I]

[0187] This is an illustrative flowchart of a variation of the method for obtaining a sample. [Figure 82J]

[0187] This is a flowchart of an exemplary variation of a method for culturing cells. [Figure 82K]

[0187] This is a flowchart illustrating an example variation of the method for changing the culture medium. [Figure 82L]

[0187] This is a flowchart illustrating an example variation of a method for controlling gas. [Figure 82M]

[0187] This is a flowchart illustrating an exemplary variation of a method for controlling pH. [Figure 83]

[0188] This is an illustrative flowchart of a modified method for performing electroporation of cells. [Figure 84]

[0189] This is a flowchart of another exemplary variation of a method for performing electroporation of cells. [Figure 85]

[0190] This is a schematic diagram illustrating an exemplary variation of a fluid connector. [Figure 86]

[0191] This is a schematic diagram illustrating an exemplary variation of a fluid connector port. [Figure 87]

[0192] This is a schematic diagram of an exemplary variation of a fluid connector connection process. [Figure 88]

[0193] This is a schematic diagram of an exemplary variation of a fluid connector connection process. [Figure 89]

[0194] This is a schematic diagram illustrating an exemplary variation of a fluid connector. [Figure 90A]

[0195] This is a side view of an exemplary deformation of a fluid connector. [Figure 90B]

[0195] Figure 90A is a perspective view of the fluid connector. [Figure 90C]

[0195] This is a side cross-sectional view of the fluid connector shown in Figure 90A. [Figure 91A]

[0196] This is a side view of an exemplary deformation of a fluid connector. [Figure 91B]

[0196] Figure 91A is a perspective view of the fluid connector. [Figure 91C]

[0196] This is a side cross-sectional view of the fluid connector shown in Figure 91A. [Figure 91D]

[0197] This is a side view of an exemplary deformation of a fluid connector. [Figure 91E]

[0197] Figure 91D is a perspective view of the fluid connector shown. [Figure 91F]

[0197] This is a side cross-sectional view of the fluid connector shown in Figure 91D. [Figure 92A]

[0198] This is a side view of an exemplary deformation of a fluid connector. [Figure 92B]

[0198] This is a perspective side view of the fluid connector shown in Figure 92A. [Figure 92C]

[0198] Figure 92A is a perspective view of the fluid connector shown. [Figure 92D]

[0198] This is a side cross-sectional view of the fluid connector shown in Figure 92A. [Figure 93A]

[0199] This is a perspective view of an exemplary variation of a fluid connector. [Figure 93B]

[0199] Figure 93A is a perspective view of the fluid connector. [Figure 94A]

[0200] This is a perspective view of an exemplary variation of a fluid connector. [Figure 94B]

[0200] Figure 94A is a perspective view of the fluid connector. [Figure 95A]

[0201] This is a perspective view of an exemplary variation of a fluid connector. [Figure 95B]

[0201] It is a perspective view of the fluid connector shown in FIG. 95A. [Figure 95C]

[0201] It is a detailed side view of the port with an open port configuration. [Figure 95D]

[0201] It is a detailed side view of the port with a closed port configuration. [Figure 96A]

[0202] It is a plan view of an exemplary modification of the fluid device. [Figure 96B]

[0202] It is a side view of an exemplary modification of the fluid device coupled to the robot. [Figure 96C]

[0202] It is a perspective view of an exemplary modification of the fluid device held by the robot. [Figure 97A]

[0203] It is a perspective view of an exemplary modification of the MACS module. [Figure 97B]

[0203] It is a cross-sectional perspective view of an exemplary modification of the MACS module. [Figure 97C]

[0203] It is a side cross-sectional view of an exemplary modification of the MACS module. [Figure 98]

[0204] It is a flowchart of an exemplary modification of the cell processing method. [Figure 99]

[0205] It is a flowchart of an exemplary modification of the cell processing method. [Figure 100]

[0206] It is a flowchart of an exemplary modification of the cell processing method. [Figure 101]

[0207] It is a flowchart of an exemplary modification of the cell processing method. [Figure 102]

[0208] It is a schematic diagram of an exemplary modification of the cell processing system. [Figure 103A]

[0209] It is a perspective view of an exemplary modification of the sterilizing liquid transfer device. [Figure 103B]

[0209] It is a perspective view of an exemplary modification of the sterilizing liquid transfer device.

MODE FOR CARRYING OUT THE INVENTION

[0105]

[0210] This specification describes systems and methods for processing and producing cell products for biomedical use. Cell processing methods and systems may include moving a cartridge containing cell products between multiple instruments within a work cell. One or more instruments are configured to work in conjunction with the cartridge to perform cell processing steps on the cell products, thereby enabling the system (e.g., work cell) to perform cell processing steps on the cell products. In some variations, multiple cell processing steps can be performed within a single cartridge. For example, a robotic arm may be configured to move the cartridge between multiple instruments for different cell processing steps. The cartridge may include multiple cell processing devices (e.g., modules). These devices include, for example, bioreactors, countercurrent centrifugation (CCE) modules, magnetic cell sorting devices (e.g., magnetically activated cell selection modules), electroporation devices (e.g., electroporation modules), sorting modules (e.g., fluorescence-activated cell classification (FACS) modules), acoustic flowcell modules, centrifugation modules, microfluidic enrichment modules, and combinations thereof. In some variations, the system can process two or more cartridges in parallel. For example, a bioreactor may include multiple slots configured to work with multiple cartridges simultaneously, and typically, one process step (e.g., cell culture in the bioreactor) may be the rate-limiting step in the operation of the cell processing system. The cell processing systems described herein can increase throughput by reducing operator intervention through the automation of the movement of cartridges (and cell products) between instruments using a robot. However, in some variations, cartridges may be moved manually between instruments. Furthermore, the use of multiple bioreactors may enable the system to process multiple cartridges from multiple patients simultaneously, thereby increasing the system's throughput.Furthermore, automated cell processing systems can facilitate the transfer of sterile fluid between cartridges and instruments or other components of the system. These other components may include, for example, fluid connectors (e.g., sterile fluid transfer ports), reagent storage containers, second cartridges, sampling containers (e.g., sterile fluid transfer devices), or combinations thereof.

[0106] Workcell

[0211] In some variations, a system for cell processing (e.g., a work cell) may include multiple instruments. Each instrument is independently configured to perform one or more cell processing operations on a cartridge. A robot may be configured to move the cartridge between each of the instruments. The instruments may include one or more of the following: bioreactor instruments, cell selection instruments (e.g., magnetically activated cell selection instruments), classification instruments (e.g., fluorescence-activated cell classification (FACS) instruments), electroporation instruments, countercurrent centrifugation (CCE) instruments, reagent storage containers, etc. The system can perform automated production of cell products.

[0107]

[0212] The cartridge may be portable and configured to facilitate automated sterile cell processing using work cells and robots. For example, the cartridge may be configured to move to one or more instruments in a work cell to perform different cell processing steps. In some variations, the instrument may be configured to move to the cartridge. In some variations, the cartridge may include multiple modules, including one or more of the following: a bioreactor module, a cell selection module (e.g., a magnetically activated cell selection module), a classification module (e.g., a fluorescence-activated cell classification (FACS) module), an electroporation module, and a countercurrent centrifugation eltriation (CCE) module. The cartridge may further include one or more of the following: a sterile fluid transfer port, a fluid transfer bus fluidically coupled to each module, and a pump fluidically coupled to the fluid transfer bus.

[0108]

[0213] In some variations, a method for processing a solution containing cell products may include a cell processing step of digesting tissue with an enzymatic reagent to release a selected cell population into solution; a cell processing step of concentrating cells using a CCE instrument; a cell processing step of washing cells using a CCE instrument; a cell processing step of selecting cells in solution using a selection instrument; a cell processing step of classifying cells in solution using a classification instrument; a cell processing step of differentiating or proliferating cells in a bioreactor; a cell processing step of activating cells using an activation reagent; a cell processing step of electroporating cells; a cell processing step of transducing cells using a vector; and a cell processing step of completing the cell product.

[0109] Cell selection system

[0214] The cell processing systems described herein may include cell selection systems configured to separate cells based on predetermined criteria. For example, cells can be separated based on physical characteristics such as size and / or density using a countercurrent centrifugation elutriation instrument. Alternatively, cells can be separated based on the presence of a predetermined antigen using, for example, a magnetically activated cell selection instrument. In some variations, a cell selection system including modules for these separation methods can facilitate one or more cell processing steps, including, but not limited to, cell concentration, cell dilution, cell washing, buffer replacement, and magnetic separation. The cell selection systems described herein can increase throughput and cell yield output in a compact, portable structure. For example, before magnetically separating cells, the cell suspension may be mixed with a magnetic reagent in excess or at a predetermined concentration (e.g., cells / ml). Similarly, after magnetically separating cells, the cells may be washed with a solution (e.g., a suitable buffer solution).

[0110]

[0215] In some variations, the cell separation system may include a rotor configured for countercurrent centrifugation of cells in a fluid, a first magnet configured to magnetically rotate the rotor and separate cells from the fluid in the rotor, a flow cell configured to communicate with the rotor and receive cells from the rotor, and a second magnet configured to magnetically separate cells in the flow cell.

[0111]

[0216] In some variations, the CCE module is integrated into a cartridge, enabling the cell processing system to separate cells based on their size and / or density. In some variations, the cell separation system may include a housing containing a rotor configured to separate cells from a fluid (e.g., to separate cells of different sizes and / or densities from cells in a fluid), and a magnet configured to magnetically rotate the rotor. The housing may be configured to move relative to the magnet or vice versa (e.g., to move the magnet relative to the housing). The CCE module described herein can perform cell separation in a compact, portable housing, and the magnet may be located outside the housing (e.g., the magnet is located inside the CCE instrument).

[0112]

[0217] In some variations, a compact rotor that can be useful for cartridge integration may include input and output fluid tubes extending from the rotor to both sides of the rotor housing. For example, the rotor may include a first side containing a first fluid tube and a second side containing a second fluid tube on the opposite side of the first side. An ertriation chamber (e.g., a cone) can be coupled between the first and second fluid tubes.

[0113]

[0218] In some variations, the method for separating cells from a fluid may include moving a rotor defining a rotation axis toward a magnet, allowing the fluid to flow through the rotor, rotating the rotor (e.g. magnetically) around the rotation axis using a magnet while the fluid flows through the rotor, and moving the rotor toward the direction away from the magnet.

[0114]

[0219] In some variations, the method for separating cells from a fluid may involve introducing the fluid containing the cells into a flow cell. The set of cells can be labeled with magnetic particles. The set of cells can be magnetically attracted to a magnetic array during the residence time and then flow out of the flow cell after the residence time.

[0115]

[0220] In some variations, the flow cell may include an elongated cavity having a certain cavity height, and the magnet array may include multiple magnets, each separated by a certain distance. A predetermined ratio between the cavity height and the separation distance can optimize the magnetic separation of cells within the flow cell.

[0116] Electroporation

[0221] In some variations, the electroporation modules described herein may be configured to facilitate one or more of the transfection and introduction of cells. As detailed herein, a fluid volume containing cells (e.g., a first batch) can be physically separated from subsequent fluid volumes containing cells (e.g., a second batch, a third batch) by gas (e.g., an air gap). Applying separate electroporation signals (e.g., voltage pulses, waveforms) to each of the individual fluid batches can improve electroporation efficiency and thus increase throughput. In some variations, electroporation efficiency and throughput can be similarly improved by active field compensation.

[0117]

[0222] In some variations, the cell processing apparatus may include a fluid conduit configured to receive a first fluid containing cells and a second fluid (such as a gas, oil, etc.), an electrode set coupled to the fluid conduit, a pump coupled to the fluid conduit, and a controller including a processor and a memory. The controller may generate a first signal to introduce the first fluid into the fluid conduit using the pump, generate a second signal to introduce the second fluid into the fluid conduit so that the second fluid separates the first fluid from a third fluid, and may further be configured to generate an electroporation signal to perform electroporation of the cells in the fluid conduit using the electrode set.

[0118]

[0223] In some variations, a method of performing electroporation of cells may include receiving a first fluid containing cells in a fluid conduit, receiving a second fluid containing a gas in the fluid conduit so that the first fluid is separated from a third fluid, and applying an electroporation signal to the first fluid to perform electroporation of the cells.

[0119]

[0224] In some variations, a method of performing electroporation of cells may include receiving a first fluid containing cells in a fluid conduit, applying a resistance measurement signal to the first fluid using an electrode set, measuring the resistance between the first fluid and the electrode set, and applying an electroporation signal to the first fluid based on the measured resistance.

[0120] Bioreactor

[0225] In some variations, the bioreactor may include an enclosure including a bottom and sidewalls, and a gas permeable membrane coupled to one or more of the bottom and sidewalls of the enclosure. The gas permeable membrane can assist in cell culture. In some variations, the cell processing system may include a bioreactor and a stirrer coupled to the bioreactor. The stirrer may be configured to stir the bioreactor based on orbital motion.

[0121] Fluid connector

[0226] Currently, in cell therapy manufacturing, there are no automated sterile fluid connector solutions for multiple uses that allow for multiple connection and disconnection cycles to and from a system. For example, conventional sterile fluid connectors are typically single-use devices and are therefore expensive and labor-intensive. Generally, the fluid connectors described herein facilitate sterile control of the fluid connector and the device coupled thereto by including multiple sealed enclosures between the sterile portion (e.g., the lumen or cavity of the fluid connector) and the external (e.g., non-sterile) surrounding environment. The fluid connectors described herein may be highly durable components that can be reused for multiple cycles while maintaining sterility and / or bioburden control. For example, the fluid connector can be sterilized with a sterilizing agent without damaging cell products or other biological materials.

[0122]

[0227] In some variations, the sterile manufacturing systems described herein can utilize one or more sterile fluid connectors and have a configuration suitable for operation by robots such as robotic arms. The sterile fluid connectors described herein enable automated, sterile, and metered fluid transfer to automate cell therapy manufacturing. Automating cell therapy manufacturing reduces the manufacturing cost per patient, lowers the risk of process failure, and can meet the commercial-scale patient demand for cell therapy. In some variations, the sterile fluid connectors can increase one or more of the following by eliminating human operators from the manufacturing process: sterility, efficiency, and speed. The automated and integrated sterilization process described herein can be applied to the fluid connectors to maintain the sterility of the system. For example, the fluid connectors can maintain sterility over multiple connection / disconnection cycles between separate sterile closed volumetric fluid devices (e.g., enclosures, containers, vessels, cartridges, instruments, bioreactors, closed vessels, sealed chambers). Thus, the systems, devices, and methods described herein can reduce the complexity of the sterilization process, reduce energy consumption, and increase sterilization efficiency.

[0123]

[0228] In some variations, the fluid connector may include a first connector configured to interlock with a second connector (e.g., a male and female connector). Each proximal end of these connectors may be configured to connect to each fluid device (e.g., fluid-communicate, form a fluid pathway) for the transfer of one or more fluids (e.g., liquids and / or gases) and biological substances (e.g., cell products) between the fluid devices. The distal ends of these connectors may include ports configured to interlock with each other. The fluid connector may also include sterilizer ports configured within the distal ends of the first and second connectors to facilitate sterilization of the chamber. The fluid connector can be sterilized as desired before or after connection to ensure sterility. In this way, the fluid connector can be reused over multiple connection and disconnection cycles.

[0124]

[0229] In some variations, a system utilizing the fluid connector described herein (e.g., a work cell) may include a robot configured to operate the fluid connector and a controller configured to control the robot to operate the first and second connectors together (without human intervention) (e.g., move, connect, open, close, disconnect) while further reducing the risk of contamination by maintaining the sterility of the fluid connector and multiple fluid devices. The fluid devices may be one or more of instruments and cartridges.

[0125] Cell processing control

[0230] This specification describes systems and methods for producing cell products for biomedical use using automated systems. Conventional semi-automated solutions for cell processing do not allow the user to define the biological process. Instead, the user selects from a limited set of predefined machine processes and process control parameters. Currently, there are no scalable manufacturing solutions for cell therapy production. For example, cell therapy production is traditionally performed in batches (i.e., one product is produced in a single laboratory / set, with the necessary processing tools located inside). This is guided by technicians following standard operating procedures (SOPs), or in some cases, processing tools (e.g., Miltenyi Prodigy, Lonza Cocoon) can perform a series of processing steps for a single patient's product in a single multi-functional processing tool. However, existing solutions (e.g., Miltenyi Prodigy) do not allow the user to define the biological process. Furthermore, the manual work required in conventional solutions increases the risk of product contamination and human error.

[0126]

[0231] In some variations, the system described herein can be used to convert a set of cell therapy biological manufacturing processes into a set of machine instructions suitable for automated execution. For example, a method for converting user-defined cell processing operations into cell processing steps to be executed by the processor of an automated cell processing system may include receiving an ordered input list of cell processing operations and running a transformation model on the ordered input list to generate an ordered output list of cell processing steps that can be executed by the system. As used herein, a transformation model may refer to an algorithm, process, or transformation rule configured to convert a set of cell processing steps into a set of machine or hardware instructions for the system. In some variations, a robot can be controlled to move one or more cartridges, each containing cell products, between instruments, and the instruments can be controlled to execute cell processing steps for each cell product. Thus, the system and method allow biologists to define manufacturing processes in biological terms and have the system convert this biological model (e.g., process definition) into a set of machine execution instructions.

[0127]

[0232] The comprehensive automation of closed systems described herein can reduce process failure rates and costs. For example, comprehensive automation can shorten manufacturing time (e.g., residence time) and increase throughput compared to conventional manual methods. For example, multiple processes (e.g., 10 or more) can be executed simultaneously. The methods described herein can further reduce the possibility of contamination and user error. Thus, the systems, apparatus, and methods described herein can increase one or more of the following in relation to cell processing: automation, reproducibility, reliability, process flexibility, instrument throughput, and process scalability, while reducing one or more of the following in relation to labor costs and process time.

[0128] I. system

[0233] Described herein are systems and apparatus configured to perform cell processing steps for producing cell products (e.g., cell therapy products). In some variations, a cell processing system may include a plurality of instruments independently configured to perform one or more cell processing operations on a cartridge (e.g., a fluid device), and a robot capable of moving the cartridge between each of these instruments. The use of robots and controllers can enhance one or more of the automation, efficiency, and sterility of the cell processing system.

[0129]

[0234] In some variations, the system for cell processing may include multiple instruments, each independently configured to perform one or more cell processing operations on a cartridge. The robot can move the cartridge between each of these instruments. In some variations, the system may consist of a work cell including an enclosure.

[0130]

[0235] Figure 1A is a block diagram of a cell processing system 100 including a work cell 110 and a controller 120. In some variations, the work cell 110 may include one or more of the following: instruments 112, cartridges 114 (e.g., consumables, fluid devices), a robot 116 (e.g., a robotic arm), a reagent storage container 118, a fluid connector 132, a sterilizer source 134, a fluid source 136, a pump 138, a sensor 140, and a sterilizer transfer device 142. In some variations, the controller 120 may include one or more of the following: a processor 122, memory 124, a communication device 126, an input device 128, and a display 130.

[0131]

[0236] In some variations, the work cell can include a fully or at least partially enclosed housing within which one or more cell processing steps are performed in a fully or at least partially automated process. In some variations, the work cell may be an open system that does not include an enclosure and can be configured for use in a clean room, a biosafety cabinet, or other sterile location. In some variations, to reduce manual labor in the cell processing steps, a robot 116 can be used to move the cartridge 114. In some variations, the work cell can be configured to perform the transfer of sterilizing liquid into and out of the cartridge in a fully or partially automated process. For example, one or more fluids can be stored in the sterilizing liquid transfer device 142. In some variations, the sterilizing liquid transfer device can be a portable consumable that is movable within the system 100. The sterilizing liquid transfer devices and fluid connectors described herein enable automatic, sterile, and quantitative fluid transfer for automating cell therapy manufacturing. In some variations, the enclosure of the work cell can be configured to meet the standards of the International Organization for Standardization (ISO) standard ISO7 or higher (e.g., ISO6 or ISO5). The advantage of meeting ISO7 or higher standards is that the system can be used in facilities that do not meet the ISO7 standard (i.e., are not clean rooms or other adequately filtered air spaces). Optionally, this facility can also be an ISO8 or ISO9 facility. In some variations, the volume of the work cell is less than about 800 m 3 less than, about 700 m 3 less than, about 600 m 3 less than, about 500 m 3 less than, about 300 m 3 less than, about 250 m 3 less than, about 200 m 3 less than, about 150 m 3 less than, about 100 m 3 less than, about 50 m 3 less than, about 25 m 3 less than, about 10 m3 Less than, and approximately 5m 3 This can include values ​​less than, as well as all ranges and sub-values ​​between these values.

[0132]

[0237] In some variations, the robot 116 may be configured to manipulate consumable cartridges 114 and fluid connectors 132 between different instruments to perform a predetermined cell processing step sequence. In some variations, it is also possible to receive the same consumable cartridge 114 in different instruments 112 and / or process multiple cartridges 114 in parallel.

[0133]

[0238] In some variations, the cartridge 114 can contain cell products from different donors or cell products for different recipients. Cell products from a single donor may be divided into multiple cartridges 114 if necessary to generate enough product for therapeutic use, or if the donor provides product to multiple recipients (e.g., for allogeneic transplantation). Cell products for a single recipient may also be divided into multiple cartridges 114 if necessary to generate enough product for therapeutic use in that recipient. If necessary to generate several cell products through specific genetic recombination, cell products for a single recipient may be divided into multiple cartridges 114 and then optionally recombined in a specific ratio for therapeutic use in that recipient. For example, fluid connectors 132 can be connected between two or more cartridges 114 to transfer cell products and / or fluids between these cartridges 114. Furthermore, fluid connectors 132 can be connected between the fluid-carrying component set of system 100 (e.g., cartridge 114, reagent storage container 118, fluid source 136, sterile fluid transfer device 142, fluid tubes, containers, etc.). For example, a first fluid connector can be connected between the first cartridge and the sterile fluid transfer device, and a second fluid connector can be connected between the sterile fluid transfer device and the second cartridge.

[0134]

[0239] As shown in Figure 1B, the cartridge 114 may include one or more of the following, as detailed herein: a bioreactor 150, a cell separation system 152, an electroporation module 160, a fluid transfer bus 162, a sensor 164, and a fluid connector 166. The cell separation system 152 may include one or more of the following: a rotor 154, a flow cell 156, and a magnet 158. In some variations, the magnet 158 ​​may include one or more magnets and / or a magnet array. For example, the cell separation system 152 may include a first magnet configured to magnetically rotate the rotor 154 and a second magnet (e.g., a magnet array) configured to magnetically separate cells within the flow cell 156.

[0135] Workcell

[0240] In some variations, the work cell 110 may include at least a partially enclosed enclosure (e.g., housing) in which one or more automated cell processing steps are performed. For example, the work cell 110 may be configured to transfer sterile fluid in and out of the cartridge 114 in a fully or partially automated process. In some variations, the work cell 110 may be configured without an enclosure for use in a cleanroom, biosafety cabinet, or other appropriately clean or sterile location. In some variations, the work cell 100 may include a feedthrough access biosafety cabinet, quality control equipment, pumps, consumables (e.g., fluid devices), fluid connectors, consumable feedthroughs, and a sterilization system (e.g., sterilizer source and / or generator, fluid source, heater / desiccator, aerator).

[0136]

[0241] Figure 2A is a block diagram of a cell processing system including a work cell 203. The work cell 203 may include an enclosure 202 having four walls, a bottom, and a roof. The work cell may be divided into an internal zone 204 with feedthrough access 206 and quality control (QC) equipment 212. An air filtration inlet (not shown) can provide ISO 7 or higher air quality to the internal zone 204 by performing high-efficiency particulate air (HEPA) filtration. This air filtration can maintain sterile cell processing in an ISO 8 or ISO 9 manufacturing environment. The work cell 203 may also have an air filter at the air outlet to maintain the ISO rating of the room. In some variations, the work cell 203 may further include, within the internal zone 104, a bioreactor instrument 214, a cell selection instrument 216 (e.g., MACS), an electroporation instrument (EP) 220, a countercurrent centrifugation (CCE) instrument 222, a sterile fluid transfer instrument 224 (e.g., a fluid connector), a reagent storage container 226, and a sterilization system 260. The reagent storage container 226 may be accessible to the user via a sample pickup port 228. A robot 230 (e.g., a support arm, a robotic arm) may be configured to move one or more cartridges 250 (e.g., consumables) from one instrument to another, and / or move one or more cartridges 250 to or from the reagent storage container. In some variations, the work cell 203 may include one or more movable barriers 213 (e.g., access means, doors) configured to facilitate access to one or more of the instruments within the work cell 203.

[0137]

[0242] In some variations of the method according to this disclosure, a human operator can insert one or more empty cartridges 250 into the feedthrough 206 via the cartridge port 207. The cartridges 250 can be pre-sterilized, or the feedthrough 206 can sterilize the cartridges 250 using ultraviolet radiation (UV) or with a chemical sterilizer provided as vapor, spray, or washing solution. The feedthrough 206 chamber may optionally be configured to automatically spray, wash, irradiate, or otherwise treat the cartridges (e.g., using ethanol and / or isopropyl alcohol solution, vaporized hydrogen peroxide (VHP)) to maintain the sterility of the internal zone 204 (e.g., ISO 7 or higher). The cartridges 250 can be transferred to a biosafety cabinet 206, where input cell products can be provided and inserted into the cartridges 250 via a sterilization solution transfer port. The user can then return the cartridges 250 to the feedthrough 206 (by a robot 230) and initiate automated processing using a computer processor in a computer server rack (e.g., controller 120). The robot 230 may be configured to move the cartridge 250 to multiple instruments and stations using the components of the work cell 200 in a predetermined sequence. At the end of cell processing, the cartridge 250 containing the processed cell products can be returned to the feedthrough 206 for retrieval by the user. In some variations, the outer surface of the enclosure 202 may include input / output devices 208 (e.g., displays, touchscreens).

[0138]

[0243] Figure 2B is a perspective view of a work cell 205 of the cell processing system. Figure 2C is a perspective view of the cell processing system showing a cartridge 250 (e.g., one of the cartridges described herein) introduced into a work cell 205 (e.g., one of the work cells described herein). Multiple cartridges may be inserted into the work cell 205 simultaneously, and one or more cell processing operations may be performed in parallel on these cartridges.

[0139]

[0244] In some variations, the height of workcell 205 may include values ​​and ranges greater than approximately 1 m, between approximately 1 m and approximately 3 m, between approximately 1 m and approximately 5 m, between approximately 3 m and approximately 10 m, between approximately 5 m and approximately 20 m, between approximately 10 m and approximately 30 m, between approximately 20 m and approximately 100 m, greater than approximately 100 m, and all values ​​and ranges in between. In some variations, one or more of the length and width of workcell 205 may include values ​​and ranges greater than approximately 1 m, between approximately 1 m and approximately 5 m, between approximately 3 m and approximately 10 m, between approximately 5 m and approximately 20 m, between approximately 10 m and approximately 30 m, between approximately 20 m and approximately 100 m, greater than approximately 100 m, and all values ​​and ranges in between.

[0140]

[0245] Figure 2D is a schematic diagram of a variation of the work cell 200. The work cell 200 may include an enclosure 202 having four walls, a bottom, and a roof. The work cell may be divided into an internal zone 204 with feedthrough access 206, a biosafety cabinet (BSC) 208, a computing server rack 210 (e.g., controller 120), and quality control (QC) equipment 212. An air filtration inlet (not shown) can perform high-efficiency particulate air (HEPA) filtration to provide ISO 7 or higher air quality to the internal zone 204. This air filtration can maintain sterile cell processing in an ISO 8 or ISO 9 manufacturing environment. The work cell may also have an air filter at the air outlet to maintain the ISO rating of the room. In some variations, the work cell 200 may further include, within the internal zone 204, instruments 211 (e.g., located in a universal instrument bay), a bioreactor instrument 214, a cell selection instrument 216 (e.g., MACS, a cell selection system), a cell classification instrument 218 (e.g., FACS), an electroporation instrument (EP) 220, a countercurrent centrifugation (CCE) instrument 222, a sterile fluid transfer instrument 224 (e.g., a fluid connector), a reagent storage container 226, and a sterilization system 260 including one or more of a sterilizer source, a fluid source, and a pump. The reagent storage container 226 may be accessible to the user via a sample pickup port 228. A robot 230 (e.g., a support arm, a robotic arm) may be configured to move one or more cartridges 250 (e.g., consumables) from one instrument to another or to the reagent storage container.

[0141]

[0246] In some variations, a human operator can insert one or more cartridges 250 into the feedthrough 206. The cartridges 250 can be pre-sterilized, or the feedthrough 206 can sterilize the cartridges 250 using ultraviolet radiation (UV) or with a chemical sterilizer provided as a spray or washing solution. The feedthrough 206 chamber can optionally be configured to automatically spray, wash, irradiate, or otherwise treat the cartridges (e.g., using an ethanol and / or isopropyl alcohol solution) to maintain the sterility of the internal zone 204 (e.g., ISO 7 or higher) or the biosafety cabinet 208 (e.g., ISO 5 or higher). The cartridges 250 are transferred to the biosafety cabinet 206, where input cell products can be supplied and inserted into the cartridges 250 using a sterilization fluid transfer device 224 (e.g., a fluid connector). The user can then return the cartridges 250 to the feedthrough 206 and initiate automated processing using a computer processor in a computer server rack 210 (e.g., controller 120). The robot 230 may be configured to move the cartridge 250 to multiple instruments and stations in a predetermined sequence. The components of the work cell 200 are controlled by a computer processor in a computer server rack 210. Additionally or alternatively, the sequence in which the cartridge 250 moves within the work cell 200 does not have to be predetermined. For example, the movement of the cartridge 250 may be determined based on one or more of the results of a previous step, sensor values, predetermined thresholds (e.g., based on a quality control system), etc. At the end of cell processing, the cartridge 250 containing the processed cell product can be returned to the feedthrough 206 for retrieval by the user. Additionally or alternatively, the cell product 250 containing the processed cell product may be transferred (via a fluid connector) to a second cartridge (e.g., a single-use cartridge) and stored in a reagent storage container 226 for retrieval by the user.

[0142]

[0247] In some variations, cells and starting reagents from a patient can be inserted into a cartridge (e.g., a single-use cartridge) by a human operator in a biosafety cabinet that is located separately from or integrated with the work cell. In some variations, the cartridge described herein, containing the cell products and reagents, is closed so that it can be moved through a non-sterile field without contamination. An automated decontamination routine may also be performed on the cartridge. For example, the cartridge may be placed in a feedthrough that facilitates decontamination of the cartridge before it enters the ISO7 environment of the work cell.

[0143]

[0248] Figure 2E is a schematic plan view of another variation of work cell 201. Work cells 200, 201, and 203 may include an enclosure 202 having four walls, a bottom, and a roof. The work cell may be divided into an internal zone 204 with feedthrough access 206, a biosafety cabinet (BSC) 208, a computing server rack 210 (e.g., controller 120), and quality control (QC) equipment 212. An air filtration inlet (not shown) can perform high-efficiency particulate air (HEPA) filtration to provide ISO 7 or higher air quality to the internal zone 204. This air filtration can maintain sterile cell processing in an ISO 8 or ISO 9 manufacturing environment. The work cell may also have an air filter at the air outlet to maintain the ISO rating of the room. In some variations, the work cell 200 may further include, within the internal zone 104, instruments 211 (e.g., located in a universal instrument bay), a bioreactor instrument 214, a cell selection instrument 216 (e.g., MACS), a cell classification instrument 218 (e.g., FACS), an electroporation instrument (EP) 220, a countercurrent centrifugation (CCE) instrument 222, a sterile fluid transfer instrument 224, and a reagent storage container 226. The reagent storage container 226 may be accessible to the user via a sample pickup port 228 (e.g., a door that facilitates bulk loading of the sterile fluid transfer instrument 224). A robot 230 (e.g., a support arm, a robotic arm) may be configured to move one or more cartridges 250 (e.g., consumables) from one instrument to another or to the reagent storage container.

[0144]

[0249] In some variations of the method according to this disclosure, a human operator can insert one or more empty cartridges 250 into the feedthrough 206. Additionally or alternatively, pre-filled cartridges may be inserted into the feedthrough 206. The cartridges 250 can be pre-sterilized, or the feedthrough 206 can sterilize the cartridges 250 using ultraviolet radiation (UV) or using a chemical sterilizer provided as a spray or washing solution. The feedthrough 206 chamber may optionally be configured to automatically spray, wash, irradiate, or otherwise treat the cartridges (e.g., using an ethanol and / or isopropyl alcohol solution) to maintain the sterility of the internal zone 204 (e.g., ISO 7 or higher) or the biosafety cabinet 208 (e.g., ISO 5 or higher). The cartridges 250 can be transferred to the biosafety cabinet 106, where input cell products can be supplied and inserted into the cartridges 250 via a sterilization transfer port. The user can then return the cartridge 250 to the feedthrough 206 and start automated processing using a computer processor in the computer server rack 210 (e.g., controller 120). The robot 230 may be configured to move the cartridge 250 to multiple instruments and stations in a predetermined sequence. The components of the work cell 200 are controlled by the computer processor in the computer server rack 210. At the end of cell processing, the cartridge 250 containing the processed cell products can be returned to the feedthrough 206 for retrieval by the user.

[0145]

[0250] In some variations, one or more components of a sterilization system (e.g., a sterilizing agent source, a pump) can be coupled to a work cell. For example, Figure 3 is a block diagram of a cell processing system 300 including a work cell 310, a sterilization system 320, a fluid connector 330, and a fluid device 340. In some variations, the fluid device 340 may include a main (e.g., consumables) feedthrough and a fluid device (e.g., reagent) feedthrough. The sterilization system 320 may include a sterilizing agent source 322, a pump 324, and a heater (e.g., a desiccant / dryer) 326. For example, the heater 326 may be configured to ventilate under a predetermined set of conditions. The sterilization system 320 can be coupled to one or more of the work cell 310, the fluid connector 330, and the fluid device 340, and may be in fluid communication with them. In some variations, a robot (not shown) may be configured to operate and control the cell processing system 300. For example, the fluid connector 330 may be coupled to one or more of the fluid device 340 and instruments (not shown). By circulating one or more sterilizing agents and fluids (e.g., hot air, vaporized hydrogen peroxide (VHP)) using the sterilization system 320, one or more of the work cell 310, fluid connector 330, and fluid device 340 can be sterilized and / or ventilated. In some variations, the sterilization system 320 may include one or more of vaporized hydrogen peroxide (VHP), electron beam (e-beam) sterilization, dry thermal decontamination, and steam-in-place sterilization. In some variations, the sterilization system 320 may provide a sterility assurance level (SAL) of at least 10⁻³ SAL.

[0146]

[0251] Figures 4A and 4B show perspective views of the cell processing system 400, including cartridges 400 and 402, feedthroughs 410 and 412, and fluid connectors 420 and 422 (e.g., sterile fluid transfer devices). For example, cartridge 400 is shown in Figure 16A within the feedthrough 410, after a robot (not shown) has moved cartridge 400 to the fluid connector 420.

[0147] robot

[0252] In general, a robot may include any mechanical device capable of moving a cartridge from one position to another. For example, a robot may include a mechanical manipulator (e.g., an arm) that is in a fixed position or mounted on a linear rail or a two-dimensional or three-dimensional rail system. In one variation, the robot includes a robotic shuffling system. In another variation, the robot includes a wheeled device. In some variations, the system includes two or more robots of the same or different types (e.g., two robotic arms, each independently configured to move a cartridge between instruments). The robot may also include end effectors for precise manipulation of various cartridges, or for scanning barcodes or reading radio-frequency identification tags (RFID).

[0148]

[0253] Figure 5 is a perspective view of the cell processing system 500. In this system, a robotic arm moves consumable cartridges between slots of various instruments, each configured to perform a different cell processing step. In some variations, the same consumable cartridge can be received by various instruments. The system 500 may include a modular design to accommodate various instrument configurations. In some variations, multiple cartridges can be processed in parallel. Each cartridge can contain cells from different donors or cell products for different recipients. For example, if a donor provides products to multiple recipients (e.g., for allogeneic transplantation), cell products from a single donor can be divided into multiple cartridges to generate a predetermined amount of cell products for use in therapy. In some variations, cell products for a single recipient can be divided into multiple cartridges to generate a predetermined amount of product for use in therapy in that recipient. In some variations, cell products for a single recipient can be divided into multiple cartridges to generate a predetermined amount of several cell products through specific genetic recombination and recombined in a specific ratio for use in therapy in that recipient.

[0149] cartridge

[0254] Generally, the cell processing systems described herein may comprise one or more cartridges containing one or more modules configured to work in conjunction with one or more instruments. A robot (e.g., a robotic arm) may be configured to move the cartridge and / or instruments to perform one or more cell processing steps. For example, a cartridge may include a bioreactor module and / or fluid connectors (e.g., sterile fluid transfer ports) coupled by the robot to a bioreactor instrument in a work cell. Once a predetermined processing step is completed, the cartridge can be moved by the robot to another instrument in the work cell, and another cartridge can be coupled to the bioreactor instrument. Thus, portable cartridges and shareable instruments can increase the efficiency, throughput, and flexibility of the cell manufacturing process.

[0150]

[0255] In some variations, the cartridge may optionally provide a self-contained device capable of performing one or more cell processing steps. Modules may be integrated into a fixed configuration within the cartridge. Additionally or alternatively, modules may be configurable or movable within the cartridge, allowing various cartridges to be assembled from shared modules. Similarly, the cartridge may be a single closed unit with fixed components for each module. Alternatively, the cartridge may include configurable modules coupled by configurable fluid, mechanical, optical, and electrical connections. In some variations, one or more sub-cartridges, each containing a set of modules, may be configured to be assembled to perform various cell processing workflows. Each module may be provided in a separate housing or integrated with other modules within a cartridge or sub-cartridge. For simplicity, this disclosure generally presents modules as separate groups of components, but they can be arranged in any suitable configuration. For example, components of different modules may be scattered, and each module may be defined by a set of connected components that collectively perform a predetermined function. However, the components of each module may or may not be physically grouped within the cartridge. In some variations, a single cell product can be processed using multiple cartridges. This can be done by transferring the cell product from one cartridge of the same or different type to another, and / or by dividing the cell product into many cartridges, and / or by pooling multiple cell products in a small number of cartridges.

[0151]

[0256] Generally, each instrument in the system interacts with one or more modules on a cartridge. For example, the electroporation module on the cartridge (if present) is moved by the system to the electroporation instrument, which then interacts with the electroporation instrument to perform the electroporation step on the cell products, and can also interact with common components such as components of the fluid bus line (e.g., pumps, valves, sensors, etc.). The advantage of such a segmented module / instrument design is that expensive components (e.g., motors, sensors, heaters, lasers, etc.) can be kept within the system's instruments while processing multiple cartridges. In such variations, the use of disposable cartridges eliminates the need to sterilize cartridges between uses. Furthermore, multiple instruments can be used simultaneously and in parallel by multiple cell manufacturing processes, potentially increasing the use of shared instruments (e.g., electroporation instruments, CCE instruments, MACS instruments, sterile fluid transfer instruments, FACS instruments, etc.). In contrast, conventional semi-automatic instruments (e.g., Miltenyi Prodigy) have instrument components that cannot be used simultaneously in parallel while idle.

[0152]

[0257] Figure 6 is a schematic diagram of a cartridge 600, which can be a consumable made from cost-effective materials that are useful for recycling or limited use. The cartridge 600 may include a liquid transfer bus 624 fluidly coupled to a small bioreactor module 614a, a large bioreactor module 614b, a cell selection module 616, a cell classification module 618, an electroporation module 620, and a countercurrent centrifugation ertriation (CCE) module 622. In some variations, the cell selection module 616 may be a magnetically activated cell selection (MACS) module. The cell classification module 618 may include a fluorescence-activated cell classification (FACS) module. The cartridge 600 may include a housing 602 that makes the cartridge self-contained and optionally protects the contents from contamination. Sterile liquid transfer ports (SLTPs) 606a-606k may be fluidly coupled to storage sections 607a-607k. Each storage section may be a flexible bag or a rigid container. In some variations, a flexible bag can be configured to hold a large quantity of material and allow for fluid transfer, but it may collapse if fluid leaks out and expand if fluid flows in, so the transferred fluid will not be replaced by a liquid or gas to maintain pressure within the storage area.

[0153]

[0258] In some variations, the liquid transfer bus 624 includes valves V1 to V28 and may further include tubing that fluidically links these valves to each other and to each module. Valves coupled to four fluid lines as shown are 4 / 2 (4-port, 2-position) valves, and valves coupled to three fluid lines as shown are 3 / 2 (3-port, 2-position) valves. The internal flow paths of the valves are shown in the description. The cartridge may further include a first pump 632a and a second pump 632b. Each of these pumps has tubing exposed outside the housing 602, allowing each pump to interact with a pump actuator (e.g., rotor) in some fixtures of the system (e.g., work cell). The liquid transfer bus 624 may be fluidly coupled to a storage unit 607d and a product bag, which are fluidly coupled to the STLP 606d and product input tubing lines 627a to 627b. The operator can introduce cell products into the storage unit 607d by connecting the product input tube line 627a or 627b to an external cell source (e.g., a bag of cells collected from a donor). The SLTP 606d may be configured to allow a system according to this disclosure (e.g., work cell 110) to automatically add fluid to the storage unit 607d. For example, one or more fluid-carrying containers, such as storage units 607a-607k or bags, can receive fluid using the SLTP. Additionally or alternatively, the SLTP may be configured to periodically sample one or more of the fluid-carrying containers. The cartridge may further include collection bags 626a-626c fluidically coupled to the liquid transfer bus 624 via valves V17-V19. The cartridge 600 may be configured to allow the operator to remove the collection bags 626a-626c after the system has completed cell processing.

[0154]

[0259] Figure 7 is a schematic diagram of another variation of cartridge 700. For example, cartridge 700 may include fewer mechanism sets than cartridge 600. Cartridge 700 may include a fluid transfer bus 724, which may be fluidically coupled to a bioreactor module 714, a countercurrent centrifugation (CCE) module 722, and a module 716 selected from a cell selection module, a cell classification module, an electroporation module, or any other cell processing module. Cartridge 700 may include a housing 702 and sterile fluid transfer ports (SLTPs) 706a-706f (e.g., fluid connectors). Each of the SLTPs 706a-706f is fluidically coupled to storage sections 707a-707f, each of which can be a flexible bag or a rigid container. SLTP 706g is fluidically coupled to the bioreactor module 714, allowing the system or operator to directly access the bioreactor. The storage section 707c can be fluidically coupled to the SLTP 707c and the product input tubing line 727. In some variations, the liquid transfer bus 724 includes 14 valves V1-V3, V9, V11-V12, V17-V23, and V28, and may further include tubing that fluidly couples these valves to each other and / or to each module. The cartridge may further include collection bags 726a-726c that are fluidly coupled to the liquid transfer bus 724 via valves V17-V19. The cartridge may further include a pump 732. The pump 732 has tubing exposed outside the housing 702, allowing each pump to work in conjunction with a pump actuator in the system (e.g., a work cell).

[0155]

[0260] Figures 8A and 8B show side and top views of different variations of the cartridge, respectively. In some variations, the cartridge 800 may include a bioreactor 814, a pump 816, and a countercurrent centrifugation (CCE) module 822. The cartridge 800 may include empty spaces 818, 819, and 820 configured to accommodate one or more additional modules, such as a cell selection module, a cell classification module, an electroporation module, and a miniature bioreactor module. In some variations, the empty spaces may define an empty volume of the cartridge reserved for accommodating modules at another time. In some variations, the cartridge 800 may include two or more additional bioreactors and / or storage sections within the empty spaces 818, 819, and 820. Along near the surface of the cartridge 800, fluid connectors 806a to 806j (e.g., SLTPs) may be present, fluidically coupled to the storage sections 807a to 807f. Storage sections 807b and 807e may contain a fluid (e.g., buffer solution or culture medium). Along the top surface, there are product input tube lines 827a to 827d which can be fluidically connected to storage sections 807a, 807b, 807e, and 807f, respectively. The liquid transfer bus 824 can fluidly connect the STLP, storage sections, and product input tube lines to the module via tubing.

[0156]

[0261] In some variations, the external dimensions of the housing 802 may include approximately 225mm × 280mm × 385mm, 225mm × 295mm × 385mm, and 450mm × 300mm × 250mm, as well as all values ​​and sub-ranges between these. In some variations, the volume of the cartridge 800 may include approximately 10%, 20%, 30% or less, as well as all ranges and sub-values ​​between these. In some variations, the volume of the cartridge 800 may include approximately 10%, 20%, 30%, 50%, 100%, 200% or more, as well as all ranges and sub-values ​​between these.

[0157]

[0262] In some variations, the cartridge 800, as shown in the side view of Figure 8C and the perspective view of Figure 8D, may include a MACS module 818. For example, the bioreactor module 814 may include ports 815a to 815f. These include pH and dissolved oxygen (DO) sensors (ports 815a and 815b), a gas input line 815c, an output line 815d each having a sterile filter at the rear of the connector, and a coolant input line 815e and output lines 815f from the bioreactor instrument interface when working in conjunction with the bioreactor module 814 (for heat exchange). For example, the gas input line 815c may be configured for gas transfer to a fluid (e.g., headspace gas control or via a gas-permeable membrane).

[0158]

[0263] Figure 9 shows a side cross-sectional view of cartridge 900. In some variations, cartridge 900 may include an enclosure (e.g., housing), a bioreactor 914, one or more pumps 916, a valve 930, a cell selection module 917, and a countercurrent centrifugation ertriation (CCE) module 922. In some variations, the cell selection module 616 may be a magnetically activated cell selection (MACS) module 917. The cartridge may further include a collection bag 926. Optionally, cartridge 900 may include a blank space configured to accommodate one or more additional modules, such as a cell selection module, a cell classification module, or an electroporation module 918. In some variations, cartridge 900 may include one or more bioreactors and / or storage units within the blank space.

[0159]

[0264] In some variations, the cartridge may include one or more valves. In some variations, the valve 1000 on the cartridge may be configured to receive an actuator 1010 provided by the instrument (shown in Figure 10A). When the cartridge is inserted into the instrument, the valve 1000 may dock with the actuator 1010 (shown in Figure 10B) and be configured so that the valve 1000 switches from one position to another as the actuator 1010 rotates. In some variations, the valve may be configured to clamp a portion of a flexible tube. The pinch valve may include a closing configuration, and an external actuator may be configured to open and close the valve in conjunction with the pinch valve (e.g., using a linear-movement solenoid). The valve itself may be configured to be disposable, but the actuator may be integrated into an instrument configured to process cartridges repeatedly.

[0160] Reagent storage container

[0265] In some variations, the system includes one (or more) reagent storage containers. Within this container are stored reagents, including, but not limited to, cell culture media, buffers, cytokines, proteins, enzymes, polynucleotides, transfection reagents, non-viral vectors, viral vectors, antibiotics, nutrients, cryoprotectants, solvents, cytomaterials, and pharmaceutically acceptable excipients. Additionally or alternatively, waste materials may be stored within the reagent storage container. In some variations, samples being processed, extracted from one or more cartridges, may be stored within the reagent storage container. The reagent storage container may include one or more temperature-controlled compartments (e.g., a freezer, condenser, water bath, warming chamber, or other, with temperatures such as approximately -80°C, -20°C, 4°C, 25°C, 30°C, 37°C, and 42°C). The temperatures within these compartments can be varied during the cell manufacturing process to heat or cool the reagents. In variations of the method of this disclosure, cartridges may be moved to the reagent storage container by robot (or manually by an operator). The reagent storage container works in conjunction with one or more sterile fluid transfer ports on the cartridge to distribute reagents or substances into the cartridge. Optionally, fluid is added to or removed from the cartridge before, during, or after the addition or removal of reagents. In some variations, the system includes a sterile fluid transfer device, which is similarly configured to transfer fluid in and out of the cartridge automatically, manually, or semi-automatically. The operator can manually supply reagents to the sterile fluid transfer station, or a robot can supply reagents (e.g., from a feedthrough or other location). In some cases, the robot moves one or more reagents from the reagent storage container to the sterile fluid transfer station. The reagent storage container may optionally have an automated door to allow robotic access to sterile fluid transfer devices and / or other reagent storage containers, each under independent closed-loop temperature control. These devices and storage containers may be configured for pick-and-place movement by the robot. In some variations, the reagent storage container may include one or more sample pickup areas.For example, a robot may be configured to move one or more reagents between one or more sample pickup areas.

[0161]

[0266] A variety of materials can be used to construct the cartridge and cartridge housing, including metal, plastic, rubber, and / or glass, or combinations thereof. The cartridge, its components, and its housing can be formed by molding, machining, extrusion, 3D printing, or any combination thereof. The cartridge may include commercially available components (e.g., tubing, valves, accessories), which can be mounted on or integrated with custom components or devices. The cartridge housing may constitute an additional layer of enclosure to further protect the sterility of the cell products. The operator can perform cartridge loading or unloading in an ISO 5 or more advanced environment using aseptic techniques to ensure the sterility of the cartridge contents when the cartridge is open. In some variations, the operator can perform cartridge loading or unloading using manual aseptic connections (e.g., aseptic tube welding). Robotic systems can also perform aseptic loading and unloading of liquids into and from the cartridge using sterile liquid transfer devices and sterile liquid transfer ports on the cartridge.

[0162] Countercurrent centrifugal ertriation

[0267] Countercurrent centrifugal eltriation (CCE) is a technique used to separate cells based on characteristics such as size and / or density. Countercurrent centrifugal eltriation combines centrifugation and countercurrent eltriation. Centrifugation corresponds to the sedimentation process under the influence of a centrifugal force field, while countercurrent eltriation corresponds to the separation process by washing. Separation takes place in a conical (e.g., biconical, funnel-shaped) eltriation chamber. Two opposing forces act on particles (e.g., cells) in the fluid delivered to the eltriation chamber: a centrifugal force that propels the fluid away from the axis of rotation and a fluid velocity (e.g., countercurrent) that propels the fluid toward the axis of rotation. By varying the flow rate and centrifugal force, separation of particles (e.g., cells) can be achieved. For example, particles can be separated based on characteristics such as size and density, as will be described in more detail herein.

[0163]

[0268] Countercurrent centrifugation (CCE) can perform multiple operations useful for cell therapy manufacturing workflows, including, but not limited to, cell washing, cell concentration, medium / buffer exchange, transduction, and separation of leukocytes from other blood components (e.g., platelets and erythrocytes). In some variations, the fluid source for the cell separation process (e.g., apheresis bag) may include suspensions of leukocytes, erythrocytes, platelets, and plasma. To isolate target immune cells, leukocytes can be isolated and subsequently magnetically labeled for magnetic separation. The leukocyte separation step may be performed in a CCE module to separate cells based on size and density, and magnetic separation may be performed in a MACS module. In some variations, integrating the CCE module into a cartridge allows the cell processing system to separate cells based on one or more of the following: cell cycle progression (e.g., G1 / M phase cells, which are larger than G0, S, and G2 phase cells) and cell type (e.g., leukocytes versus erythrocytes and / or platelets).

[0164]

[0269] Generally, a rotor configured to rotate may include an ertriation chamber (e.g., a cone, a bicone). A fluid containing a suspension of cells can be injected into the rotor under continuous flow. Once the cells are introduced into the cone (e.g., a bicone), they move according to their sedimentation rate to a position on a gradient where the effects of two forces acting on the cells balance. Smaller cells with a low sedimentation rate (e.g., platelets) can be rapidly washed toward the axis of rotation as the flow velocity increases. Such small cells can be ejected from the cone (e.g., swept away). Relatively larger (or denser) cells (e.g., red blood cells) flow through the cone at a relatively slow speed, reaching equilibrium at the ertriation boundary where centrifugal and tensile forces balance, and the flow velocity is relatively slow as the cone expands. The largest or densest cells (e.g., white blood cells) remain near the chamber entrance where centrifugal force and fluid velocity are high. By gradually increasing the flow rate, progressively larger or denser cell fractions (e.g., platelets → red blood cells → white blood cells) can be continuously output from the rotor. By continuously increasing the flow rate, all cells are eventually elutriated from the cone.

[0165]

[0270] Figure 56 is a block diagram of a cell separation system 5600, which includes a work cell 5610 and at least one cartridge 5620. In some variations, the work cell 5610 may include one or more of the following: a countercurrent centrifugation ertriation (CCE) instrument 5632 (e.g., a first magnet), a magnetically activated cell selection (MACS) instrument 5642 (e.g., a magnet array, a second magnet), a fluid connector 5652, a pump 5654, an imaging system including an optical sensor 5660 and an illumination source 5662, a sensor 5664, and a processor 5670. In some variations, the cartridge 5620 may include one or more of the following: a CCE module 5630 (e.g., a rotor), a MACS module 5640 (e.g., a flow cell), and a fluid connector 5650 (e.g., a sterile fluid transfer port, a fluid transfer bus). For example, a cartridge for cell processing may include a fluid transfer bus and multiple modules, each module being fluidically linked to the fluid transfer bus. These modules may include any of the CCE modules or MACS modules described herein. In some variations, a robot (not shown) may be configured to move the cartridge 5620 between various positions within the work cell 5610 to perform various cell processing steps.

[0166]

[0271] In some variations, the imaging system (e.g., optical sensor 5660, illumination source 5622) may be configured to generate image data corresponding to one or more of the CCE module 5630 and MACS module 5640. For example, as detailed herein, image data of the fluid flow through the rotor of the CCE module 5630 can be used to analyze the fluid flow rate and / or the rotational speed of the rotor. In some variations, the optical sensor 5660 may be, for example, a CMOS / CCD sensor with a resolution of about 100 μm, a working distance of about 40 mm to about 100 mm, and a focal length of less than about 8 mm. The optical sensor 5660 may be configured to operate synchronously with the illumination source 5662. In some variations, the optical sensor 5660 may include one or more of a colorimeter, a turbidity sensor, and an optical density sensor. In some variations, the illumination source 5662 may operate as a stroboscopic light source configured to output light pulses synchronized with the rotational speed of the rotor of the CCE module 5630.

[0167]

[0272] In some variations, the sensor 5664 may include one or more of the following: an optical density sensor configured to measure the intensity of a fluid; a leak detector configured to detect moisture and / or leaks; an inertial sensor configured to measure vibrations; a pressure sensor (e.g., a photoelectron sensor) configured to measure pressure in a fluid line; a bubble sensor configured to detect the presence of bubbles in a fluid tube; a colorimetric sensor; a vibration sensor, etc.

[0168]

[0273] In some variations, the fluid connector 5652 may include one or more valves configured to control the fluid flow between the work cell and the cartridge 5620. The processor 5670 may correspond to a controller (e.g., a processor and memory) as detailed herein. The processor 5670 may be configured to control one or more of the following: the CCE device 5632, the MACS device 5642, the pump 5654, the fluid connector 5652 (e.g., valves), the optical sensor 5660, the illumination source 5662, and the sensor 5664.

[0169]

[0274] In some variations, the cell processing system 5600 may comprise a cartridge 5600 containing a rotor of a CCE module 5630 configured for countercurrent centrifugation of cells in a fluid. A first magnet of the CCE instrument 5632 may be configured to magnetically rotate the rotor to separate the cells from the fluid in the rotor. The cartridge may further comprise a flow cell of a MACS module 5640, which is coupled to the rotor and configured to receive cells from the rotor. A second magnet of the MACS instrument 5642 may be configured to magnetically separate the cells in the flow cell.

[0170]

[0275] In some variations, the illumination source 5662 may be configured to illuminate cells. The optical sensor 5660 may be configured to generate image data corresponding to cells. In some variations, the system 5600 may include one or more of the following: an oxygen deficiency sensor, a leak sensor, an inertia sensor, a pressure sensor, and a bubble sensor. In some variations, the system 5600 may include one or more valves and pumps.

[0171]

[0276] Figure 57 is a side cross-sectional view of a counterflow centrifugal ertriation (CCE) module 5700. The CCE module 5700 includes a housing 5710 (e.g., an enclosure), a rotor 5720 configured to rotate relative to the housing 5710, and one or more fluid ports 5730 (e.g., a fluid inlet, a fluid outlet). In some variations, the CCE module 5700 may be portable and configured to move within a work cell 5610 and a cartridge 5620. For example, a robot may move the CCE module 5700 between various fixtures in the work cell 5610.

[0172]

[0277] Figure 58 is a side cross-sectional view of a magnetically activated cell selection (MACS) module. The MACS module includes a housing 5810 (e.g., enclosure), a first fluid port 5820 (e.g., fluid inlet), a second fluid port 5830 (e.g., fluid outlet), and a flow cell 5810 coupled between the first fluid port 5820 and the second fluid port 5830. As detailed herein, the flow cell 5810 may include a cavity (e.g., chamber) containing one or more channels (e.g., linear channels, layered fluid flow channels). In some variations, the cavity of the flow cell 5810 may be substantially empty. For example, the flow cell 5810 may not have mesh, beads, meandering channels, etc. In some variations, the flow cell 5810 may have a longitudinal axis aligned perpendicular to the ground. In other words, the flow cell 5810 is oriented vertically, and the first fluid port 5820 is positioned at a higher elevation than the second fluid port 5830, so that gravity can facilitate the fluid flow through the flow cell 5810. In some variations, the MACS module 5800 is portable and can be configured to move within the work cell 5610 and cartridge 5620. For example, a robot can move the MACS module 5630 between various fixtures in the work cell 5610.

[0173]

[0278] Figures 59A and 59B are perspective views of a system 5900 for cell processing (e.g., a CCE system). The system 5900 comprises a CCE module 5930 (e.g., a cartridge) including a housing 5931 and a rotor 5910, a CCE instrument 5932, an optical sensor 5960, and an illumination source 5962. In some variations, the CCE instrument 5932 may include a magnet configured to magnetically rotate the rotor 5910 within the CCE module 5930. To facilitate illumination by the illumination source 5962 and image data generation by the optical sensor 5960, one or more portions of the housing 5931 and rotor 5910 can be made optically transparent.

[0174]

[0279] In some variations, the cell processing system 5900 may comprise a cartridge 5930 including a housing 5931 containing a rotor 5910 configured to separate cells from a fluid. An instrument 5932 containing a magnet may be configured to magnetically rotate the rotor 5910 in conjunction with the cartridge 5930. The cartridge 5930 may be configured to transfer cell products between multiple instruments. In some variations, the housing 5931 can house the rotor 5910. In some variations, the housing 5931 may include one or more openings 5937 configured to facilitate visualization (e.g., imaging) of the rotor 5910. Figures 59A and 59B show the magnet 5932 in proximity to but not mounted on the housing 5931. Figure 59C is a perspective view of the rotor 5910 and housing 5931, where the magnet 5932, optical sensor 5960, and illumination source 5962 are not shown.

[0175]

[0280] In some variations, the cartridge 5930 (e.g., housing 5931, 5910) may include consumable components such as disposable components, limited-use components, or single-use components. In some variations, the magnet 5932 may include durable components that can be reused multiple times. In some variations, the magnet 5932 may be detachably coupled to the housing 5931. For example, the housing 5931 can be moved relative to the magnet 5932 to facilitate magnetic coupling between the magnet 5932 and multiple cartridges 5930. Additionally or alternatively, the configuration may be such that the magnet 5932 is moved relative to the housing 5931.

[0176]

[0281] Figure 59D is a side cross-sectional view of the CCE module 5930. In some variations, the housing 5931 of the rotor 5910 may include a first side 5933 containing a first fluid port 5912 (e.g., a first fluid tube) and a second side 5935 containing a second fluid port 5914, the second side 5935 being opposite the first side 5933. The rotor 5910 (including a conical or biconical rotor as detailed herein) may be coupled between the first fluid port 5912 and the second fluid port 5914. In some variations, the CCE module 5930 may include an air gap 5902 between the housing 5931 and the magnet 5932. That is, the cartridge 5930 and the magnet 5932 may be coupled non-contact. As a result, the cartridge does not need to be mechanically coupled with the magnet 5932 to perform countercurrent centrifugal eltriation. Therefore, the rotor 5910 has low alignment sensitivity with the magnet 5932, and vibrations between the rotor 5910 and the magnet 5932 may also be small. Furthermore, the space between the rotor 5910 and the magnet 5932 allows the second fluid port 5914 to extend near the second side 5935 of the housing 5931, thereby enabling fluid to flow on each side of the rotor 5910.

[0177]

[0282] In some variations, countercurrent centrifugal eltriation can be performed by moving the magnet 5932 toward (or toward) the rotor 5910 by the system 5900. The rotor may define an axis of rotation (e.g., coaxial with the first fluid port 5912 and the second fluid port 5914). Fluid can flow through the rotor through the first fluid port 5912 and the second fluid port 5914. While fluid is flowing through the rotor 5910, the magnet 5932 can magnetically rotate the rotor about the axis of rotation. The rotor can move toward the magnet. For example, moving the rotor 5910 may involve using a robot (not shown) to move the rotor 5910 forward and backward relative to the magnet 5932.

[0178]

[0283] In some variations, the fluid can flow into the rotor 5910 through a first fluid port 5912 along a first side 5933 of the rotor 5910. After counterflow centrifugal etractoration via the rotor 5910, the fluid can flow out of the rotor 5910 through a second fluid port 5914 along a second side 5935 of the rotor 5910.

[0179]

[0284] In some variations, countercurrent centrifugal ertriation can be visualized by an optical sensor 5960 and an illumination source 5962 to manage and modify cell separation in real time based on predetermined criteria in a closed loop, with the aim of maximizing ertriation efficiency. In some variations, the optical sensor 5960 may be configured to image any part of the rotor through which the fluid flows (e.g., a first fluid tube, a second fluid tube, a third fluid tube, a first bicone, a second bicone). For example, the optical sensor 5960 can be used to generate image data of one or more fluids and cells within the rotor 5910. In some variations, the illumination source 5962 can be used to illuminate one or more fluids and cells. For example, the optical sensor can image the output of a cone to identify non-target cells on which ertriation is being performed.

[0180]

[0285] In some variations, one or more of the rotor rotation speed and fluid flow rate can be selected, at least partially based on image data. For example, the rotor may have a rotation speed of up to 6,000 RPM. For example, the fluid may have a flow rate of up to approximately 150 ml / min while the rotor is rotating. In some variations, the rotor may be moved toward the illumination source 5962 and the optical sensor 5960. Additionally or alternatively, the rotor 5910 may be moved toward the area away from the illumination source 5962 and the optical sensor 5960.

[0181]

[0286] Figure 59E is a side cross-sectional view of the rotor 5910 including a first fluid port 5912 (e.g., fluid tube, inlet) and a second fluid port 5914 (e.g., fluid tube, outlet). In some variations, the first fluid port 5912 and the second fluid port 5914 may extend toward each other and / or parallel to the axis of rotation of the rotor 5910. In some variations, the first fluid port 5912 and the second fluid port 5914 are located on opposite sides of the rotor 5910, thereby simplifying the design of the fluid routing and cartridge, and further reducing manufacturing costs. For example, each of these ports contains only one lumen, thus simplifying the fluid seal. Conventionally, a drive motor is fixed and mechanically coupled to the second side of the rotor, so a complex fluid path (including inlets and outlets) is formed on the first side of the rotor. Figures 59F and 59G are side cross-sectional views of the rotor 5910 located within a housing 5931.

[0182]

[0287] Figure 60A is a plan view of a rotor 6000 that may be used with any of the CCE systems, CCE modules, cartridges, housings, and combinations thereof described herein. The rotor 6000 may include a first fluid tube 6010, a cone 6020 (e.g., a bicone), a second fluid tube 6030, a magnetic section 6040 (e.g., magnets), and a housing 6050. Fluid may flow sequentially through the first fluid tube 6010, the cone 6020, and the second fluid tube 6030. In some variations, the magnetic section 6040 may include one or more magnets. In some variations, the rotor 6000 may define a rotation axis 6060. In some variations, at least a portion of the first fluid tube 6010 and at least a portion of the second fluid tube 6030 may extend parallel to the rotation axis (e.g., in a direction that penetrates the plane of paper in Figure 60A). In some variations, at least a portion of the first fluid tube 6010 and at least a portion of the second fluid tube 6030 may be coaxial.

[0183]

[0288] In some variations, the cone 6020 may include a bicone having a first cone with a first base and a second cone with a second base, wherein the first base faces the second base. In some variations, the bicone may include a cylinder (or any other shape) that lies between and / or is in fluid communication with the first and second cones. For example, one or more cones of the rotor may include a generally stepped shape. For example, one or more cones may include stacked circular steps. In some variations, the cone of the rotor may include a single cone.

[0184]

[0289] In some modifications, at least a portion of the rotor can be made optically transparent to facilitate visualization and / or imaging of the rotor 6000 and / or the fluid (e.g., cells) within the rotor 6000. For example, the cone 6020 and the portions of the first fluid tube 6010 and the second fluid tube 6030 can be made transparent.

[0185]

[0290] In some variations, the cone can contain a volume of approximately 10 ml to approximately 40 ml. In some variations, the cone can have a cone angle of approximately 40 degrees to approximately 60 degrees.

[0186]

[0291] In some variations, the cone includes a first cone (e.g., a distal cone) and a second cone (e.g., a proximal cone), and the first cone can be larger than the second cone. In some variations, the length of the first cone may be approximately 60 mm to 90 mm. In some variations, the length of the proximal cone may be approximately 15 mm to 40 mm. In some variations, the diameter of the cone (e.g., the maximum diameter of the cone) may be approximately 15 mm to 40 mm.

[0187]

[0292] In some variations, the rotor 6000 may have an asymmetrical shape. In some variations, a first part of the rotor 6000 (e.g., a first end) may include a cone 6020, and a second part (e.g., a second end) may include a paddle shape.

[0188]

[0293] In some variations, the cone can have a length of at least about 4 cm (e.g., about 9 cm to about 12 cm), a cone diameter of about 5 cm or less (e.g., about 3 cm to about 5 cm), a fluid flow rate of up to about 100 ml / min (e.g., about 60 ml / min to about 100 ml / min), and a rotational speed of less than about 3000 RPM. The shapes of the first and second cones can be generally linear (as opposed to convex or concave).

[0189]

[0294] Figures 60B and 60C are perspective views of the rotor 6002, including the first fluid tube 6012, the cone 6022, the second fluid tube 6032, and the housing 6052, while Figure 60D is a side view. Figure 60E is a perspective view of the rotor 6002 positioned within the housing 6090.

[0190]

[0295] Figure 60F is a plan view of a rotor 6004 having two cones (e.g., two bicones). The rotor 6004 is configured to ertriate cells (e.g., erythrocytes, leukocyte-removed transfusion products) in a second cone for the purpose of recirculating buffer for reuse. The rotor 6004 may include a housing 6052, a first fluid tube 6012, a first cone 6022 coupled to the first fluid tube 6012, a second fluid tube 6023 coupled to the first cone 6022, a second cone 6024 coupled to the second fluid tube 6023, and a third fluid tube 6032 coupled to the second cone 6024. The first cone 6022 may contain a first volume, and the second cone 6024 may contain a second volume that is larger than the first volume. In some variations, the ratio of the second volume to the first volume can be approximately 2:1 to approximately 5:1. The fluid can flow sequentially through the first fluid tube 6012, the first cone 6022, the second fluid tube 6023, the second cone 6024, and the third fluid tube 6032. In some variations, the rotor 6004 may include a magnetic portion 6042.

[0191]

[0296] In some variations, the first cone 6022 may include the first bicone, and the second cone 6024 may include the second bicone. In some variations, the first bicone may include a third cone including the first base and a fourth cone including the second base opposite the first base. In some variations, the second bicone may include a fifth cone including the third base and a sixth cone including the fourth base opposite the third base.

[0192]

[0297] In some variations, parts of the rotor 6004, such as the first cone 6022, the second cone 6024, and at least portions of the first fluid tube 6012, the second fluid tube 6023, and the third fluid tube 6032, may be optically transparent. In some variations, the first fluid tube 6012 may include an inlet, and the third fluid tube 6032 may include an outlet.

[0193]

[0298] In some variations, cells enter the first cone 6022 and eltriation of red blood cells (RBCs) 6030 can be performed in the second cone 6024. Since the second cone 6024 is spread far from the axis of rotation (center of the housing 6052), the RBCs 6030 can be concentrated at the inlet 6025 of the second cone 6024 by centrifugation. Because the volume of the second cone 6024 is large, the velocity of the fluid (e.g., buffer) is reduced, which can reduce the force acting on the RBCs 6030 in the second cone 6024. By recirculating the fluid (e.g., buffer), eltriation of high concentrations of RBCs can be performed using less fluid (e.g., buffer). In some variations, white blood cells 6040 can be collected from the first cone 6022. An optical sensor may be configured to image the first cone 6022 to generate imaging data used to identify the boundary between the WBCs 6040 and the RBCs 6030. In some variations, it is possible to remove small particles (e.g., platelets) using less fluid (e.g., buffer solution) by passing the recirculated fluid through a filter.

[0194]

[0299] Figure 60G is a plan view of rotor 6005 having two cones (e.g., two bicones) and configured to perform eltriation of cells (e.g., red blood cells) in the second cone, and Figure 60H is a side view. The rotor having two cones can facilitate the recirculation of the buffer for reuse. Rotor 6006 may include a housing 6052, a first fluid tube 6012, a first cone 6022 coupled to the first fluid tube 6012, a second cone 6024 coupled to the first cone 6022, and a fluid tube 6032 (e.g., an outlet) coupled to the second cone 6024.

[0195]

[0300] Figure 60I is a perspective view of rotor 6006 including cone 6024 and housing 6054. Figure 60J is a perspective view of rotor 6007 including cone 6026 and housing 6056. Figure 60K is a schematic plan view of rotor 6008 and its corresponding dimensions. Figure 60L is an image of a set of rotors with various dimensions.

[0196]

[0301] Figures 11A to 11C show another variation of the countercurrent centrifugal ertriation (CCE) module 1100. Figure 11A is a perspective view of the cartridge 1110 including the CCE module 1100 in an extended configuration configured to receive a CCE instrument. Figures 11B and 11C are side cross-sectional views of the CCE module 1100 in a retracted configuration and an extended configuration, respectively. In some variations, the CCE module may include a conical element having inner and outer surfaces, fixedly attached to the distal end of a linear member having inner and outer surfaces. The proximal end of the linear member may be rotatably mounted to a pivot point to allow the linear member to expand, retract, and / or rotate. For example, Figure 11C shows the linear member expanding out of the cartridge housing and then rotating to generate centrifugal force. Cell products can be transported to a conical element through the inner and outer surfaces of the linear member (optionally located inside the tube) against the centrifugal force generated by the rotation of the linear member, and supplied to the opening at the distal end of the inner surface of the conical element. Due to the backflow of the solution and the sedimentation of cells under centrifugal force, cells in the cell product can be separated based on the ratio of hydrodynamic cross-section to mass. Then, by increasing the flow rate and / or decreasing the rotation of the linear member, cells can be selectively returned to the proximal end of the linear member through the void on the inner surface of the linear member. These selected cells can be guided into a tube that returns them to the cartridge. After performing the concentration and / or washing steps, the linear member can be stored in the housing in the storage configuration shown in Figure 11B.

[0197] Magnetic cell selection

[0302] In general, the systems and methods described herein allow for the selection of cells based on magnetically labeled cells corresponding to cells having a given antigen. For example, a cell suspension of interest can be immunologically labeled with magnetic particles (e.g., magnetic beads) configured to selectively bind to the surface of the target cells. When the cell suspension flows through a flow cell, the labeled cells can generate a large magnetic moment. The flow cell may be positioned in close proximity to a magnetic array (e.g., permanent magnets, electromagnets). This magnetic array generates a gradient magnetic field within the flow cell to attract the labeled cells for separation, capture, recovery, and purification. The magnetic array may cover the entire volume of the flow cell by generating non-uniform magnetic fields at the edges and interfaces of the individual magnets, and may be configured such that the tensile force applied by the fluid flowing through the flow cell is equal to the magnetic force.

[0198]

[0303] Figures 61A to 61C are schematic diagrams of a magnetic cell separation (e.g., magnetically activated cell selection) system and process. The magnetic cell separation system may include a flow cell 6110 with an inlet 6130 and an outlet 6132, a magnetic array 6120, a first fluid source 6140 (e.g., input sample source), a second fluid source 6142 (e.g., buffer source), a third fluid source 6150 (e.g., target cell storage), a fourth fluid source 6152 (e.g., waste storage), and a valve set 6134. As shown in step 6100, cell sets 6160, 6170, including labeled cells 6160 (e.g., magnetically labeled cells) and unlabeled cells 6170, can flow into the flow cell 6110. For example, cell set 6160 can be labeled with a magnetically activated cell selection (MACS) reagent. The MACS reagent can be cultured in the cell set to label (e.g., attach, bind) cells to the MACS reagent. As detailed herein, the magnetic array 6120 can be positioned outside the flow cell 6110 so as to be movable relative to the flow cell 6110. For example, the magnetic array 6120 may move away from the flow cell 6110 to facilitate the outflow of the cell set 6160 from the flow cell 6110. Conventional flow cells have meandering pathways containing mesh and / or beads for capturing cells. However, it is difficult to recover labeled cells from conventional flow cell configurations. In contrast, the flow cell 6110 described herein does not include meandering pathways such as beads and mesh, and therefore allows for efficient and continuous selection using either positive or negative selection. In some variations, the flow cell may include generally layered channels as detailed herein.

[0199]

[0304] In step 6102, the magnetic array 6120 may magnetically attract the cell set 6160 toward the magnetic array 6120 over a predetermined residence time and / or based on the measured amount of magnetically separated cells. In some variations, the residence time can be at least 1 minute (e.g., at least 2 minutes, at least 3 minutes, at least 5 minutes). Unlabeled cells 6170 are not magnetically attracted to the magnetic array 6120 and may exit the flow cell 6110 through the outlet 6132 and flow into the fourth fluid source 6152. In some variations, the fluid (e.g., cells 6160, 6170) within the flow cell may be statically held within the flow cell 6110 for a certain residence time before the fluid (e.g., cells 6170) flows out of the outlet 6132. In some variations, the longitudinal direction of the flow cell 6110 can be oriented substantially perpendicular to the ground so that the fluid flow through the flow cell 6110 is facilitated by gravity. In step 6104, the magnetic coupling between the magnet array 6120 and the cells 6160 is released after the residence time, allowing the cells 6160 to flow into the third storage unit 6150.

[0200]

[0305] In some variations, due to static friction, cells may remain attached to the surface of the flow cell even after the magnet array 6120 has been removed. Therefore, gas may be flowed through the flow cell 6110 to assist in the collection of cells into the third storage unit 6150. A gas flow through the flow cell can improve cell recovery more than a forceful flow of liquid through the flow cell. Since the interface created by the gas (e.g., bubbles, air gap) is maintained by gravity, it is possible to implement relatively wide flow cells, which can further improve cell recovery than flow cells arranged horizontally. The MACS modules described herein may be configured to perform positive and / or negative selection by changing the order of the steps.

[0201]

[0306] Additionally or alternatively, an optical sensor may be configured to image the flow cell to generate imaging data used to identify the amount of cells magnetically attracted to the magnetic array. When a predetermined amount of cells is measured by the optical sensor, the fluid containing the labeled cells can be drained out of the flow cell.

[0202]

[0307] Figure 62A is a perspective view of MACS module 6200 in a first configuration. MACS module 6200 (and any of the MACS modules described herein) can be a component of any of the cartridges described herein. For example, a cartridge for cell processing may include a liquid transfer bath and several modules, each module being fluidically linked to the liquid transfer bath. MACS module 6200 may include a flow cell 6210, an inlet 6230, and an outlet 6232, each containing an elongated cavity with a certain cavity height. MACS module 6200 may further include a magnet array 6220 containing several magnets. Each magnet may be separated by a certain separation distance, as shown in Figures 62G, 63D, and 63E, but the magnets in the magnet array 6220 shown in Figures 62A to 62E are in contact with adjacent magnets.

[0203]

[0308] Figure 62G is a schematic diagram of a flow cell 6210 and a magnet array 6220. In some variations, the flow cell 6210 may include a cavity height 6202 and a cavity width 6204. The fluid may be configured to flow through the flow cell 6210 in a first direction 6206. The magnet array 6220 may include a plurality of magnets, each having a width 6222. In some variations, adjacent magnets may be separated by a predetermined separation distance 6224. Each magnet pair may have the same or different separation distances 6224. As shown in Figure 62G, the orientation (e.g., poles) of the magnets in the magnet array 6220 may include a predetermined pattern.

[0204]

[0309] In some variations, the ratio of cavity height 6202 to separation distance 6224 may include approximately 20:1 to approximately 1:20, approximately 10:1 to approximately 1:10, approximately 5:1 to approximately 1:5, and approximately 3:1 to approximately 1:3, as well as all values ​​and subranges in between. In some variations, the actuator 6240 (e.g., linear, rotary) may be configured to move the magnet array 6220 relative to the flow cell 6210. In some variations, the orientation (e.g., poles) of the magnets in the magnet array 6220 may include a predetermined pattern (e.g., Halbach array).

[0205]

[0310] In some variations, the magnet array 6220 can move relative to the flow cell 6210 or vice versa. Figure 62A shows the MACS module 6200 in an open configuration, and Figure 62B shows the MACS module 6200 in a closed configuration. Figure 62B is a perspective view of the MACS system 6200 in a second configuration in which labeled cells can be magnetically attracted toward the magnet array 6220. In the second configuration, magnetic field lines generated by the magnet array traverse the flow channel, applying magnetophoresis to the magnetically labeled cells injected into the channel. Figure 62C is a side cross-sectional view of the MACS system 6200 including the magnet array 6220. Figure 62D is a perspective view of the MACS system 6200 in the second configuration. Figure 62E is a plan view of the flow cell 6210 and magnet array 6220 of the MACS system. Figure 62F is a plan view of the flow cell 6210 of the MACS system.

[0206]

[0311] Figures 63A to 63E are perspective views of sets 6300, 6310, 6320, 6330, and 6340 of magnet arrays. One or more of the size, intensity, shape, spacing, and orientation of the magnets in the magnet array can be set to generate a magnetic field that attracts magnetically labeled cells. Additionally or alternatively, the magnet array may include a high-permeability material configured to increase or decrease the magnetic field intensity and magnetic field gradient within the flow cell. This material may be placed between the magnets and the flow cell. Additionally or alternatively, this material may be placed within and / or on one or more sides of the flow cell.

[0207]

[0312] Figures 64A and 64B are perspective and side cross-sectional views, respectively, of the MACS module 6400, which includes a flow cell 6410 and a magnet array 6420. The flow cell 6410 includes a set of linear channels 6412, 6414, and 6416, which include a first channel 6412 parallel to a second channel 6416, and a third channel 6416 in fluid communication with each of the first channel 6412 and the second channel 6416. As shown in Figure 64B, the third channel 6416 is positioned between the first channel 6412 and the second channel 6414 to define a volume in which the fluids from the first channel 6412 and the second channel 6416 interact (e.g., mix). In some variations, the flow cell 6410 may include a first inlet 6430 that is coupled to a first channel 6412 and configured to receive a first fluid 6460 (e.g., cells). A second inlet 6431 may be coupled to a second channel 6414 and configured to receive a second fluid 6470 (e.g., buffer solution). The flow cell 6410 may also include a first outlet 6432 coupled to the first channel 6412 and a second outlet 6433 coupled to the second channel 6414.

[0208]

[0313] The magnet array 6420 is positioned outside the flow cell 6400 and can be moved relative to the flow cell 6400 as described herein. In some variations, the longitudinal axis of the flow cell 6410 can be perpendicular to the ground so that the fluid flows in a generally vertical direction.

[0209]

[0314] In some variations, the first channel 6412 may have different dimensions from the second channel 6414. For example, the first cavity height of the first channel 6412 may be greater than the second cavity height of the second channel 6414. For example, the ratio of the first cavity height to the second cavity height may include approximately 1:1 to approximately 3:7, approximately 1:1 to approximately 2:3, and approximately 2:3 to approximately 3:7, as well as all values ​​and subranges in between. Because the cavity height of the first channel 6412 is greater than that of the second channel 6414, the fluid flowing through the first channel 6412 may have a slower flow velocity than that of the second channel 6414. In some variations, the third channel 6416 may include the length and diameter of the third channel 6416, and this ratio may include approximately 2:1 to approximately 6:1, approximately 2:1 to approximately 3:1, approximately 3:1 to approximately 4:1, approximately 4:1 to approximately 5:1, approximately 5:1 to approximately 6:1, and approximately 3:1 to approximately 5:1, as well as all values ​​and subranges in between.

[0210]

[0315] As shown in Figure 64B, the first fluid 6462 can flow through the flow cell 6410 in a generally first direction. The magnetic attraction force generated by the magnet array 6420 pulls the magnetically labeled cells 6416 in the first fluid 6462 from the first channel 6412 towards the second channel 6414 (for example, towards the magnet array 6420), so that the cells 6416 can separate from the rest of the first fluid 6462 in the third channel 6416. Similarly, the second fluid 6470 (for example, buffer solution) can flow through the second channel 6414. As the cells 6416 flow towards the magnet array 6420, they displace the second fluid 6470 flowing through the third channel 6416 so that a portion of it can flow into the first channel 6412. In this way, the magnetically labeled cells 6416 are magnetically separated from the first fluid 6462, and the second fluid 6470 can facilitate the removal of the first fluid 642 that does not contain the cells 6416.

[0211]

[0316] In some variations, a set of fluid loops can be coupled to the flow cell to enable multiple cell separation cycles. Figure 64C is a schematic diagram of a MACS module including a flow cell 6410, a first fluid tube 6480 coupled to the inlet 6430 and outlet 6432 of the flow cell 6410. The first fluid tube 6480 may be configured to receive a cell set from the outlet 6432 of the flow cell 6410 and recover and / or recirculate it via the inlet 6430 of the flow cell 6410. A second fluid tube 6490 may be coupled to the inlet 6431 and outlet 6433 of the flow cell 6410 to recirculate fluids such as buffer and unrecovered magnetically labeled cells. The second fluid tube 6490 may be configured to receive fluids from the flow cell 6410 that do not contain a cell set. Based on the number of cycles performed, higher purity labeled cells can be recovered. For example, one cell separation cycle may produce approximately 80% cell purity, a second cell separation cycle approximately 96%, a third cell separation cycle approximately 99.2%, and a fourth cell separation cycle approximately 99.84%.

[0212]

[0317] In some variations, centrifugal force can be applied to the magnetic cell separation process independently of the fluid flow rate to further attract labeled cells towards the magnetic array in order to maintain throughput. Figures 65A to 65C are schematic diagrams of MACS module 6500 that use centrifugal force to facilitate the cell separation process. Figure 65A shows a flat flow cell 6510 configured to be wound to form a roughly cylindrical shape 6512. The flow cell 6510 may include a curved channel 6520.

[0213]

[0318] Figure 65B shows a cylindrical flow cell 6510 concentrically surrounded (e.g., nested within) a cylindrical magnet array 6530. In Figure 65B, only a cross-section of the magnet array 6530 is shown for clarity. The flow cell 6510 can be separated from the magnet array 6530 by a predetermined distance. Thus, the flow cell 6510 can be configured to rotate about its longitudinal axis (6550) to generate an outward centrifugal force toward the magnet array 6530 relative to the fluid 6540 in the flow path 6520. During the cell separation process, the fluid may be subjected to the set of forces shown in Figure 65C. These forces include bulk fluid forces 6560 in the axial direction (e.g., bulk flow), centrifugal forces 6570 outward from the center of rotation (e.g., proportional to the net buoyancy of the particle system), and magnetic forces 6580 outward from the center of rotation (e.g., proportional to the net magnetic attraction of the particle system). In some variations, labeled cells may have a higher density than unlabeled cells. Therefore, the centrifugal force preferentially pushes the labeled cells toward the magnet 6530, further increasing the specificity and efficiency of cell separation.

[0214]

[0319] Figures 66A to 66C are schematic diagrams of a cell separation system and process. The magnetic cell separation system may include a flow cell 6610 containing a channel 6620 (shown schematically flattened for clarity) and a magnetic array 6630. As shown in step 6600, cell sets 6640, 6642, containing labeled cells 6640 (e.g., magnetically labeled cells) and unlabeled cells 6642, can flow into the channel 6620 of the flow cell 6610. For example, cell set 6640 can be labeled with a magnetically activated cell selection (MACS) reagent. The magnetic array 6630 may be positioned outside the flow cell 6610 so as to be movable relative to the flow cell 6610. For example, the magnetic array 6630 can be moved away from the flow cell 6610 to facilitate the outflow of cell sets 6640 from the flow cell 6610.

[0215]

[0320] In step 6602, the flow cell 6650 can be rotated to generate centrifugal force that pushes the cells 6640, 6642 toward the magnet array 6630. In some variations, the longitudinal direction of the flow cell 6610 can be oriented substantially perpendicular to the ground so that the fluid flow through the flow cell 6610 is facilitated by gravity. In step 6604, the magnet array 6630 can magnetically attract the cell set 6640 toward the magnet array 6630 for a predetermined residence time as described herein. Unlabeled cells 6642 are not magnetically attracted to the magnet array 6630 and can exit the flow cell 6610 and flow into, for example, a waste container. In some variations, the fluid (e.g., cells 6160, 6170) within the flow cell can be statically retained within the flow cell 6110 for a residence time before the fluid (e.g., cells 6170) flows out of outlet 6132. In some variations, the magnetic coupling between the magnet array 6630 and the cells 6640 is released after a residence time, allowing the cells 6640 to be harvested.

[0216]

[0321] Figures 12A and 12B show the magnet and MACS module 1210 of the MACS device 1200, which includes the magnet. The magnet is shown in the ON configuration in Figure 12A and in the OFF configuration in Figure 12B.

[0217] Bioreactor

[0322] The bioreactors described herein may include vessels configured for culturing mammalian cells. Generally, cells and gene therapy products can be grown within a bioreactor to produce clinical volumes that can later be administered to patients. The growth rate and achievement of cell growth can be optimized by controlling numerous biological and environmental factors. The bioreactor modules described herein enable one or more of the monitoring, regulation, and / or control of cell growth (for example, to facilitate consistent and efficient cell proliferation).

[0218]

[0323] Figure 67A is a schematic diagram of a cell processing system 6700 (e.g., a bioreactor module) comprising one or more of the following: a bioreactor 6710, one or more sensors 6720, a stirrer 6730, a temperature controller 6740, and a gas controller 6750. In some variations, the sensors 6720 may be configured to monitor (e.g., measure, detect, determine) one or more features of the bioreactor module 6700 and the cells within the bioreactor 6710. For example, the sensors 6720 may include one or more of the following: a pH sensor, a dissolved oxygen (DO) sensor, a temperature sensor, a glucose sensor, a lactose sensor, a cell density sensor, a humidity sensor, or a combination thereof. One or more of the sensors may be non-invasive optical sensors.

[0219]

[0324] Figures 67B to 67D are schematic diagrams of a cell processing system including a work cell 6760, a bioreactor system 6700 (e.g., a bioreactor instrument), a cartridge 6770, a stirrer 6730, and a fluid connector 6780. In some variations, the cartridge 6770 for cell processing may include a fluid transfer bus and several modules (e.g., a bioreactor module, a CCE module, a MACS module, an EP module). Each module may be fluidically linked to the fluid transfer bus. The bioreactor module may include at least one bioreactor.

[0220]

[0325] The bioreactor apparatus 6700 may be configured to work in conjunction with the cartridge 6770. In some variations, the bioreactor apparatus 6700 may include a stirrer 6730 configured to connect to the bioreactor. The stirrer may be configured to agitate a cell medium containing cells. In some variations, the fluid connector 6780 may be configured to connect the bioreactor system 6700 and the work cell 6760.

[0221]

[0326] Figure 67B illustrates a cartridge 6770 containing a bioreactor placed within a work cell 6760. The bioreactor 6700 can be separated from the work cell 6760. As shown in Figure 67C, once the fluid connector 6780 connects the work cell 6760 to the bioreactor 6700 (e.g., to create a sterile channel), the cartridge 6770 can be moved into the bioreactor 6700. For example, as shown in Figure 67D, the cartridge 6770 can be connected to a stirrer 6730 (e.g., placed on the stirrer 6730) and then stirred. In some variations, the fluid connector 6780 may include a set of foldable sidewalls (e.g., accordion-like) configured to receive and disperse the stirring of the stirrer 6730 while preventing such movement from being transmitted to the work cell 6760. That is, the fluid connector 6780 may function as a bellows section to maintain the connection between the work cell 6760 and the bioreactor 6700 without stirring the work cell 6760. In some variations, the fluid connector 6780 can connect a bioreactor (e.g., cartridge 6770) to a liquid transfer bus.

[0222]

[0327] In some variations, the agitator may be configured to generate motion (e.g., orbital, rotational, linear) relative to the bioreactor to mix the culture when it is necessary to promote the interaction between reagents and cells. For example, orbital motion may be used to generate a homogeneous culture volume so that a small sample taken from this culture can represent the entire culture. In some variations, the agitator 6730 may include one or more impellers. The agitator 6730 may be configured to mix at varying intensities during a given culture cycle.

[0223]

[0328] In some variations, such as the toroidal bioreactor described herein, which includes a geometric shape capable of promoting a continuous and gentle fluid flow around the entire circumference, the orbital motion promotes increased interaction within the cell culture, thereby supporting homogeneous mixing while minimizing the shear stress transmitted to the cells.

[0224]

[0329] In some variations, the temperature controller 6740 may be configured to control the temperature of the bioreactor and the corresponding process. The temperature controller 6740 can be coupled to the bioreactor. For example, the temperature controller 6740 can ensure that the cell culture is heated to physiological conditions as desired and cooled to slow metabolic processes (e.g., to keep cells dormant) by controlling the temperature of the cell culture between approximately 2°C and approximately 40°C. For example, the temperature controller 6740 may include a circulating coolant coupled to a heat exchanger coupled to a thermal interface (e.g., a heating plate / cooling plate).

[0225]

[0330] In some variations, the gas regulator 6750 may be coupled to the bioreactor and configured to control the gas composition of the bioreactor and the corresponding process using one or more of clean dry air (CDA), carbon dioxide, and nitrogen. The gas regulator 6750 may be coupled to the bioreactor. For example, the sensor 6720 and the gas regulator 6750 may provide closed-loop gas control of the bioreactor module 6700. In some variations, the CDA may include oxygen, such as pure oxygen. In some variations, the gas regulator may include a manifold coupled to one or more gas sources. The manifold may include a solenoid coupled to a valve (e.g., a limiting orifice) configured to control the gas flow through the bioreactor 6710. The solenoid may be configured to pulse the amount and composition of the gas received through the manifold. Additionally or alternatively, one or more of a proportional valve and a mass flow controller (MFC) may be configured to measure and control the gas flow to the manifold. In some variations, the gas regulator 6750 may include one or more sensors for measuring the gas mixture and / or flow rate. Additionally or alternatively, these sensors may be configured for closed-loop control of the gas flow through the gas regulator.

[0226]

[0331] In some variations, the pH of the bioreactor 6710 can be controlled using the gas regulator 6750, based on the pH measured by the pH sensor. For example, the gas regulator 6750 can control the free hydrogen ions and pH of the cell culture by controlling the CO2 concentration of the gas in contact with the cell culture according to the measured pH. In some variations, the pH of the bioreactor 6710 should be approximately 5.5 to 8.5. To adjust the pH, one or more of the following can be used: the CO2 composition of the gas in the bioreactor 6710, the buffer solution, and reagents (e.g., acids, bases). In some variations, the dissolved oxygen concentration in the bioreactor 6710 can be approximately 0% to 21%. The dissolved oxygen concentration can be adjusted using the nitrogen composition of the gas in the bioreactor 6710. For example, the dissolved oxygen concentration can be adjusted by controlling both the agitator in the bioreactor and the flow rate and composition of the gas in contact with the cell culture.

[0227]

[0332] In some variations, the gas regulator 6750 can be used to control the oxygen concentration in the bioreactor 6710 (for example, to below atmospheric levels) using dissolved oxygen measured by a dissolved oxygen sensor. For example, the gas regulator 6750 can create a hypoxic state by controlling the nitrogen concentration of the gas in contact with the cell culture.

[0228]

[0333] Figures 68A and 68B are cross-sectional perspective views of a bioreactor 6800 comprising an enclosure 6810 including a bottom 6812, side walls 6814, and a top 6816. A gas-permeable membrane 6820 can be bonded to one or more of the bottom 6812 and side walls 6814 of the enclosure 6810. In some variations, the enclosure 6810 may include a first chamber 6830 having a first volume and a second chamber 6832 having a second volume. The first chamber 6830 is separated from the second chamber 6832, and the first volume is smaller than the second volume. In some variations, the first chamber 6830 may be concentrically nested within the second chamber 6832. For example, the nested arrangement of the chambers can increase the overall working volume range (e.g., 100:1). The first chamber 6830 may include a recessed shape with an angled bottom to facilitate fluid retention at the center of the first chamber 6830 during suction. In some variations, the bottom 6812 may be placed on a temperature controller (not shown), such as a thermoelectric element. In some variations, the enclosure 6810 may be made of a thermally conductive material such as metal (e.g., aluminum).

[0229]

[0334] In some variations, the bioreactor 6800 can be coupled to a gas regulator (not shown) to facilitate gas transfer (e.g., into and out of the culture) through a gas-permeable membrane 6820. The gas-permeable membrane 6820 may be configured to hold the cell culture. Gases can diffuse through the surface of the culture in contact with the gas-permeable membrane, enabling increased oxygenation of the cell culture and removal of gaseous metabolic byproducts, thereby increasing the potential for metabolic activity. For example, the gas-permeable membrane 6820 allows dissolved oxygen to diffuse into the culture in close proximity to a cell bed where oxygen can be consumed. In some variations, the bioreactor can be coupled to both a first gas regulator to facilitate gas transfer through the gas-permeable membrane and a second gas regulator to facilitate control of the headspace gas composition.

[0230]

[0335] In addition to gas transfer, the gas reactors described herein may be configured to efficiently control the temperature of the cell culture using conductive thermal interfaces (e.g., gas-permeable membrane 6820, enclosure 6810) along both the bottom and side walls of the bioreactor.

[0231]

[0336] In some variations, the first chamber 6830 may contain a working volume of approximately 10 ml to approximately 100 ml. In some variations, the first chamber 6830 may contain a total volume of approximately 10 ml to approximately 130 ml. In some variations, the second chamber 6832 may contain a working volume of approximately 100 ml to approximately 1000 ml. In some variations, the second chamber 6832 may contain a total volume of approximately 100 ml to approximately 1400 ml. In some variations, the first chamber 6830 may have a diameter of approximately 10 mm to approximately 100 mm and a height of approximately 10 mm to approximately 100 mm. In some variations, the second chamber 6832 may have a diameter of approximately 100 mm to approximately 250 mm and a height of approximately 10 mm to approximately 100 mm.

[0232]

[0337] As shown in Figure 68B, the bottom 6822 of the gas-permeable membrane 6820 may have an angle of approximately 3 to 10 degrees relative to the bottom 6812 of the enclosure 6810. Similarly, Figures 69A and 69B show inclined bases. For example, due to the inclination of the bottom 6822, the chambers 6830 and 6832 deepen towards the center of the bioreactor 6800. This promotes cell growth near the center of the bioreactor 6800 and can support one or more of the following: cell sampling, cell transfer, cell recovery, etc. In some variations, the orbital motion of the bioreactor 6800 can promote cell aggregation near the center of the bioreactor 6800, thereby increasing intercellular interactions.

[0233]

[0338] In some variations, the gas-permeable membrane 680 may include curved surfaces. In some variations, the gas-permeable membrane may include a set of patterned curved surfaces. For example, a set of patterned curved surfaces may have a radius of curvature ranging from approximately 50 mm to approximately 500 mm.

[0234]

[0339] In some variations, the bioreactor may be configured to facilitate monitoring between chambers (e.g., temperature, pH, dissolved oxygen) and fluid flow (e.g., gas composition, fluid transfer). As shown in Figure 68C, the enclosure 6810 may include one or more nesting surfaces surrounding the longitudinal axis (e.g., center) of the enclosure 6810. For example, the nesting surfaces may include a set of concentric annular bodies. The enclosure 6810 may have an annular shape. Figure 68C is a perspective view of the enclosure 6810 including a set of openings 6818 (e.g., holes, openings, slits, slots), and Figure 68D is a bottom view. In some variations, the openings 6818 allow for gas transfer and / or heat transfer between the components and chambers of the bioreactor 6800. Additionally or alternatively, one or more sensors may be coupled to the openings 6818. For example, the openings 6818 may be coupled to non-contact sensors (e.g., pH, DO), such as an optical sensor (not shown) configured to determine a fluorescent spot placed on the surface of the bioreactor. In some variations, one or more of the sensors and fluid connectors may be introduced through the opening 6818.

[0235]

[0340] In some variations, the gas permeable membrane extends along the bottom 6812 and side walls 6814 of the enclosure 6810, as shown in Figure 68B. In some variations, the gas permeable membrane extends only along the bottom 6812 of the enclosure 6810. Figure 68E is a perspective view of the gas permeable membrane 6820, whose outer surface includes one or more protrusions 6824 (e.g., protrusions, spacers, ribs, etc.), and Figure 68F is a side view. The protrusions 6824 are also illustrated in the perspective view of Figure 68G and the bottom view of Figure 68H. The protrusions 6824 contact the enclosure 6810 and define the cavity between the enclosure 6810 and the gas permeable membrane 6820. In other words, the projection 6824 may be configured to mechanically separate the enclosure 6810 from a portion of the gas-permeable membrane 6820 in order to facilitate heat transfer from the enclosure 6810 to the cell culture. In some variations, the gas-permeable membrane may include polydimethylsiloxane (PDMS) (e.g., silicone), fluoroethylene propylene (FEP), polyolefin (PO), polystyrene (PS), ethyl vinyl acetate (EVA), and its thickness may include about 0.1 mm to about 0.4 mm, about 0.2 mm to about 0.3 mm, about 0.25 mm, and all ranges and sub-values ​​in between.

[0236]

[0341] Figure 69A is a side cross-sectional view of the bioreactor enclosure 6910. The enclosure 6910 includes a first chamber 6912, a second chamber 6914, and a column 6916 extending along the longitudinal axis of the enclosure 6910. Figure 69B is a cross-sectional perspective view of the enclosure 6910 showing its nested curved surface. The column 6916 may be configured to promote cell culture in combination with agitation such as orbital motion.

[0237]

[0342] Figure 70 is an exploded perspective view of the bioreactor 7000, including the enclosure 7010, the gas permeable membrane 7020, and the upper part 7030. The upper part 7030 may be made of a material such as polyethylene.

[0238]

[0343] Figure 71A is a plan view of the bioreactor 7100, including the first chamber 7110 and the second chamber 7120. Figure 71B is a side cross-sectional view of the bioreactor 7100.

[0239]

[0344] Figures 13A and 13B are perspective views of the cartridge 1300 and the bioreactor instrument interface 1310. In Figure 13B, the bioreactor instrument interface 1310 is coupled to the cartridge 1300.

[0240]

[0345] Figure 14 is a perspective view of a bioreactor apparatus 1410 including a set of cartridges 1400, 1402, and 1404 and cavities 1420, 1422, and 1424 configured to receive each cartridge. In some variations, each cartridge can be docked to allow for simultaneous growth, culture, and stabilization steps.

[0241] Electroporation module

[0346] In some variations, the electroporation module may be configured to facilitate the intracellular transport of macromolecules (e.g., transfection by electroporation). The electroporation module may include a continuous-flow or batch-mode chamber and one or more electrode sets for applying a DC or AC current to the chamber. A discharge or current source from one or more capacitors can generate a current in the chamber sufficient to facilitate the transport of polynucleotides, proteins, nucleoprotein complexes, or other macromolecules into the cell. As with other modules described herein, one or more components used in the process step (here, electroporation) may be provided on a cartridge or in an instrument working with the cartridge. For example, one or more capacitors and / or batteries may be provided in the module or instrument on the cartridge. In some variations, as described, for example, in Garcia et al. Sci.Rep. 6:21238 (2016), the electroporation module may be configured to apply an electric field to a cell suspension under continuous flow in a microfluidic device.

[0242]

[0347] Additionally or alternatively, intracellular transport of macromolecules may be achieved by other methods, such as mechanoporation. Throughout this disclosure, it should be understood that variations including an electroporation module may, alternatively or additionally, include a mechanoporation module or other modules configured to perform any suitable method of delivering macromolecules into cells. Mechanoporation can be achieved, for example, by applying transient fluid pressure to a solution containing cells, or by applying physical pressure to cells (e.g., with microneedles). For example, international patent applications WO2017 / 041051 and WO2017 / 123663 present exemplary methods of mechanoporation by passing a cell suspension through a constriction, and these applications are incorporated herein by reference. Mechanoporation can also be achieved by creating vortices in a cell suspension in a microfluidic device.

[0243]

[0348] Figure 72 is a schematic diagram of an electroporation module 7200 (e.g., an electroporation system). The electroporation module 7200 includes an electroporation chamber 7210 (which may include fluid tubes), a pump 7220, an inlet 7230, an outlet 7232, a pinch valve set 7234, a first fluid source 7240 (e.g., a fluid storage section, a cell storage section), a second fluid source 7242 (e.g., a vent, a gas source), a sensor set 7250 (e.g., a bubble sensor), a controller (e.g., a processor and memory) configured to control the module 7200, and a signal generator 7270 configured to send an electroporation signal (e.g., a voltage pulse) to the electroporation chamber 7210.

[0244]

[0349] In some variations, the fluid tube 7210 may be configured to receive a first fluid containing cells and a second fluid. An electrode set can be connected to the fluid tube 7210. A pump can be connected to the fluid tube 7210. The controller 7260 may be configured to generate a first signal to introduce the first fluid into the fluid tube 7210 using the pump 7220, a second signal to introduce the second fluid into the fluid tube 7210 so that the first fluid is separated from the third fluid by the second fluid, and an electroporation signal to perform electroporation of cells in the fluid tube 7210 using the electrode set.

[0245]

[0350] In some variations, the second fluid may include gas or oil. In some variations, the controller may be configured to generate a third signal to introduce a third fluid into the fluid tube 7210. The third fluid may be separated from the first fluid by the second fluid. In some variations, the cartridge for cell processing may include a liquid transfer bus and several modules, such as an electroporation module 7200. Each module may be fluidically linked to the liquid transfer bus.

[0246]

[0351] The sensor set 7250 may be configured to measure fluid changes within a fluid tube, such as a change from a first fluid to a second fluid (e.g., from liquid to air) within the fluid tube. Module 7200 may further include a valve set configured to ensure that the fluid does not flow back into the electroporation module 7210 and / or the fluid source 7240. The electroporation chamber 7210 may include a cavity configured to hold the fluid on which electroporation is to be performed, and a set of electrodes for applying an electroporation signal to the fluid. For example, the signal generator 7270 may generate a square valve pulse as detailed herein.

[0247]

[0352] In some variations, the electroporation module 7200 (e.g., valve 7234, pump 7220, sensor 7250, and controller 7260) may be configured to control the fluid flow through the electroporation chamber 7210 discontinuously (e.g., in a batch process). For example, a first cell batch may be electroporated and physically separated from a second cell batch by an intermediate fluid such as air or a fluid such as oil. Cell batch separation can reduce mixing between transfected and untransfected cells and further ensure a fixed batch volume. That is, a fluid gap can form a visually verifiable boundary between cell batches to reduce diffusion and mixing between electroporated and unelectroporated cells. Cell batch separation can reduce the time cells are exposed to certain cytotoxic reagents (e.g., electroporation buffers), thereby improving performance.

[0248]

[0353] In some variations, cell batch electroporation can be performed in a substantially static state (e.g., a state where there is substantially no fluid flow). In contrast, conventional continuous-flow electroporation has a fluid flow rate limit that correlates with transfection efficiency. The batch processing described herein allows for the transfer of cell batches into and out of the electroporation chamber 7210 at a predetermined rate, thereby increasing the overall throughput of the system 7200 without reducing electroporation efficiency. Furthermore, the electroporation system 7200 does not use the precisely controlled flow rate / pulse rate required in continuous-flow electroporation systems.

[0249]

[0354] Figure 73 is an exploded perspective view of an electroporation module 7300, which may include electrodes 7310, fluid tubes 7320 (e.g., electroporation chambers), substrates 7330 (e.g., alloy busbars), housings 7340, and fasteners 7350. In some variations, the fluid tubes 7320 may be configured to hold a fluid volume of approximately 0.4 ml to approximately 3.5 ml. The electroporation module 7300 has a parallel plate design. In some variations, the electrodes may include stainless steel and be separated by insulating gaskets. In some variations, polishing the electrodes and / or coating them with a non-reactive material (e.g., gold, platinum) can reduce the gradual accumulation of biological material (e.g., charged molecules, DNA, proteins) on the electrode surface.

[0250]

[0355] Generally, a method for performing electroporation of cells may include receiving a first fluid containing cells into a fluid tube, receiving a second fluid into the fluid tube to separate the first fluid from a third fluid, and performing electroporation of cells by applying an electroporation signal to the first fluid. In some variations, the third fluid may be separated from the first fluid by the second fluid within the fluid tube. In some variations, the first fluid may be substantially static when the electroporation signal is applied.

[0251]

[0356] Figures 74A and 74B are schematic diagrams of variations of the electroporation processes 7400 and 7402. Method 7400 may include introducing cells into the electroporation chamber 7450 (7410). For example, in step 7412, a first fluid can be injected into the electroporation chamber by opening valve v1 and generating negative pressure with a pump (valves v2 and v3 are closed). In step 7414, a second fluid (e.g., gas, oil) can separate the first fluid from the third fluid to produce a first cell batch for electroporation. For example, valve v1 and v3 can be closed while valve v2 is open and the pump is generating negative pressure. In some variations, the injection volume may be approximately 1 ml to 3 ml, and the injection time may be approximately 8 seconds to 15 seconds (at a rate of approximately 20 ml / min). In step 7420, the electroporation of cells in the first fluid can be performed with each valve closed and the pump turned off. In step 7430, valves v1 and v2 are closed, valve v3 is opened, and with the pump generating positive pressure, the cells of the first fluid can be flowed out of the electroporation chamber 7450.

[0252]

[0357] Figure 74B shows another configuration in which a pump placed between the input and the electroporation chamber may be configured to inject in one direction. Method 7402 may include introducing cells into the electroporation chamber 7450 (7411). For example, in step 7416, a first fluid can be injected into the electroporation chamber by opening valves v1 and v4 and the pump generating positive pressure (valves v2 and v3 are closed). In step 7418, a second fluid (e.g., gas, oil) can separate the first fluid from the third fluid to produce a first batch of cells to be electroporated. For example, valves v1 and v3 can be closed while valves v2 and v4 are open and the pump is generating positive pressure. In step 7422, the electroporation of cells in the first fluid can be performed with each valve closed and the pump turned off. In step 7432, valves v1 and v4 are closed, valves v2 and v3 are opened, and with the pump generating positive pressure, the cells of the first fluid can be flowed out of the electroporation chamber 7450 and output.

[0253]

[0358] In some variations, the impedance / resistance between electrodes in an electroporation system can increase over time due to electrode passivation / degradation caused by the adhesion of charged biological materials (e.g., charged molecules, DNA, proteins) to the electrode surface. Active electric field compensation can be used to ensure that a consistent electric field strength is applied to cells across multiple cell batches. This reduces the need to modify the electrode surface to reduce passivation.

[0254]

[0359] Figure 75 is a circuit diagram of a resistor divider network for the electroporation process 7500. For example, voltage V chip A cell set can be placed in an electroporation chamber 7510 to which a fluid resistance R can be applied. bThis corresponds to the fluid resistance (e.g., cell mixture). Assuming uniform cell distribution and that electroporation is performed on each fluid batch of the same volume, the fluid resistance R b It must be consistent. i V corresponds to the resistance between the fluid and the electrode, which increases over time throughout the electroporation process. In conventional electroporation processes, V ps R is constant. However, R i As this increases over time, the voltage applied to the fluid decreases over time, leading to a decrease in electric field strength.

[0255]

[0360] Fluid resistance R b Due to fluctuations and the limited number of pulses that can be applied, interpolation to compensate for the decrease in electric field strength may not accurately compensate for electrode passivation.

[0256]

[0361] In some variations, a method for performing electroporation of cells may include receiving a first fluid containing cells into a fluid tube, applying a resistance measurement signal to the first fluid using an electrode set, measuring the resistance between the first fluid and the electrode set, and applying an electroporation signal to the first fluid based on the measured resistance. In some variations, a second fluid containing gas can be received into the fluid tube before applying the electroporation signal to the fluid. The first fluid can be separated from the third fluid by the second fluid.

[0257]

[0362] Figures 76A to 76D are plots of measurement waveforms and electroporation waveforms 7600, 7602, 7604, and 7606. Figure 76A shows a first resistance measurement pulse 7620 with low voltage and wide pulse width. Figure 76B shows a second resistance measurement pulse 7622 with high voltage and narrow pulse width. Figure 76C shows a third resistance measurement pulse 7624 with a continuous low voltage waveform for continuously monitoring impedance changes over time. Figure 76D shows a fourth resistance measurement pulse 7626 with a low AC voltage waveform for continuously monitoring impedance changes over time. Each resistance measurement pulse avoids inducing electroporation in cells by reducing voltage and / or pulse width. By monitoring the voltage and current of the applied resistance measurement pulse, changes in resistance can be measured and the electroporation pulse applied to the cell batch can be compensated accordingly.

[0258]

[0363] In some variations, the electroporation signal may consist of approximately 1 to 50 pulses, with a voltage of approximately 100V to 700V, a pulse width of approximately 100μs to 1ms, a pulse interval of approximately 5 seconds to 30 seconds, a resistive pulse voltage of approximately 10V to 40V, and a resistive pulse width of approximately 10μs to 50μs.

[0259]

[0364] For example, performing 8-batch electroporation allows for the reception of one electroporation pulse per batch. Each electroporation pulse may have an electric field strength of approximately 0.5 kV / cm to approximately 2.0 kV / cm. The resistance measurement pulse applied before each batch may have an electric field strength of less than approximately 0.2 kV / cm so as not to induce electroporation with this resistance measurement pulse.

[0260] Sterilization fluid transfer device

[0365] Generally, the sterile fluid transfer devices described herein may be configured to store fluids for transfer to other components of a cell processing system, such as cartridges or bioreactors. In some variations, the sterile fluid transfer device may include a portable consumable configured to be moved using a robot. For example, a robot may be configured to move the sterile fluid transfer device from a reagent storage container to an ISO7 space for the sterile fluid transfer device within the cell processing system. The sterile fluid transfer device enables automated, sterile, and quantitative fluid transfer to automate the production of cell therapies.

[0261]

[0366] Figures 103A and 103B are perspective views of a sterile fluid transfer device 10300, including a fluid cavity 10310 (e.g., a container, vessel), a fluid connector 10320 (e.g., a fluid connector), and a pump 10330. Fluid stored in the fluid cavity 10310 can be transferred into and out of the sterile fluid transfer device 10300 via the fluid connector 10320 using the pump 10330. In some variations, the sterile fluid transfer device 10300 may include an engagement mechanism 10340 (e.g., a robot mount) to facilitate robot arm control.

[0262] Fluid connector

[0367] In general, the sterile fluid connectors described herein enable fluid transfer, which may be at least one of sterile fluid transfer, fully automated fluid transfer, and precisely metered fluid transfer (e.g., precise control of the volume of fluid being transferred), by forming a sterile fluid path between at least two fluid devices. In some variations, a robot may be configured to connect the fluid connector between at least two of a plurality of instruments and one or more cartridges. In some variations, a robot may be configured to operate a fluid controller to open and close a set of ports and valves on the fluid connector. Operating the fluid connector with a robot and controller can facilitate the automation and sterility of cell processing systems.

[0263]

[0368] In some variations, the system may include a robot configured to operate a fluid connector as described herein, and a controller including memory and a processor. The controller may be coupled to the robot. The controller may be configured to generate a port signal using a robot arm to connect a first port to a second port, a first valve signal using a robot arm to move a first valve in parallel with respect to a second valve, and a second valve signal to move the first and second valves to an open configuration.

[0264]

[0369] In some variations, a fluid pump can be coupled to the sterilizer source, and the controller may be configured to generate a first fluid signal to circulate the fluid within the chamber via the sterilizer port. The controller may be configured to generate a second fluid signal to circulate the sterilizer within the chamber via the sterilizer port, at least to sterilize the chamber. The controller may be configured to generate a third fluid signal to remove the sterilizer from the chamber.

[0265]

[0370] In some variations, the controller may be configured to generate a port signal for coupling a first port to a second port using a robotic arm, a first valve signal for translating a first valve relative to a second valve using a robotic arm, and a second valve signal for moving both the first and second valves to an open configuration.

[0266]

[0371] The fluid connector can further accommodate multiple connection cycles in a sterile system and can be controlled without human intervention. For example, the fluid connector may include one or more of the following: an engagement mechanism to facilitate robot arm control, and an alignment mechanism to ensure proper connection between connector components. Figure 15 is a block diagram of an exemplary variation of a fluid connector system 1500, including a fluid connector 1510, a first fluid device 1520, a second fluid device 1522, a sterilizer source 1530, a fluid source 1532, a robot (e.g., a robot arm) 1540, and a controller 1550. The fluid connector 1510 can be detachably coupled (e.g., connected and / or disconnected, attached and detached) to each of the first fluid device 1520, the second fluid device 1522, the sterilizer source 1532, the fluid source 1532, and the robot 1540. In some variations, the fluid device may include one or more of the following: a cartridge and a sterilizer transfer device. For example, the sterilizer transfer device may be in fluid communication with the cartridge via the fluid connector. As detailed herein, separate parts of the fluid connector 1510 (e.g., male connector, female connector) can be detachably coupled to one another. The robot 1540 may be configured to physically manipulate (e.g., detachably couple) one or more of the fluid connector 1510, the first fluid device 1520, the second fluid device 1522, the sterilizer source 1530, and the fluid source 1532 in a predetermined manner. For example, the robot 1540 can connect the fluid connector 1510 between the first fluid device 1520 and the second fluid device 1522. The robot 1540 can also connect the sterilizer source 1530 and / or the fluid source 1532 to the sterilizer port of the fluid connector 1510. In some variations, the robot 1540 can initiate a sterilization process of one or more parts of the fluid connector 1510 using, for example, a sterilizer from the sterilizer source 1530 by controlling one or more valves and / or ports of the fluid connector 1510. The controller 1550 can be coupled to one or more of the robot 1540, the sterilizer source 1530, and the fluid source 1532 to control one or more of the fluid transfer and sterilization processes.

[0267]

[0372] Figure 16A is a schematic diagram of an exemplary variation of the fluid connector 1600. The fluid connector 1600 may include a lumen extending along its length. This lumen is positioned between the first fluid device 1630 and the second fluid device 1640 to allow fluid flow through the fluid connector 1600. In some variations, the first fluid device 1630 and the second fluid device 1640 can be connected and disconnected aseptically using the fluid connector 1600. The fluid devices 1630 and 1640 may include closed sterile devices, which may be of the same type or different types. For example, the fluid devices 1630 and 1640 may include one or more sterile fluid transfer devices and consumables. In some variations, the fluid connector 1600 may include a first connector 1610 including a first proximal end 1612 and a first distal end 1614. The first proximal end 1612 may be configured to connect to the first fluid device 1630. The first distal end 1614 may include the first port 1616, the first housing 1617, and the first valve 1618. The first housing 1617 may be configured to receive the first port 1616 in a closed configuration, as detailed herein.

[0268]

[0373] The fluid connector 1600 may further include a second connector 1620, which includes a second proximal end 1622 and a second distal end 1624. The second proximal end 1622 may be configured to connect to a second fluid device 1640. The second distal end 1624 may include a second port 1626, a second housing 1627, and a second valve 1628. The second housing 1627 may be configured to receive the second port 1626 in a closed configuration. In Figure 16A, the first connector 1610 includes a sterilizer port 1650 configured to connect to a sterilizer source (not shown). Additionally or alternatively, the second connector 1620 may also include a sterilizer port 1650. As detailed herein, when the second port 1626 is coupled to the first port 1616, the sterilizer port 1650 may be configured to have fluid communication with the first distal end 1614 and the second distal end 1624.

[0269]

[0374] In some variations, the fluid devices 1630, 1640 may include a sterilizer chamber and a sterilizer port configured to receive a sterilizer. The sterilizer chamber may house a fluid device connector (not shown) configured to connect to the proximal end of the first connector 1610 or the second connector 1620. The fluid devices 1630, 1640 may receive a sterilizer in the same manner as the fluid connector 1600.

[0270]

[0375] Figure 16B is a detailed schematic diagram of the first connector 1610, including the first port housing 1617 and the chamber 1615. The chamber 1615 may be defined by a cavity surrounded by one or more distal ends 1614, 1624. For example, the chamber 1615 in Figure 16B may include the portion of the first connector 1610 (e.g., the first distal end 1614) between the first valve 1618 and the first port 1616 in a closed configuration. In some variations, the first chamber 1615 is approximately 1 cm 3 ~about 5cm 3 The chamber may have a volume of the above. When the first connector 1610 is coupled to the second connector 1620 and the ports 1616 and 1626 are in an open configuration (for example, shown in Figure 16D), the chamber 1616 may include the portion of the fluid connector 1600 between the first valve 1618 and the second valve 1628 (for example, the first distal end 1614 and the second distal end 1624). The chamber 1615 may include a closed volume configured to receive a fluid such as a sterilizer from the sterilizer port 1650. In some variations, the sterilizer port 1650 may include an inlet 1652 and an outlet 1654. Methods using the fluid connector will be described in more detail in relation to Figures 16C to 16L and Figure 27.

[0271]

[0376] In some variations, the fluid connector 1600 may include one or more alignment and robot engagement mechanisms configured to facilitate robot operation, as detailed herein. In some variations, the fluid connector 1600 may be connected to one or more sensors, pumps, and valves to facilitate fluid transfer and monitoring.

[0272]

[0377] In some variations, the components of the fluid connector in contact with the fluid may be USP Class VI compliant for cell processing and / or GMP applications. In some variations, the components of the fluid connector may consist of materials including, but are not limited to, cyclic olefin copolymers (COC), polychlorotrifluoroethylene, polyetherimide, polysulfone, polystyrene, polycarbonate, polypropylene, silicone, polyetheretherketone, polymethyl methacrylate, nylon, acrylic, polyvinyl chloride, vinyl, phenolic resin, petroleum polymers, glass, polyethylene, terephthalate, metals, stainless steel, titanium, aluminum, cobalt-chromium, chromium, silicates, glass, alloys, ceramics, carbohydrate polymers, quasi-minerals, and combinations or composites thereof.

[0273]

[0378] Figures 17A to 18D show the external and internal deformations of a fluid connector. Figure 17A is a front perspective view of a fluid connector 1700 in a closed port configuration. Figure 17B is a rear perspective view of the fluid connector 1700, and Figure 17C is a rear view. Generally, a fluid connector may include multiple internal seals to reduce contamination and assist in sterilization, and an alignment mechanism to assist in proper alignment of the fluid connector components.

[0274]

[0379] The fluid connector 1700 may include a lumen extending along its length. In some variations, the fluid connector 1700 may comprise a first connector 1710 including a first proximal end 1712 and a first distal end 1714. The first proximal end 1712 may be configured to connect to a first fluid device (not shown for clarity). The first proximal end 1712 may include a Luer connector or any other suitable connector. The first distal end 1714 may include a first port 1716 and a first housing 1717. In Figure 17A, the first housing 1717 is shown keeping the first port 1716 in a closed configuration. The first connector 1710 further includes sterilizer ports 1750, 1752 configured to connect to a sterilizer source (not shown for clarity). In some variations, the sterilizer ports may include an inlet and an outlet. In some variations, the sterilizer port may optionally include one or more of a check valve and a particle filter configured to reduce contamination within the sterilizer port when not connected to a robot or actuator. The first connector 1710 may include a first alignment mechanism 1760, such as a set of protrusions on the first distal end 1714 of the first connector 1710. The alignment mechanism can ensure that the operation of the fluid connector is not affected by small positioning errors caused by robot operation.

[0275]

[0380] The fluid connector 1700 may further include a second connector 1720, which includes a second proximal end 1722 and a second distal end 1724. The second proximal end 1722 may be configured to couple to a second fluid device (not shown for clarity). The second proximal end 1722 may include a Luer connector or any other suitable connector. The second distal end 1724 may include a second port 1726 and a second housing 1727. In Figure 17A, the second housing 1727 is shown keeping the second port 1726 in a closed configuration. The second connector 1720 may further include a second alignment mechanism 1762, such as a set of holes on the second distal end 1724 of the second connector 1720. The second alignment mechanism 1762 may be coupled to the first alignment mechanism 1760 in a predetermined axial rotation configuration to assist in the engagement of the first connector 1710 and the second connector 1720.

[0276]

[0381] The first port 1716 and the second port 1726, respectively, are housed in the first housing 1717 of the first distal end 1714 and the second housing 1727 of the second distal end 1724, respectively. Since ports 1716 and 1726 are not detachable from the fluid connector 1700, robot control is facilitated, and thus the risk of failure of automated operation by the robot is reduced.

[0277]

[0382] In some variations, the first connector 1710 may include a first robot engagement mechanism 1770, and the second connector 1720 may include a second robot engagement mechanism 1772. The robot engagement mechanisms 1770 and 1772 may be configured to be operated by a robot such as a robot arm (e.g., robot 1540). In some variations, as shown in Figures 17A to 17F, the robot engagement mechanisms 1770 and 1772 may be operably coupled to a first port 1716 and a second port 1726, respectively, and configured to actuate ports 1716 and 1726 into closed port configurations and open port configurations. Additionally or alternatively, a user may manually actuate the robot engagement mechanisms 1770 and 1772 to actuate ports 1716 and 1726, respectively.

[0278]

[0383] Figure 17D is a front perspective view of the fluid connector 1700 in an open port configuration. Figure 17E is a rear perspective view of the fluid connector 1700 in an open port configuration, and Figure 17F is a rear view. In the open port configuration, the first valve 1718 of the first connector 1710 and the second valve 1728 of the second connector 1720 are shown in Figure 17D.

[0279]

[0384] Figure 18A is a side view of the fluid connector 1800 in a separate configuration, and Figure 18B is a side cross-sectional view. In some variations, the fluid connector 1800 may comprise a first connector 1810 comprising a first housing 1817 including a first port 1816, a sterilizer port 1850 configured to couple to a sterilizer source (not shown), and a first alignment mechanism 1860 configured to couple to a corresponding alignment mechanism (not shown) of a second connector 1820. The fluid connector 1800 may comprise a second connector 1820 comprising a second housing 1827 including a second port 1826. The first connector 1810 and the second connector 1820 can be aligned axially, and the alignment mechanism can assist in the rotational alignment of the first connector 1810 relative to the second connector 1820. The first valve 1818 may include a first valve shaft 1819, and the second valve 1828 may include a second valve shaft 1829.

[0280]

[0385] Figure 18C is a side view of a fluid connector 1800 in which the first housing 1817 and the second housing 527 are combined into one, but the first port 1816 and the second port 1826 are both in a closed configuration, so the first connector 1810 and the second connector 1820 are not in fluid communication. Figure 18D is a side cross-sectional view. The first alignment mechanism on each connector 1810, 1820 may be configured to ensure axial and / or rotational alignment between the first connector 1810 and the second connector 1820.

[0281]

[0386] Figure 18E is a side view of the fluid connector 1800 in an open port configuration, and Figure 18F is a side cross-sectional view. The first port 1817 and the second port 1827 are each transitioned from a closed configuration to an open configuration. This creates a closed internal volume within the distal ends of each connector 1810 and 1820. Since the first valve 1818 and the second valve 1828 are each in a closed configuration, the fluid flow between the first connector 1810 and the second connector 1820 is blocked and is suppressed to half portions by the automatic shut-off valves on both sides.

[0282]

[0387] Figure 18G is a side view of a fluid connector 1800 in an open valve configuration in which a first valve 1818 is coupled to a second valve 1828, and Figure 18H is a side cross-sectional view. For example, the second valve 1828 can be moved parallel to the first valve 1818 along the longitudinal axis of the second connector 1820. As illustrated in Figures 18G and 18H, the second connector 1820 can be compressed axially to move the second valve 1828 parallel to the first valve 1818. The first valve 1818 coupled to the second valve 1828 forms a radial seal, and the first valve shaft 1819 and the second valve shaft 1829 are in contact, enabling fluid communication between the first connector 1810 and the second connector 1820.

[0283]

[0388] Figures 19 to 26B are schematic diagrams of variations of a fluid connector system for coupling fluid devices. In some variations, the fluid connector may include a first connector configured to couple to any one of a plurality of second connectors. Figure 19 is a schematic diagram of an exemplary variation of a fluid connector system 1900, which includes a first connector 1910, a plurality of second connectors 1920, 1921, 1922, a first fluid device 1930 (e.g., a sterile fluid transfer device), a second fluid device 1940 (e.g., consumables), and a robot 1960 (e.g., a robotic arm, a 3DOF robot). The first connector 1910 may be coupled in fluid communication with the first fluid device 1930, and the second connectors 1920, 1921, 1922 may be coupled in fluid communication with the second fluid device 1940. The first connector 1910 and the second connectors 1920, 1921, 1922 may each include a port 1916 configured to connect to a corresponding port, as detailed herein. The robot 1960 may include one or more end effectors 1962, 1964 configured to operate and / or connect to one or more of the first fluid device 1930 and the first connector 1910. For example, the first connector 1910 may include one or more sterile ports 1950 configured to connect to an end effector 1962 (e.g., a gripper). Similarly, the first fluid device 1930 may include one or more fluid ports 1952 configured to connect to an end effector 1964.

[0284]

[0389] In some variations, to facilitate efficient shared fluid connections between fluid devices, fluid connectors, and sterilization systems, the robot 1960 may be configured to connect to one or more of a sterile agent source, a fluid source, and a pump. For example, Figure 96A is a plan view of a fluid device 9600 (e.g., a sterile fluid transfer device) including a fluid port 9610 configured to connect to a fluid source (not shown) and a sterilization port 9620 configured to connect to a sterile agent source (not shown). Figures 96B and 96C are a side view and a perspective view of the fluid device 9600 connected to the robot 9650, respectively. In some variations, the robot 9650 may include one or more fluid tubes 9660 configured to connect to one or more of the fluid port 9610 and sterilization port 9620 of the fluid device 9600.

[0285]

[0390] In some variations, the fluid connector may include a third connector positioned between the first and second connectors. Figure 20A is a schematic diagram of an exemplary variation of a fluid connector system 2000, which includes a first connector 2010, a plurality of second connectors 2020, 2021, 2022, a third connector 2070 (e.g., instruments, sterilization enclosures), a first fluid device 2030 (e.g., sterilization fluid transfer device), a second fluid device 2040 (e.g., consumables), and a robot 2060 (e.g., robotic arm, 3DOF robot, 1DOF robot). The first connector 2010 may be coupled in fluid communication with the first fluid device 2030, and the second connectors 2020, 2021, 2022 may be coupled in fluid communication with the second fluid device 2040. The third connector 2070 may be coupled between the first connector 2010 and one of the second connectors 2020, 2021, and 2022. The third connector 2070 may include a lumen configured to receive and circulate a sterilizer through one or more portions of the first connector 2010, the second connectors 2020, 2021, and 2022, and the third connector 2070. In some variations, the first fluid device 2030 and one or more of the first connectors 2010 can be simplified by non-removably coupling the sterilization port 2052 to the sterilizer source and / or fluid source.

[0286]

[0391] The robot 2060 may include one or more end effectors 2062, 2064, 2066 configured to operate and / or couple to one or more of the first fluid device 2030, the first connector 2010, and the third connector 2070. For example, the first fluid device 2030 may include one or more fluid ports 2050 configured to couple to end effector 2062. Similarly, the third connector 2070 may include one or more sterilization ports 2052 configured to couple to the robot 2060 (e.g., end effector 2064). In some variations, to facilitate efficient shared fluid connections between the fluid device, fluid connectors, and sterilization systems, the robot 2060 may be configured to couple to one or more of the sterile agent source, fluid source, and pump.

[0287]

[0392] Figures 20B and 20C are schematic diagrams of the fluid connector connection process. In Figure 20B, at 2002, the third connector 2070 can be connected to the distal end of the first connector 2010. At 2004, the distal end of the second connector 2020 can be connected to the third connector 2070. At 2006, the second connector 2020 can be translated through the third connector 2070 to directly connect the second connector 2020 to the first connector 2010.

[0288]

[0393] In Figure 20C, in 2002, the third connector 2070 can be coupled to the distal end of the first connector 2010 and the distal end of the second connector 2020. In 2005, the first connector 2010 and the second connector 2020 can each be moved parallel to each other through the third connector 2070. In 2007, the second connector 2020 can be further moved parallel to the first connector 2010, so that the first connector 2010 can be directly coupled to the second connector 2010. Figure 20C further shows that the first port 2090 and the second port 2092 can transition between a closed port configuration and an open port configuration.

[0289]

[0394] In some variations, the fluid connector may include a third connector positioned between the first and second connectors. The third connector may be coupled to a second robot different from the first robot coupled to the first connector. Figure 21 is a block diagram of an exemplary variation of a fluid connector system 2100, including a first connector 2110, a plurality of second connectors 2120, 2121, 2122, a third connector 2170 (e.g., instruments, sterilization enclosures), a first fluid device 2130 (e.g., sterilization fluid transfer device), a second fluid device 2140 (e.g., consumables), a first robot 2160, and a second robot 2166. The first connector 2110 may be coupled in fluid communication with the first fluid device 2130, and the second connectors 2120, 2121, 2122 may be coupled in fluid communication with the second fluid device 2140. The third connector 2170 may be coupled between the first connector 2110 and one of the second connectors 2120, 2121, and 2122. The third connector 2170 may include a lumen configured to receive and circulate a sterilizer through one or more portions of the first connector 2110, the second connectors 2120, 2121, and 2122, and the third connector 2170. In some variations, the first fluid device 2130 and one or more of the first connectors 2110 can be simplified by non-removably coupling the third connector 2170 to the sterilizer source and / or fluid source.

[0290]

[0395] The first robot 2160 may include one or more end effectors 2162, 2164 configured to operate and / or couple to one or more of the first fluid device 2130 and the first connector 2110. For example, the first fluid device 2130 may include one or more fluid ports 2150 configured to couple to the end effector 2162. The third connector 2170 may couple to the second robot 2166 (e.g., a 3DOF robot). In some variations, to facilitate efficient shared fluid connections between the fluid device, fluid connector, and sterilization system, the robots 2160, 2166 may be configured to couple to one or more of the sterile agent source, fluid source, and pump.

[0291]

[0396] In some variations, the fluid connector may include a sterilizer source coupled to a plurality of second connectors. Figure 22 is a block diagram of an exemplary variation of a fluid connector system 2200, which includes a first connector 2210, a plurality of second connectors 2220, 2221, 2222, a first fluid device 2230 (e.g., a sterilizer transfer device), a second fluid device 2240 (e.g., consumables), a robot 2260, a sterilizer source 2290 including one or more valves, and a sterilizer switch 2292. The first connector 2210 may be coupled in fluid communication with the first fluid device 2230, and the second connectors 2220, 2221, 2222 may be coupled in fluid communication with the second fluid device 2240. The robot 2260 may include one or more end effectors 2262, 2264 configured to operate and / or connect to one or more of the first fluid device 2230 and the first connector 2210. For example, the first fluid device 2230 may include one or more fluid ports 2250 configured to connect to the end effector 2262. In some variations, the sterilizer source 2290 may be connected to the switch 2292. To facilitate efficient shared fluid connection between the fluid device, fluid connector and sterilization system, the switch 2292 may be connected to each of the second connectors 2220, 2221, and 2222. In some variations, a sterilizer conduit can be routed from the switch 2292 through the second fluid device 2240 to each of the second connectors 2220, 2221, and 2222.

[0292]

[0397] In some variations, the fluid device may include one or more sterilizer valves coupled to a plurality of second connectors. Figure 23 is a block diagram of an exemplary variation of the fluid connector system. Figure 23 is a block diagram of an exemplary variation of the fluid connector system 2300, which includes a first connector 2310, a plurality of second connectors 2320, 2321, 2322, a first fluid device 2330 (e.g., a sterilizer transfer device), a second fluid device 2340 (e.g., consumables), a robot 2360, a sterilizer valve set 2390 disposed within the housing of the second fluid device 2340, and a sterilizer switch 2392. The first connector 2310 may be coupled in fluid communication with the first fluid device 2330, and the second connectors 2320, 2321, 2322 may be coupled in fluid communication with the second fluid device 2340. The robot 2360 may include one or more end effectors 2362, 2364 configured to operate and / or couple to one or more of the first fluid device 2330 and the first connector 2310. For example, the first fluid device 2330 may include one or more fluid ports 2350 configured to couple to the end effector 2362. In some variations, the sterilizer valve 2390 may be coupled to the switch 2392. To facilitate efficient shared fluid connection between the fluid device, fluid connector, and sterilization system, the switch 2392 may be coupled to each of the second connectors 2320, 2321, and 2322 via the sterilizer valve 2390. In some variations, a sterilizer conduit can be routed from the switch 2392 to each of the second connectors 2320, 2321, and 2322 via the second fluid device 2340.

[0293]

[0398] In some variations, the fluid connector may include a sterilizer source coupled to a plurality of second connectors, each of which may have a sterilizer port (e.g., a sterilizer valve) and a sterilizer conduit through a fluid device. Figure 24A is a block diagram of an exemplary variation of a fluid connector system 2400, including a first connector 2410, a plurality of second connectors 2420, 2421, 2422, a first fluid device 2430 (e.g., a sterilizer transfer device), a second fluid device 2440 (e.g., consumables), a robot 2460, and a sterilizer switch 2492 coupled to a sterilizer source (not shown). The first connector 2410 may be coupled in fluid communication with the first fluid device 2430, and the second connectors 2420, 2421, 2422 may be coupled in fluid communication with the second fluid device 2440. The robot 2460 may include one or more end effectors 2462, 2464 configured to operate and / or connect to one or more of the first fluid device 2430 and the first connector 2410. For example, the first fluid device 2430 may include one or more fluid ports 2450 configured to connect to the end effector 2462.

[0294]

[0399] In some variations, each of the second connectors 2420, 2421, and 2422 may include sterilizer ports 2494, 2496, and 2498, each containing a valve coupled to the distal end of the second connectors 2420, 2421, and 2422. In some variations, a sterilizer conduit can be routed from switch 2492 through a second fluid device 2440 to each of the sterilizer ports 2494, 2496, and 2498. In some variations, a sterilizer source (not shown) may be coupled to switch 2492. To facilitate efficient shared fluid connection between the fluid device, fluid connectors, and sterilization system, switch 2492 may be coupled to each of the second connectors 2420, 2421, and 2422 via the sterilizer ports 2494, 2496, and 2498.

[0295]

[0400] Figure 24B is a schematic diagram of the fluid connector connection processes 2402, 2404, and 2406 when the first connector 2410 is coupled to the second connector 2420. For example, when the first connector 2410 and the second connector 2420 are separated and disconnected (2402), the sterilizer port 2494 is in a closed valve configuration. Figure 24C is a detailed schematic diagram of the sterilizer valve 2494. In some variations, in 2404 and 2406, when the first connector 2410 is coupled to the second connector 2420, the valve 2494 may transition to an open valve configuration.

[0296]

[0401] In some variations, the multiple second connectors may include one or more pneumatic sterilizer valves and sterilizer pathways through fluid devices. Figure 25A is a block diagram of an exemplary variation of a fluid connector system 2500, which includes a first connector 2510, a plurality of second connectors 2520, 2521, 2522, a first fluid device 2530 (e.g., a sterilizer transfer device), a second fluid device 2540 (e.g., consumables), a robot 2560, and a sterilizer switch 2592 coupled to a sterilizer source (not shown). The first connector 2510 may be coupled in fluid communication with the first fluid device 2530, and the second connectors 2520, 2521, 2522 may be coupled in fluid communication with the second fluid device 2540.

[0297]

[0402] In some variations, each of the second connectors 2520, 2521, and 2522 may include pneumatic sterilizer ports 2594, 2596, and 2598, each containing a valve coupled to the distal end of the second connectors 2520, 2521, and 2522. In some variations, sterilizer conduits can be routed from switch 2592 through a second fluid device 2540 to each of the sterilizer ports 2594, 2596, and 2598. In some variations, a sterilizer source (not shown) may be coupled to switch 2592. To facilitate efficient shared fluid connection between the fluid device, fluid connectors, and sterilization system, switch 2592 may be coupled to each of the second connectors 2520, 2521, and 2522 via the sterilizer ports 2594, 2596, and 2598.

[0298]

[0403] The robot 2560 may include a first fluid device 2530, a first connector 2510, and one or more end effectors 2562, 2564 configured to operate and / or couple to one or more of the sterilizer ports 2594, 2596, 2598. For example, the first fluid device 2530 may include one or more fluid ports 2550 configured to couple to end effector 2562. Similarly, the sterilizer ports 2594, 2596, 2598 may be configured to couple to end effector 2562 to pneumatically actuate them. By pneumatically acting the sterilizer ports, it may be possible to reduce the number of check valves between the sterilizer ports 2594, 2596, 2598 and switch 2592 and form a sterilizer conduit.

[0299]

[0404] Figure 25B is a schematic diagram of the fluid connector connection processes 2502 and 2504 when the first connector 2510 is coupled to the second connector 2520. For example, when the first connector 2510 and the second connector 2520 are separated and disconnected (2502), the sterilizer port 2594 is in a closed valve configuration. Figure 25C is a detailed schematic diagram of the sterilizer valve 2594. In some variations, when the first connector 2510 is coupled to the second connector 2520 in 2504 and the valve 2594 is pneumatically operated, the valve 2594 may transition to an open valve configuration.

[0300] Liquid transfer bus

[0405] Generally, to enable the transfer of one or more of cell products (i.e., one or more solutions containing cell products), fluids, and reagents between modules, the modules of a cartridge can be connected to each other directly or fluidically via one or more liquid transfer buses. In some variations, the liquid transfer bus may include a portion of the cartridge configured to control the flow and distribution of cell products between modules and storage units. The liquid transfer bus may include a fluid manifold, fluid tubing (e.g., pipes), and one or more valves (but not limited to, one or more of 2 / 2 valves, 3 / 2 valves, 3 / 3 valves, 4 / 2 valves, and rotary selector valves).

[0301]

[0406] To transfer cell products, reagents, or fluids within a cartridge, any pump or other structure can generate a pressure difference between the fluid in one part of the cartridge and the fluid in another part of the cartridge. For example, a cartridge may have one or more pumps, be pre-filled with pressurized fluid contained behind a valve, or be connected to a fluid source or fluid sink. A cartridge may include one or more mechanical pumps (e.g., linear pumps, peristaltic pumps, gear pumps, screw pumps, plunger pumps) or may include multiple parts of a single pump (i.e., the pump can be coupled with a pump actuator). External pressure may be applied to the cartridge, the tubing within the cartridge, or the bag within the cartridge (i.e., pressurizing either the liquid in the bag or the headspace gas in the bag). In some variations, the arrangement of the cartridge components can facilitate gravity-based fluid transfer within the cartridge (e.g., pumping by gravity). One advantage of the disclosed variations is that operator intervention may be reduced, although the systems and methods of this disclosure may use manual operation within the designed workflow or as an aid to automatic operation in the event of imperfect system operation. For example, a process step may involve manual intervention, such as fluid input or output. The operator may intervene in the automated process to correct device operation (e.g., manually compressing a bag to forcefully flush any remaining fluid through the system). The fluid may include liquids and / or gases, and compressed gas supplied from outside or inside the pressurized chamber can be used to generate a liquid flow, such as the transfer of a solution containing cell products from one module to another.

[0302]

[0407] In some variations, the fluid transfer bus may be configured to deliver one or more cell products to each of a series of modules in an order set by the cartridge design or determined by the operation of the system by one or more processors. Similarly, some variations of the cartridge may have the advantage that the order of cell processing steps and process parameters of cell therapy processing steps are controlled by a controller rather than set by the cartridge. In some variations, the fluid transfer bus may be controlled to deliver cell products to modules in any of a variety of sequences or to bypass one or more modules (for example, by configuring the state of one or more valves attached to the fluid bus). In some variations, one module may be used more than once in the cell processing method. Optionally, this method may include performing one or more washing steps. For example, the countercurrent centrifugation elutriation (CCE) module may be used more than once. In an exemplary method, the method may include culturing cell products in a first bioreactor module, transferring the cell products to a CCE module for enrichment of a desired cell type, transferring the cell products to a second bioreactor module for a second culture step, washing the CCE module with a washing solution, and transferring the cell products to the CCE module for a second enrichment step.

[0303]

[0408] In some variations, one or more liquid transfer buses may be fluidly coupled to a plurality of bags or storage units used for dispensing solutions or reagents, storing cell products, or collecting waste liquid or discarded reagents.

[0304]

[0409] In some variations, the cartridge may include one or more pumps that can be fluidly coupled to a liquid transfer bus and / or one or more modules. One or more pumps may include a motor operably coupled to control the circuit and a power source (e.g., a battery or an electrical connector for an external power source). In some variations, the pump may be divided into a pump on the cartridge and a pump actuator on one or more devices of the system. The pump may be an opening in the cartridge, with tubing arranged around this opening, and the pump may be configured to receive a pump actuator (e.g., a peristaltic rotor). The cartridge can be made more compact and simplified by separating the pump components that come into contact with cell products (e.g., tubing) from the pump components that perform the operation of the cell products (i.e., a pump actuator such as a peristaltic rotor). For example, Figures 26A and 26B show the pump head 2610 and pump 2610 of the cartridge in a separated configuration (Figure 13A) and a coupled configuration.

[0305]

[0410] In some variations, one or more pumps 146 (e.g., fluid pumps) can generate a predetermined fluid flow rate for circulating a sterilizer and / or fluid. In some variations, the pumps may include one or more positive displacement pumps (e.g., peristaltic pumps, diaphragm pumps, syringe pumps), centrifugal pumps, or combinations thereof. One or more fluid sources can be connected to the pumps.

[0306]

[0411] In some variations, the pump may be configured to receive a pump signal (generated by the controller) configured to circulate a sterilizing agent for a sufficient residence time to sterilize at least a portion of the fluid connector. For example, the pump may be configured to circulate the sterilizing agent for at least 10 seconds. In some variations, the pump may be configured to receive a pump signal configured to circulate a non-sterilizing gas (e.g., an inert gas, air) to remove the sterilizing agent.

[0307]

[0412] In some variations, peristaltic flow can be generated in discontinuous flow pumps (e.g., peristaltic pumps) when, for example, a tube comes into contact with or is released between rollers. In some variations, peristaltic flow can be compensated for to generate a substantially continuous flow using closed-loop feedback from a flow sensor. For example, a flow sensor can be coupled to a fluid tube to measure the flow rate. A controller can receive the measured flow rate and generate a pump signal to the pump based on a proportional correction function configured to reduce the "ripple" measured by the flow sensor. Additionally or alternatively, the controller may perform periodic error correction on the pump signal to reduce periodic errors that may be unique to each pump. For example, a flow sensor can measure and determine the periodic error of the pump. A pump signal with periodic error correction may correspond to a waveform that includes the inverse shape of the error. The resulting pump flow can compensate for fluctuations in flow rate.

[0308] controller

[0413] In some variations, system 100 may include a controller 120 (e.g., a computing device) which includes one or more of the following: a processor 122, memory 124, communication device 126, input device 128, and display 130. The controller 120 may be configured to control (e.g., operate) the work cell 110. The controller 120 may include multiple devices. For example, the work cell 110 may house one or more components of the controller 120 (e.g., the processor 122, memory 124, and communication device 126), while one or more components of the controller 120 (e.g., the input device 128 and display 130) may be provided remotely from the work cell 110.

[0309] Processor

[0414] The processor described herein (e.g., processor 122) can process data and / or other signals to control one or more components of the system (e.g., work cell 110, controller 120). The processor may be configured to receive, process, compile, compute, store, access, read, write, and / or transmit data and / or other signals. Additionally or alternatively, the processor may be configured to control one or more components of a device and / or one or more components of a controller (e.g., console, touchscreen, personal computer, laptop, tablet, server).

[0310]

[0415] In some variations, the processor may be configured to access or receive data and / or other signals from one or more of the work cells 110, servers, controllers 120, and storage media (e.g., memory, flash devices, memory cards, databases). In some variations, the processor may be any suitable processing device configured to operate and / or execute an instruction set or code, and may include one or more data processors, image processors, graphics processing units (GPUs), physical processing units, digital signal processors (DSPs), analog signal processors, mixed-signal processors, machine learning processors, deep learning processors, finite state machines (FSMs), compression processors (e.g., data compression to reduce data rate and / or memory requirements), cryptographic processors (e.g., for secure wireless data transfer), and / or central processing units (CPUs). The processor may be, for example, a general-purpose processor, a field-programmable gate array (FPGA), an application-specific integrated circuit (ASIC), a processor board, etc. The processor may be configured to operate and / or execute application processes and / or other modules, processes, and / or functions associated with the system.The underlying device technologies can be provided in a wide variety of component types (e.g., metal-oxide semiconductor field-effect transistor (MOSFET) technologies such as complementary metal-oxide semiconductor (CMOS), bipolar technologies such as emitter-coupled logic (ECL), polymer technologies (e.g., silicon-conjugated polymers and metal-conjugated polymer metal structures), and analog-digital hybrids).

[0311]

[0416] The systems, devices, and / or methods described herein may be implemented by software (running on hardware), hardware, or a combination thereof. Hardware modules may include, for example, general-purpose processors (or microprocessors or microcontrollers), field-programmable gate arrays (FPGAs), and / or application-specific integrated circuits (ASICs). Software modules (running on hardware) may be expressed in a wide variety of software languages ​​(e.g., computer code), including structured text, typed documents, C, C++, C#, Java®, Python, Ruby, Visual Basic®, and / or other object-oriented languages, procedural languages, or other programming languages, and development tools. Examples of computer code include, but are not limited to, microcode or microinstructions, machine instructions such as those generated by a compiler, code used to generate web services, and files containing high-level instructions that a computer executes using an interpreter. Additional examples of computer code include, but are not limited to, control signals, encryption code, and compression code.

[0312] memory

[0417] The cell processing systems and devices described herein may include memory (e.g., memory 124) configured to store data and / or information. In some variations, the memory may include one or more of the following: random access memory (RAM), static RAM (SRAM), dynamic RAM (DRAM), memory buffer, erasable programmable read-only memory (EPROM), electrically erasable read-only memory (EEPROM), read-only memory (ROM), flash memory, volatile memory, non-volatile memory, or combinations thereof. In some variations, the memory may store instructions for a processor to execute device-related modules, processes, and / or functions, such as image processing, image display, sensor data, data and / or signal transmission, data and / or signal reception, and / or communication. Some variations described herein may relate to computer storage products comprising a non-temporary computer-readable medium (which may also be called a non-temporary processor-readable medium) having instructions or computer code for performing various operations performed by a computer. Computer-readable media (or processor-readable media) are inherently non-transient in that they do not contain transient propagating signals (e.g., propagating electromagnetic waves that carry information over a transmission medium such as space or a cable). Computer code (which may also be called code or algorithms) may be designed and constructed for one or more specific purposes. In some variations, memory may be configured to store received data and / or data generated by the controller and / or work cell. In some variations, memory may be configured to store data temporarily or permanently.

[0313] Input device

[0418] In some variations, the display may include and / or be operably coupled to an input device 128 (e.g., a touchscreen) configured to receive user input data. For example, user input to the input device 128 (e.g., a keyboard, buttons, or touchscreen) may be received and processed by the system 100's processor (e.g., processor 122) and memory (e.g., memory 124). The input device may include at least one switch configured to generate user input. For example, the input device may include a touch surface for the user to make an input corresponding to the user input (e.g., touching the touchscreen with a finger). The input device including the touch surface may be configured to detect contact and movement on the touch surface using any of a plurality of touch sensitivity techniques, including capacitive, resistive, infrared, optical imaging, dispersed signaling, acoustic pulse recognition, and surface acoustic wave techniques. In a variation of an input device including at least one switch, the switch may have at least one of the following: a button (e.g., a hard key, a soft key), a touch surface, a keyboard, an analog stick (e.g., a joystick), a directional pad, a mouse, a trackball, a jog dial, a step switch, a rocker switch, a pointer device (e.g., a stylus), a motion sensor, an image sensor, and a microphone. The motion sensor can receive user movement data from an optical sensor and classify the user's gestures as user input. The microphone can receive voice data and recognize the user's voice as user input.

[0314]

[0419] In some variations, the cell processing system may optionally include one or more output devices in addition to a display, such as an acoustic device and a tactile device. The acoustic device can output any system data, alarms, and / or notifications audibly. For example, the acoustic device can output an audible alarm if a malfunction is detected. In some variations, the acoustic device may include at least one of a speaker, a piezoelectric acoustic device, a magnetostrictive speaker, and / or a digital speaker. In some variations, a user can communicate with other users using the acoustic device and communication channels. For example, a user can form an acoustic communication channel (e.g., a VoIP phone).

[0315]

[0420] Additionally or alternatively, the system may include a haptic device configured to provide the user with additional sensory output (e.g., force feedback). For example, the haptic device may generate a tactile response (e.g., vibration) to confirm user input to an input device (e.g., a touch surface). As another example, haptic feedback may indicate that user input is being overridden by the processor.

[0316] Communication devices

[0421] In some variations, the controller may include a communication device (e.g., communication device 126) configured to communicate with another controller and one or more databases. The communication device may be configured to connect the controller to another system (e.g., the Internet, a remote server, a database, a work cell) by a wired or wireless connection. In some variations, the system may be in communication with other devices via one or more wired and / or wireless networks. In some variations, the communication device may include a radio frequency receiver, transmitter, and / or optical (e.g., infrared) receiver and transmitter configured to communicate with one or more devices and / or networks. The communication device may communicate by wire and / or wireless.

[0317]

[0422] A communication device may include an RF circuit configured to receive and transmit RF signals. The RF circuit converts electrical signals to electromagnetic signals and can communicate with communication networks and other communication devices via electromagnetic signals. The RF circuit may include well-known circuits for performing these functions. These circuits may include, but are not limited to, antenna systems, RF transceivers, one or more amplifiers, tuners, one or more oscillators, digital signal processors, codec chipsets, subscriber identity module (SIM) cards, memory, and the like.

[0318]

[0423] Wireless communication via any of these devices may use any of several communication standards, protocols, and technologies.These include, but are not limited to, GSM (Global System for Mobile Communications), EDGE (Enhanced Data GSM Environment), High-Speed ​​Downlink Packet Access (HSDPA), High-Speed ​​Uplink Packet Access (HSUPA), EV-DO (Evolution, Data-Only), HSPA, HSPA+, DC-HSPDA (Dual-Cell HSPA), LTE (Long Term Evolution), Near Field Communication (NFC), Wideband Code Division Multiple Access (W-CDMA), Code Division Multiple Access (CDMA), Time Division Multiple Access (TDMA), Bluetooth, WiFi (Wireless Fidelity) (e.g., IEEE 802.11a, IEEE 802.11b, IEEE 802.11g, IEEE 802.11n, etc.), and VoIP (Voice over Internet). This includes Wi-MAX, email protocols (e.g., IMAP (Internet message access protocol) and / or POP (post office protocol)), instant messaging (e.g., XMPP (extensible messaging and presence protocol), SIMPLE (Session Initiation Protocol for Instant Messaging and Presence Leveraging Extensions), IMPS (Instant Messaging and Presence Service)), and / or Short Message Service (SMS), EtherCAT, OPC UA (OPC Unified Architecture), or any other suitable communication protocol.In some variations, the devices described herein can communicate directly with each other without transmitting data over a network (e.g., via NFC, Bluetooth, WiFi, RFID, etc.).

[0319]

[0424] In some variations, the systems, devices, and methods described herein may be in communication with other wireless devices via one or more networks, each of which may be any type of network (e.g., wired network, wireless network). This communication may or may not be encrypted. A wireless network may refer to any type of digital network that is not connected by any kind of cable. Examples of wireless communication in a wireless network include, but are not limited to, cellular, radio, satellite, and microwave communications. However, wireless networks may be connected to wired networks in order to interact with the Internet, other carrier voice and data networks, business networks, and personal networks. Wired networks are typically carried on copper twisted pair wires, coaxial cables, and / or fiber optic cables. Many different types of wired networks exist, including wide area networks (WANs), metropolitan area networks (MANs), local area networks (LANs), Internet area networks (IANs), campus area networks (CANs), global area networks (GANs) such as the Internet, and virtual private networks (VPNs). From this point forward, "network" refers to any combination of wireless, wired, public, and private data networks that are typically interconnected via the Internet to provide integrated networking and information access systems.

[0320]

[0425] Cellular communications can encompass technologies such as GSM, PCS, CDMA, or GPRS, W-CDMA, EDGE, or CDMA2000, LTE, WiMAX, and 5G networking standards. Some wireless network deployments combine networks from multiple cellular networks or use a mixture of cellular, Wi-Fi, and satellite communications.

[0321] display

[0426] Image data can be output to a display of the cell processing system (e.g., display 130). In some variations, the display may include at least one of the following: light-emitting diodes (LEDs), liquid crystal displays (LCDs), electroluminescent displays (ELDs), plasma display panels (PDPs), thin-film transistors (TFTs), organic light-emitting diodes (OLEDs), electronic paper / electronic ink displays, laser displays, and / or holographic displays.

[0322] II. Method

[0427] In general, the systems and devices described herein can perform one or more cell processing steps to produce cell products. Figure 28 is a flowchart of a cell processing method 2800. Method 2800 may include concentrating a selected cell population in a solution (e.g., a fluid) (2802). For example, the solution can be transported to a CCE module in a cartridge via a liquid transfer bus. A robot can be operated to move the cartridge to the CCE instrument, and the CCE module can be coupled with the CCE instrument. The CCE instrument can be operated to concentrate the selected cell population into the CCE module. Additionally or alternatively, for any of the steps described herein, the cell products may be delivered (manually or automatically) into and out of the cartridge via a sterile liquid transfer port. In some variations, the cartridge may be sterilized (manually or automatically) at a feedthrough port.

[0323]

[0428] In some modifications, a selected population of cells in the solution can be washed (2804). For example, the solution can be transported to the cartridge's CCE module via a liquid transfer bus. A robot can be operated to move the cartridge to the CCE instrument, and the CCE module can be linked to the CCE instrument. The CCE instrument can be operated to cause the CCE module to remove culture medium from the solution, add culture medium to the solution, and / or replace the culture medium in the solution.

[0324]

[0429] In some variations, it is possible to select a population of cells in a solution (2806). For example, the solution can be transported to the selection module of a cartridge via a liquid transfer bus. A robot can be operated to move the cartridge to the selection instrument, and the selection module can be linked with the selection instrument. The selection instrument can be operated to select the chosen population of cells and have the selection module select it.

[0325]

[0430] In some variations, cell populations in solution can be classified (2808). For example, the solution can be transported to a classification module in a cartridge via a liquid transfer bus. A robot can be operated to move the cartridge to a classification device, and the classification module can be linked to the classification device. The classification device can be operated to classify cell populations into the classification module.

[0326]

[0431] In some variations, the solution can be transported to the bioreactor module via a liquid transfer bus and allowed to stand (2810). For example, a robot can be operated to move the cartridge to the bioreactor apparatus, and the bioreactor module can be operated in conjunction with the bioreactor apparatus. The bioreactor apparatus can be operated to maintain the cells in the bioreactor module under a predetermined set of conditions.

[0327]

[0432] In some modifications, cells can be grown in solution (2812). For example, the solution can be transported to the bioreactor module of the cartridge via a liquid transfer bus. A robot can be operated to move the cartridge to the bioreactor apparatus, and the bioreactor module can be linked to the bioreactor apparatus. The bioreactor apparatus can be operated to grow cells in the bioreactor module by cell replication.

[0328]

[0433] In some variations, the tissue can be digested by transporting an enzyme reagent via a liquid transfer bus to a module containing a solution that includes the tissue, thereby releasing a selected cell population into the solution (2814).

[0329]

[0434] In some variations, a selected cell population in a solution can be activated by transporting an activation reagent to a module containing a solution of cell products via a liquid transfer bus (2816).

[0330]

[0435] In some variations, the solution can be transported to the electroporation module of the cartridge via a liquid transfer bus and receive an electroporation signal for electroporating cells in the solution (2818). For example, a robot can be operated to move the cartridge to an electroporation apparatus, and the electroporation module can be linked to the electroporation apparatus. The electroporation apparatus can be operated to cause the electroporation module to perform electroporation of a selected population of cells in the presence of genetic material.

[0331]

[0436] In some variations, transduction of a selected cell population in solution can be performed by transporting an effective amount of vector to a module containing a solution of cell products via a liquid transfer bus (2820).

[0332]

[0437] In some variations, the formulation solution can be transported via a liquid transfer bus to a module containing cell products to produce the final cell product (2822). For example, the final cell product can be transported to one or more product collection bags. In some variations, completing the cell product may include one or more of the following steps: washing the cells, concentrating the cells, exchanging the cell buffer for the formulation buffer, and placing a predetermined amount of cells in the formulation buffer into one or more product collection bags and / or containers.

[0333]

[0438] In some variations, the cells can be harvested by manually or automatically removing the cell products from the cartridge (2824).

[0334]

[0439] In some variations, the cell products may include one or more of the following: immune cells, genetically modified chimeric antigen receptor T cells, genetically modified T cell receptor (TCR) cells, hematopoietic stem cells (HSCs), and tumor-infiltrating lymphocytes (TILs). In some variations, the immune cells may include natural killer (NK) cells.

[0335]

[0440] A cell processing method may include a subset of cell processing steps in any appropriate order. For example, a cell processing method may include, in order, a concentration step 2802, a selection step 2806, an activation step 2816, a transduction step 2820, a proliferation step 2812, and a harvesting step 2824. In some variations, a cell processing method may include, in order, a concentration step 2802, a selection step 2806, a standing step 2810, a transduction step 2820, and a harvesting step 2824. In some variations, a cell processing method may include, in order, a tissue digestion step 2820, a washing step 2804, an activation step 2816, a proliferation step 2812, and a harvesting step 2824.

[0336]

[0441] In general, the methods described herein can reduce the cost of cartridges (which may be consumables) by eliminating the complex steps performed in cell processing operations against a set of instructions. In some variations, the cartridge contains cell products (e.g., solutions containing cells) throughout the entire manufacturing process, and various instruments can be coupled with the cartridge at appropriate times to perform one or more cell processing steps. For example, a cell processing step may include transporting cells and reagents to each module within the cartridge. The set of instructions coupled with the cartridge facilitates process flexibility, allowing for the customization of work cells using a given set of instructions for a given cell therapy product. For example, the sequence of cell processing steps can be customized for each cell product, as detailed herein in relation to Figures 35 to 55.

[0337]

[0442] In some variations, the cell product may be retained within the cartridge throughout the entire manufacturing process (e.g., workflow). Additionally or alternatively, the cell product may be removed from the cartridge for one or more cell processing steps, either manually by the operator or automatically via a fluid connector (e.g., SLTP) or other ports on the cartridge. The cell product can then be returned to the same cartridge, transferred to another cartridge, or divided among several cartridges. In some variations, one or more cell processing steps may be performed outside the cartridge. In some variations, processing within the work cell can facilitate sterile cell processing within the cartridge.

[0338]

[0443] Figure 29 is a flowchart of a cell processing method, showing cell processing steps performed on a cartridge (e.g., consumable) within a work cell including a CCE instrument module, a sterile fluid transfer (SLT) instrument module, and a bioreactor instrument module. The consumable may be configured to perform one or more cell processing steps in conjunction with any of the CCE instrument module, SLT instrument module, and bioreactor instrument module. For example, a robot (or operator) may be configured to move the cartridge between any of the modules in the work cell. A pump head within the instrument can engage with the consumable cartridge to perform fluid transfer between modules of the cartridge, fluid transfer in and out of various storage sections within the cartridge, and / or fluid transfer via ports that allow for the addition or removal of reagents to or from the cartridge.

[0339]

[0444] In some variations, the CCE instrument module may include a pump and centrifuge configured to work with a cartridge (e.g., consumables). The SLT instrument module may include one or more fluid connectors configured to work with one or more of the cartridge bag and bioreactors. The bioreactor instrument module may include one or more sensors, temperature controllers, pumps, agitators, etc., and may be configured to work with the cartridge. In some variations, cell products may be contained within the cartridge throughout the entire cell processing.

[0340]

[0445] The cell processing method shown in Figure 29 may include using a pump to move the fluid in the product bag (e.g., cells in solution) to the CCE module (e.g., rotor) of the cartridge (e.g., consumable) (2910). In some variations, the fluid can be concentrated using the CCE module (2912). For example, blood components can be collected in a garbage bag (2913). In some variations, the fluid can be washed using the CCE module (2914). For example, buffers can be collected in a garbage bag (2915). In some variations, the culture medium can be replaced using the CCE module (2916). For example, one or more buffers (e.g., formulation buffers) and culture medium can be collected in a garbage bag (2917). In some variations, the fluid can be moved to the bioreactor of the cartridge (2918).

[0341]

[0446] In some variations, the fluid connector can fill the bag with reagents (2920). In some variations, reagents (e.g., beads, vectors) can be added to the bioreactor in the cartridge (2922). In some variations, the fluid connector can remove waste from the bag (2924). In some variations, the fluid connector may optionally remove the sample from the bioreactor.

[0342]

[0447] In some modifications, cells can be moved to a bioreactor (2930). In some modifications, cells can be activated or genetically modified (2932). In some modifications, cells can be cultured (2934). In some modifications, cells can be perfused using a pump (2936). For example, used culture medium may be collected in a garbage bag (2937). In some modifications, cells can be proliferated (2938). In some modifications, cells can be harvested after a culture medium change (2940).

[0343]

[0448] Figure 30A is a flowchart of a cell processing method for autologous CAR-T cells or genetically modified TCR cells. Method 3000 may include the steps of enrichment, selection, activation, genetic recombination, proliferation, harvesting / formulation, and cryopreservation. Figure 30B is a flowchart of a cell processing method for allogeneic CAR-T cells or genetically modified TCR cells. Method 3010 may include the steps of enrichment, activation, genetic recombination (e.g., transduction, transfection), alpha / beta T cell depletion, proliferation, harvesting / pooling / formulation, and cryopreservation.

[0344]

[0449] Figure 31 is a flowchart of a method for processing hematopoietic stem cells (HSCs). Method 3100 may include the steps of concentration, selection, standing, genetic recombination, harvesting / formulation, and cryopreservation.

[0345]

[0450] Figure 32 is a flowchart of a method for processing tumor-infiltrating lymphocytes (TILs). Method 3200 may include the steps of tissue digestion, washing, selection, activation, proliferation, harvesting / preparation, and cryopreservation.

[0346]

[0451] Figure 33 is a flowchart of a method for processing natural killer (NK) CAR cells. Method 3300 may include steps of enrichment, selection, activation, genetic recombination, proliferation, harvesting / formulation, and cryopreservation.

[0347]

[0452] Figures 34A to 34C show the controllability T(T reg This is a flowchart of cell processing methods. Method 3400 may include the steps of concentration, selection, harvesting / formulation, and cryopreservation. Method 3402 may include the steps of concentration, selection, activation, genetic recombination, proliferation, selection (arbitrarily), harvesting / formulation, and cryopreservation. Method 3404 may include the steps of concentration, selection, activation / proliferation, and introducing a feeder cell culture for harvesting / irradiation. Concentration, selection, co-culture with processed feeder cells, harvesting, and cryopreservation can be performed on another cell set.

[0348]

[0453] Figures 98 to 101 are flowcharts of cell processing methods for cell therapy workflows including splitting (e.g., parallel) processing. Method 9800 may include steps of concentration, selection, activation, recombination, proliferation, formulation, and cryopreservation. For example, cell processing method 9800 (e.g., workflow) may include splitting the cell product into two or more parts after the concentration step. The split parts may be processed in parallel within a single cartridge. In some variations, one or more split parts may be transferred to two or more cartridges for parallel processing. One or more cell processing parameters (e.g., timing of process steps, type of reagents added, transfection components, etc.) may be configured independently for each split part of the cell product. In some variations, the split parts may be pooled after the proliferation step.

[0349]

[0454] Method 9900 may include steps of concentration, selection, activation, recombination, proliferation, formulation, and cryopreservation. For example, cell processing method 9900 (e.g., workflow) may include splitting the cell product into two or more parts after the activation step. The split parts may be processed in parallel within a single cartridge. In some variations, one or more split parts may be transferred to two or more cartridges for parallel processing. One or more cell processing parameters (e.g., timing of process steps, type of reagents added, transfection components, etc.) may be configured independently for each split part of the cell product. In some variations, the split parts may be pooled after the proliferation step and / or recombination step.

[0350]

[0455] Method 10000 may include steps of concentration, selection, activation, genetic recombination, proliferation, formulation, and cryopreservation. For example, cell processing method 10000 (e.g., workflow) may include splitting the cell product into two or more parts after the selection step. The split parts may be processed in parallel within a single cartridge. In some variations, one or more split parts may be transferred to two or more cartridges for parallel processing. One or more cell processing parameters (e.g., timing of process steps, type of reagents added, transfection components, etc.) may be configured independently for each split part of the cell product. In some variations, the split parts may not be pooled.

[0351]

[0456] Method 10100 may include steps of concentration, selection, activation, genetic recombination, growth, formulation, and cryopreservation. For example, cell processing method 10100 (e.g., workflow) may include splitting a cell product into two or more parts from a starting material. The separated products can be kept separate and processed in parallel as split parts in a single cartridge or multiple cartridges. One or more cell processing parameters (e.g., timing of process steps, type of reagents added, transfection components, etc.) may be configured independently for each split part. In some variations, the split parts can be pooled after the growth step.

[0352]

[0457] Figure 102 is a schematic diagram of a cell processing system 10200 configured for dissociation processing within a single cartridge. For example, methods 9800-10100 described in association with Figures 98 to 101 can be performed within cartridge 10210. In some variations, system 10200 may include a sterile fluid transfer device 10220 containing reagents 10222, and cartridge 10210 containing multiple bioreactor modules 10230, a pump module 10240, a thermal module 10245, a pressure-driven flow module 10250, a MACS module 10255, an electroporation module 10260, a FACS module 10265, a CCE module 10270, and a blank module 10280. Cartridge 10210 may further include a reagent storage section 10285, multiple product bags 10290, and a liquid transfer bus 10295. The liquid transfer bus 10295 may be configured to connect the components of the cartridge 10210 for fluid communication.

[0353]

[0458] In some variations, the insertion of cell products into and removal of cell products from the cartridge may be performed within or outside the system. In some variations, the cartridge is fitted to the patient or donor at bedside and then sent to a cell processing system within or near the hospital, or shipped to a facility where a cell processing system is installed. Similarly, the cell products may be removed from the cartridge after processing at the facility or near the recipient (patient) of the cell products. Optionally, the cell products are frozen after optionally adding one or more cryoprotectants to the cell products before, during, or after the method of this disclosure. In some variations, the system may include a freezer and / or a liquid nitrogen source. In some variations, the system may include a water bath or a heating chamber containing gas at a controlled temperature to allow for controlled thawing of the cell products. This is, for example, a water bath set to approximately 20°C to approximately 40°C. In some variations, the cartridge is made of a material that can withstand mechanical damage when refrigerated.

[0354] Automated cell processing

[0459] Described herein are methods for converting user-defined cell processing operations into cell processing steps using the automated cell processing systems and devices described herein. Some variations accept and convert cell processing operations into cell processing steps for execution by the system, subject to a predetermined set of constraints. For example, the user can input a set of biological process steps and corresponding biological process parameters to be performed by the cell processing system. Optionally, process parameters can be customized in each cartridge or in sets of cartridges.

[0355]

[0460] Figure 35 is a flowchart that provides an overall description of variations of the automated cell processing method. Method 3500 may include receiving an ordered input list of cell processing operations (3502). For example, to be executed in two or more cartridges in an automated cell processing system, two or more sets of ordered input lists of cell processing operations may be received. For example, as shown in GUI 4900 in Figure 49 and detailed herein, one or more biological process inputs (e.g., available operations) may be selected as the ordered input list of cell processing operations, such as enrichment, MACS selection, activation, transduction, transfection, proliferation, and inline analysis. Furthermore, GUI 5200 in Figure 52 shows a complete ordered input list (e.g., selected set of operations) 5220 of cell processing operations selected by the user.

[0356]

[0461] In some variations, one or more sets of cell processing parameters can be received (3504). Each set of cell processing parameters can be associated with one of the cell processing operations. Each set of cell processing parameters can define the characteristics of the cell processing step performed by the instrument in the cell processing step. For example, GUI4000 in Figure 40 shows reagent and container parameters, GUI4200 in Figure 42 shows an example of process parameters, GUI4400 in Figure 44 shows an example of pretreatment analysis, and GUI4800 in Figure 48 shows an example of an activation setting set.

[0357]

[0462] In some variations, the transformation model can be performed on an ordered input list (3506). In some variations, the transformation model may include constraints on an ordered output list determined by a given configuration of the automated cell processing system. For example, the constraints may include information about the configuration of the automated cell processing system.

[0358]

[0463] In some variations, the constraints may include one or more of the following: the type and / or number and / or condition of the instrument, the type and / or number and / or condition of the modules on the cartridge, the type and / or number of storage units on the cartridge, the type and / or number of sterile fluid transfer ports on the cartridge, and the number and location of the fluid paths between the modules, storage units and sterile fluid transfer ports on the cartridge.

[0359]

[0464] In some variations, a set of predetermined constraints can be added to the set of process control parameters. For example, the volume and / or type of reagents used may be constrained based on the size of the system and / or the product being manufactured. Other process parameter constraints may include, but are not limited to, one or more of the following, or combinations thereof: temperature, volume, time, pH, cell size, cell number, cell density, cell viability, dissolved oxygen, glucose level, onboard reagent storage volume and waste volume. For example, GUI4000 in Figure 40 indicates that the reagent has a unit volume of 30 mL, the required volume is 54 mL, and the consumable container has a unit volume of 75 mL. GUI4800 in Figure 48 indicates that the activation concentration is 12 mg / L, the activation culture time is 1600 seconds, the activation temperature is 18°C, and the gas mixture contains 21% oxygen, 78.06% nitrogen, and 0.04% carbon dioxide. By applying these constraints using a transformation model, an ordered output list of cell processing steps can be generated that influence how one or more of the robots, instruments, and cartridges operate to produce cell products.

[0360]

[0465] In some variations, the order of operations can be constrained based on hardware limitations. For example, a robot may be limited to moving one cartridge at a time. Similarly, an instrument may be limited to operating on a predetermined number of cartridges at a time.

[0361]

[0466] In some variations, as shown in GUI4900 in Figure 49, the product insertion operation must be the first operation performed and may be performed once in each process. The filling and finishing operations are always the last operations performed before product completion and may be performed once in each process.

[0362]

[0467] In some variations, the system can prevent the user from executing a set of actions in an order that the system cannot execute.

[0363]

[0468] In some variations, notifications (e.g., warnings, alarms) may be issued if the user orders a set of actions in a "non-standard" manner. For example, a notification may be issued if the same type of action is repeated consecutively (e.g., enrichment immediately after enrichment). Similarly, a notification may be issued if an action (e.g., selection, activation) that is normally used only once within a given process is used more than once.

[0364]

[0469] In some variations, the output of the transformation model may correspond to an ordered output list of cell processing steps that the system can perform (3508). For example, the transformation model can be run on a set of ordered input lists to generate an ordered output list of cell processing steps. The output list of cell processing steps can control a robot, a cartridge, and one or more instruments.

[0365]

[0470] In some variations, an ordered output list is executed by the system to control the robot to move one or more cartridges, each containing cell products, between instruments (3510). For example, a MACS selection process selected by the user may correspond to the robot 230 in Figure 2 moving cartridge 250 from, for example, another instrument to cell selection instrument 216. In some variations, the ordered output list may include instructions for the robot to load cartridges (e.g., single-use consumables) into a cell processing system (e.g., a work cell). Furthermore, the robot may be configured to move the cartridges to a first instrument position.

[0366]

[0471] In some variations, the ordered output list is further executed by the system to control one or more instruments to perform one or more cell processing steps on one or more cell products in each cartridge (3512). For example, a computing server rack 210 (e.g., controller 120) may be configured to control an electroporation module 220 configured to apply a pulsed electric field to the cell suspension in cartridge 250. In some variations, the ordered output list may include instructions for an instrument (e.g., a bioreactor) to process the product (e.g., transfer cell products from a small bioreactor module to a large bioreactor module). The instrument may also be configured to operate under a set of process parameters (e.g., a period of 9 hours, pH 6.7, temperature 37.3°C to 37.8°C, mixing mode 3). As another example, the ordered output list may include instructions for operating a sterile fluid transfer module to perform one or more of the following: removal of waste from a cartridge, addition of culture medium to a cartridge, and addition of MACS reagent to a cartridge.

[0367]

[0472] In some variations, one or more electronic batch records can be generated based on process parameters and data collected from sensors during process execution (3514). The batch records generated by the system may include process parameters, time logs, sensor measurements from instruments, QC parameters determined by QC instruments, and other records.

[0368]

[0473] Figure 36 is a flowchart that provides an overall description of the variations of method 3600 for executing the transformation model. Some variations can generate and output one or more biological functions to the user. For example, a set of configurable biological function blocks can be displayed in a graphical user interface for user selection. The GUI allows the user to select and order the biological function blocks and define biological control parameters. If desired, one or more control parameters of the biological function blocks may be modified by the user. Some variations can generate one or more biological function templates containing a default sequence of biological function blocks. If desired, one or more biological control parameters of the biological function templates may be modified by the user.

[0369]

[0474] In some variations, the cell processing system may be configured to receive and / or store one or more biological function (e.g., process) inputs from the user (3604). For example, the user may select one or more default biological function templates.

[0370]

[0475] In some variations, a biological process model (e.g., process definition) can be generated based on a biological process input (3606). In some variations, the biological process model may include one or more of the following: enrichment, isolation, MACS selection, FACS selection, activation, recombination, gene transfer, transduction, transfection, proliferation, formulation (e.g., harvesting, pooling), cryopreservation, T cell depletion, static storage, tissue digestion, washing, irradiation, co-culture, and combinations thereof.

[0371]

[0476] In some variations, a biological process model can be transformed into an instrument execution process model (3608). For example, each biological function block in a biological process model may correspond to an ordered list of cell processing system operations with corresponding hardware control parameters. An instrument execution process model may include a sequence of hardware operations corresponding to a biological process model. As described herein, the transformation model may include one or more constraints.

[0372]

[0477] Optionally, in some variations, the cell processing system may be configured to receive and / or store one or more instrument execution process inputs from the user (3610). For example, the user may, if desired, modify the converted instrument execution process model. The user may select specific hardware components for executing several steps and changing timing parameters, etc.

[0373]

[0478] In some variations, the instrument execution process can be performed to generate cell products (3612). For example, a cell processing system can process cell products through the system at runtime as defined by the instrument execution process model.

[0374]

[0479] In some variations, the instrument execution process can be performed (3612). In some variations, the instrument execution process model can be transformed back into a biological process model (3614). The progress of this biological process may be output (e.g., displayed) to the user for monitoring. For example, the instrument execution process model may include one or more criteria (e.g., pointers) that are returned to the biological process model, enabling it to report runtime execution progress against the biological process model.

[0375]

[0480] In some variations, cell products can be monitored (3616). For example, GUIs 5300 and 5400 in Figures 53 and 54 show sensor data monitored by the system for multiple products. For example, the status of numerous viable cells and processes can be graphically displayed to the user (e.g., as a function of completion rate).

[0376]

[0481] In some variations, electronic records can be generated based on monitoring data (3618). For example, one or more electronic batch records can be generated in accordance with the 21 CFR regulations.

[0377]

[0482] Figure 55 is a block diagram of an exemplary variation of a manufacturing workflow 5500, including a processing platform 5520 (e.g., system 100, work cells 110, 200, 201) configured to produce multiple cell products (e.g., a first product, a second product, a third product) in parallel. For example, a first workflow 5510 for a first product may include multiple biological processes 5512 executed in a predetermined sequence using corresponding elements 5522 (e.g., hardware) of platform 5520. Simultaneously, a second workflow 5530 for a second product may execute a predetermined sequence of biological processes 5530 using corresponding elements 5524 of platform 5520. In this way, the hardware resources of platform 5520 can be efficiently utilized to increase throughput. In some variations, approximately 2, 3, 4, 5, 6, 7, 8, 9, 10, or more cell products can be produced simultaneously on platform 5520. The conversion model may include hardware constraints to eliminate scheduling overlaps, for example, to ensure that the same instrument is not used for different products simultaneously.

[0378] Graphical User Interface

[0483] In some variations, a graphical user interface (GUI) may be configured for process design and monitoring of the generated products. Figure 37 shows a variation of GUI 3700 that includes an initial process design interface. For example, GUI 3700 may be the process design homepage. GUI 3700 may indicate that no process is selected or loaded. The user can select the generate icon 3710 (e.g., "Generate Process") to start the process design process. In some variations, one or more of the GUIs described herein may include a search bar.

[0379]

[0484] Figure 38 shows a variation of GUI 3800 related to process generation. GUI 3800 may appear after selecting the generation icon 3710 in Figure 37. For example, GUI 3800 may include a process generation window 3810 that allows the user to input and / or select one or more of the following: process name, process description, and template. In some variations, the user can select from a given list of templates. For example, the user can generate a process and save it as a template for later selection.

[0380]

[0485] Figure 39 shows a variation of GUI3900, including its relationship to an empty process. GUI3900 may be displayed after confirming that a process has been generated in GUI3800. GUI3900 displays the process name (e.g., Car T treatment), highlights the process setup icon 3910, and allows the addition of process-specific parameters such as process reagents and containers, process parameters, and pretreatment analyses. GUI3900 may further include icons for adding process reagents and containers 3920, adding process parameters 3930, and adding pretreatment analyses 3940. Once the process setup is complete, one or more process elements can be specified.

[0381]

[0486] In some variations, GUI3900 may include one or more predetermined templates for a set of biological processes (e.g., CAR-T, NK cells, HSCs, TILs, etc.). For example, a template can assist in process development and can serve as a valid starting point for process development. The template can also be modified (e.g., customized) based on user requirements.

[0382]

[0487] Figure 40 shows a variation of GUI 4000 that includes the addition of reagent and consumable containers. GUI 4000 may appear after selecting the process reagent and container addition icon 3920 in Figure 39. For example, GUI 4000 may include a reagent and container addition window 4010 that allows the user to input and / or select one or more reagents, including reagent type, manufacturer, part number, unit volume, required volume, and required reagent input (e.g., lot number, expiration date, container transfer requirement). The reagent and container addition window 4010 may include one or more of the following: input fields, selection boxes, dropdown selectors, etc. Furthermore, the reagent and container addition window 3810 allows the user to input and / or select one or more consumable containers, including manufacturer, part number, unit volume, and required container input (e.g., lot number, expiration date). In some variations, the user can select from a given list of templates. For example, the user can generate a process and save it as a template.

[0383]

[0488] Figure 41 shows a variation of GUI 4100 that includes the association of process parameters. GUI 4100 may appear after selecting the process reagent and container addition icon 3930 in Figure 39. For example, GUI 4100 may include a process parameter addition window 4110 that allows the user to input and / or select one or more parameters, including name, parameter identifier, description, data type, unit, and parameter type. The process parameter addition window 4010 may include one or more of the following: input fields, selection boxes, dropdown selectors, etc. In some variations, the user can select from a given list of templates. For example, the user can generate parameters and save them as a template. Figure 42 shows a variation of GUI 4200 that includes the association of patient weight process parameters. For example, GUI 4200 may include a process parameter addition window 4110 pre-filled with parameter information, including patient weight, data type (e.g., integer), unit (e.g., kilogram), and parameter type (e.g., input).

[0384]

[0489] Figure 43 shows a variation of GUI 4300 related to preprocessing analysis. GUI 4300 may appear after selecting the preprocessing analysis add icon 3940 in Figure 39. For example, GUI 4300 may include a preprocessing analysis add window 4310 that allows the user to input and / or select one or more parameters, including name, identifier, description, data type, and display group. The preprocessing analysis add window 4310 may include one or more of the following: input fields, selection boxes, dropdown selectors, etc. In some variations, the user can select from a given list of templates. For example, the user can generate parameters and save them as a template.

[0385]

[0490] Figure 44 shows a variation of GUI 4400 related to white blood cell count pretreatment analysis. For example, GUI 4400 may include an additional pretreatment analysis window 4410 in which pretreatment analysis information is entered, including a name (e.g., CBC white blood cell count), identifier (e.g., CBC white blood cell count), description (e.g., number of white blood cells in the sample), data type (e.g., float type), and display group (e.g., WBC).

[0386]

[0491] Figure 45 shows a variation of GUI 4500 related to process parameter calculation. GUI 4500 may appear after selecting the preprocessing analysis add icon 3940 and the "Calculate" parameter type in Figure 39. For example, GUI 4500 may include a preprocessing analysis add window 4510 that allows the user to input and / or select one or more parameters, including name, identifier, description, data type, display gro...

Claims

1. Multiple instruments, each independently configured to perform one or more cell processing operations on a cartridge, A robot capable of moving the cartridge between each of the aforementioned multiple devices, A system for cell processing that includes the following features.

2. The system according to claim 1, wherein the system is housed within a work cell.

3. The system according to claim 1 or 2, wherein the work cell is automated.

4. The system according to any one of claims 1 to 3, wherein the plurality of instruments are configured to work in conjunction with the cartridge to perform a cell processing operation on the cartridge.

5. The system according to any one of claims 1 to 4, wherein the system comprises a processor, and the processor is configured to control the robot and the plurality of devices.

6. The system according to any one of claims 1 to 5, wherein the system is configured to receive two or more cartridges.

7. The system according to any one of claims 1 to 6, wherein the system includes the cartridge.

8. The system according to any one of claims 1 to 7, wherein the cartridge includes a plurality of modules.

9. The system according to any one of claims 1 to 8, wherein the cartridge includes a bioreactor module.

10. The system according to any one of claims 1 to 9, wherein the cartridge includes a cell selection module.

11. The system according to claim 10, wherein the cell selection module is a magnetically activated cell selection module.

12. The system according to any one of claims 1 to 11, wherein the cartridge includes a classification module.

13. The system according to claim 12, wherein the classification module is a fluorescence-activated cell classification (FACS) module.

14. The system according to any one of claims 1 to 13, wherein the cartridge includes an electroporation module.

15. The system according to any one of claims 1 to 14, wherein the cartridge includes a countercurrent centrifugal ertriation (CCE) module.

16. The system according to any one of claims 1 to 15, wherein the cartridge includes one or more sterile fluid transfer ports.

17. The system according to any one of claims 1 to 16, wherein the cartridge includes a liquid transfer bus fluidly coupled to each module.

18. The system according to any one of claims 1 to 17, wherein the cartridge includes a pump fluidically coupled to the liquid transfer bus.

19. The system according to any one of claims 1 to 18, wherein the system includes a pump actuator configured to work in conjunction with the pump.

20. The system according to any one of claims 1 to 19, wherein the system includes a bioreactor device.

21. The system according to claim 20, wherein the bioreactor device includes a plurality of slots for a cartridge.

22. The system according to any one of claims 1 to 21, wherein the system includes a cell selection device.

23. The system according to claim 22, wherein the cell selection device is a magnetically activated cell selection device.

24. The system according to any one of claims 1 to 23, wherein the system includes a classification device.

25. The system according to claim 24, wherein the classification instrument is a fluorescence-activated cell classification (FACS) instrument.

26. The system according to any one of claims 1 to 25, wherein the system includes an electroporation apparatus.

27. The system according to any one of claims 1 to 26, wherein the system includes a countercurrent centrifugal elutriation (CCE) apparatus.

28. The system according to any one of claims 1 to 27, wherein the system includes a reagent storage container.

29. The system according to any one of claims 1 to 28, wherein the cartridge includes a bioreactor module and a selection module.

30. The system according to any one of claims 1 to 29, wherein the cartridge includes a bioreactor module and a CCE module.

31. The system according to any one of claims 1 to 30, wherein the cartridge includes a bioreactor module, a selection module, and a CCE module.

32. The system according to any one of claims 1 to 31, wherein the cartridge includes a bioreactor module, a selection module, and an electroporation module.

33. The system according to any one of claims 1 to 32, wherein the cartridge includes a bioreactor module, a selection module, a CCE module, and an electroporation module.

34. The system according to any one of claims 1 to 32, wherein the cartridge includes a second bioreactor module having an internal volume that is two times, five times, or ten times larger than the internal volume of the first bioreactor.

35. The system according to any one of claims 1 to 33, wherein the system includes an enclosure.

36. The system according to claim 34, wherein the enclosure is an ISO 7 cleanroom.

37. The system according to claim 34, wherein the enclosure is an ISO 6 cleanroom.

38. The system according to claim 34, wherein the enclosure is an ISO 5 cleanroom.

39. The system according to any one of claims 33 to 34, wherein the enclosure includes a feedthrough.

40. The system according to any one of claims 1 to 38, wherein the system performs automated production of cell products.

41. A cartridge for cell processing comprising a liquid transfer bus and a plurality of modules, wherein each module is fluidically coupled to the liquid transfer bus.

42. The cartridge according to claim 41, wherein the cartridge includes one or more sterile liquid transfer ports.

43. The cartridge according to any one of claims 40 to 41, wherein the cartridge includes a bioreactor module.

44. The cartridge according to any one of claims 40 to 41, wherein the cartridge includes a cell selection module.

45. The cartridge according to claim 44, wherein the cell selection module is a magnetically activated cell selection module.

46. The cartridge according to any one of claims 40 to 45, wherein the cartridge includes a classification module.

47. The cartridge according to claim 46, wherein the classification module is a fluorescence-activated cell classification (FACS) module.

48. The cartridge according to any one of claims 40 to 47, wherein the cartridge includes an electroporation module.

49. The cartridge according to any one of claims 40 to 48, wherein the cartridge includes a countercurrent centrifugal ertriation (CCE) module.

50. The cartridge according to any one of claims 40 to 49, wherein the cartridge includes a mechanoporation module.

51. The cartridge according to any one of claims 40 to 50, wherein the cartridge includes a second bioreactor module having an internal volume that is two times, five times, or ten times larger than the internal volume of the first bioreactor.

52. The cartridge according to any one of claims 40 to 51, wherein the cartridge includes a bioreactor module, a selection module, and a CCE module.

53. The cartridge according to any one of claims 40 to 52, wherein the cartridge comprises a bioreactor module, a selection module, and an electroporation module.

54. The cartridge according to any one of claims 40 to 53, wherein the cartridge comprises a bioreactor module, a selection module, a CCE module, and an electroporation module.

55. A method for processing cells, This involves moving a cartridge containing cell products between multiple instruments within a closed automated work cell. The apparatus is a method that works in conjunction with the cartridge to perform a cell processing step on the cell product.

56. The method according to claim 55, comprising performing a cell treatment step on the cell product.

57. The method according to any one of claims 55 to 56, wherein for each cell product, all cell processing steps in the method are performed in a single cartridge.

58. The method according to any one of claims 55 to 57, further comprising dividing the cell product into a plurality of cell product portions.

59. The method according to any one of claims 58, wherein the method comprises performing the cell processing step in parallel on the plurality of cell product portions.

60. The method according to any one of claims 58 to 59, further comprising combining at least two of the plurality of cell product portions.

61. The method according to any one of claims 55 to 57, wherein the work cell includes a robot configured to move a cartridge.

62. The method according to any one of claims 55 to 61, wherein the work cell includes a processor, and the processor is configured to control the robot and the plurality of devices.

63. The method according to any one of claims 55 to 62, wherein the work cell is configured to receive two or more cartridges.

64. The method according to any one of claims 55 to 63, wherein the cartridge includes a plurality of modules.

65. The method according to any one of claims 55 to 64, wherein the cartridge includes a bioreactor module.

66. The method according to any one of claims 55 to 65, wherein the cartridge includes a cell selection module.

67. The method according to claim 66, wherein the cell selection module is a magnetically activated cell selection module.

68. The method according to any one of claims 55 to 67, wherein the cartridge includes a classification module.

69. The method according to claim 68, wherein the classification module is a fluorescence-activated cell classification (FACS) module.

70. The method according to any one of claims 55 to 69, wherein the cartridge includes an electroporation module.

71. The method according to any one of claims 55 to 70, wherein the cartridge includes a countercurrent centrifugal elutriation (CCE) module.

72. The method according to any one of claims 55 to 71, wherein the cartridge includes one or more sterile fluid transfer ports.

73. The method according to any one of claims 55 to 72, wherein the cartridge includes a liquid transfer bus fluidly coupled to each module.

74. The method according to any one of claims 55 to 73, wherein the cartridge includes a pump fluidically coupled to the liquid transfer bus.

75. The method according to any one of claims 55 to 74, wherein the work cell includes a pump actuator configured to work in conjunction with the pump.

76. The method according to any one of claims 55 to 75, wherein the work cell includes a bioreactor device.

77. The method according to claim 76, wherein the bioreactor device includes a plurality of slots for a cartridge.

78. The method according to any one of claims 55 to 77, comprising performing the cell processing step in parallel for two or more cartridges.

79. The method according to any one of claims 55 to 78, wherein the work cell includes a cell selection device.

80. The method according to claim 79, wherein the cell selection device is a magnetically activated cell selection device.

81. The method according to any one of claims 55 to 80, wherein the work cell includes a sorting device.

82. The method according to claim 81, wherein the classification instrument is a fluorescence-activated cell classification (FACS) instrument.

83. The method according to any one of claims 55 to 82, wherein the work cell includes an electroporation apparatus.

84. The method according to any one of claims 55 to 83, wherein the work cell includes a countercurrent centrifugal ertriation (CCE) apparatus.

85. The method according to any one of claims 55 to 84, wherein the work cell includes a reagent storage container.

86. The method according to any one of claims 55 to 85, wherein the cartridge includes a bioreactor module and a selection module.

87. The method according to any one of claims 55 to 86, wherein the cartridge includes a bioreactor module and a CCE module.

88. The method according to any one of claims 55 to 87, wherein the cartridge includes a bioreactor module, a selection module, and a CCE module.

89. The method according to any one of claims 55 to 88, wherein the cartridge comprises a bioreactor module, a selection module, and an electroporation module.

90. The method according to any one of claims 55 to 89, wherein the cartridge comprises a bioreactor module, a selection module, a CCE module, and an electroporation module.

91. The method according to any one of claims 55 to 90, wherein the work cell includes an enclosure.

92. The method according to claim 91, wherein the enclosure is an ISO 7 cleanroom.

93. The method according to claim 91, wherein the enclosure is an ISO 6 cleanroom.

94. The method according to claim 91, wherein the enclosure is an ISO 5 cleanroom.

95. The method according to claim 91, wherein the enclosure includes a feedthrough.

96. The method according to any one of claims 55 to 95, wherein the method performs automated production of cell products.

97. The method according to claim 96, wherein the cell product is a chimeric antigen receptor (CAR) T cell product.

98. The method according to claim 96, wherein the cell product is a natural killer (NK) cell product.

99. The method according to claim 96, wherein the cell product is a hematopoietic stem cell (HSC) cell product.

100. The method according to claim 96, wherein the cell product is a tumor-infiltrating lymphocyte (TIL) cell product.

101. The method according to claim 96, wherein the cell product is a regulatory T (Treg) cell product.

102. A method for processing a solution containing cell products, which is performed in an automated system, A concentration step comprising: transporting the solution to the CCE module of the cartridge via a liquid transfer bus; operating the robot to move the cartridge to the CCE device and linking the CCE module with the CCE device; and operating the CCE device to concentrate the selected cell population in the solution into the CCE module, thereby concentrating the selected cell population; A washing step comprising washing a selected cell population in the solution by transporting the solution from the cartridge to the CCE module via the liquid transfer bus, operating the robot to move the cartridge to the CCE device and linking the CCE module with the CCE device, operating the CCE device to cause the CCE module to remove the culture medium from the solution, to add the culture medium to the solution, and / or to replace the culture medium in the solution, A selection step comprising transporting the solution to the cartridge selection module via the liquid transfer bus, operating the robot to move the cartridge to the selection device to link the selection module with the selection device, and operating the selection device to cause the selection module to select the selected cell population in the solution, thereby selecting the selected cell population, A classification step comprising transporting the solution to the sorting module of the cartridge via the liquid transfer bus, operating the robot to move the cartridge to the sorting device to link the sorting module with the sorting device, and operating the sorting device to cause the sorting module to sort the cell population in the solution, thereby classifying the cell population, A settling step includes transporting the solution from the cartridge to the bioreactor module via the liquid transfer bus, operating the robot to move the cartridge to the bioreactor device, linking the bioreactor module with the bioreactor device, and operating the bioreactor device to maintain the cells in the bioreactor module. A proliferation step comprising: transporting the solution from the cartridge to the bioreactor module via the liquid transfer bus; operating the robot to move the cartridge to the bioreactor device; linking the bioreactor module with the bioreactor device; operating the bioreactor device to proliferate the cells in the solution in the bioreactor module by cell replication, thereby proliferating the cells; A tissue digestion step comprising transporting an enzyme reagent to a module containing a solution containing tissue via the liquid transfer bus, and releasing a selected cell population into the solution by the enzyme reagent digesting the tissue, An activation step comprising activating a selected cell population in the solution by transporting an activation reagent to a module containing the solution containing the cell product via the liquid transfer bus, An electroporation step comprising: transporting the solution to the electroporation module of the cartridge via the liquid transfer bus; operating the robot to move the cartridge to the electroporation apparatus to link the electroporation module with the electroporation apparatus; operating the electroporation apparatus to cause the electroporation module to perform electroporation of the selected cell population in the presence of the vector; Transduction step includes performing transduction of a selected cell population in the solution by transporting an effective amount of vector to a module containing the solution containing the cell product via the liquid transfer bus, A filling / completion step includes transporting the formulation solution to a module containing the cell product via the liquid transfer bus to produce the final cell product, and transporting the final cell product to one or more product collection bags, A method comprising one or more cell processing steps, selected from and performed sequentially in any order.

103. The method according to claim 102, wherein the cartridge is sterilized manually or automatically at the feed-through port.

104. The method according to claim 102, comprising manually or automatically introducing one or more of the fluid and the cell products into the cartridge via a sterile liquid transfer port.

105. The method according to claim 102, comprising a collection step of manually or automatically removing the cell product from the cartridge.

106. The method according to claim 102, wherein the cell product includes immune cells.

107. The method according to claim 106, in order: the concentration step, the selection step, the activation step, the transduction step, the proliferation step, and the harvesting step.

108. The method according to claim 106, wherein the immune cells are genetically modified chimeric antigen receptor T cells.

109. The method according to claim 106, wherein the immune cells are genetically modified T cell receptor (TCR) cells.

110. The method according to claim 106, wherein the immune cells include natural killer (NK) cells.

111. The method according to claim 102, wherein the cell product includes hematopoietic stem cells (HSCs).

112. The method according to claim 111, wherein the method comprises, in order, the concentration step, the selection step, the standing step, the transdermal introduction step, and the sampling step.

113. The method according to claim 102, wherein the cell product comprises tumor-infiltrating lymphocytes (TILs).

114. The method according to claim 113, wherein the method comprises, in order, the tissue digestion step, the washing step, the activation step, the proliferation step, and the collection step.

115. A countercurrent centrifugal ertriation (CCE) module comprising a conical element having an inner and outer surface, fixedly attached to the distal end of a linear member having an inner and outer surface, The proximal end of the linear member is rotatably mounted to a pivot point to enable the linear member to be extended, retracted, and rotated, in a CCE module.

116. Enclosure and, Multiple instruments, each independently configured to perform one or more cell processing operations on a cartridge, A robot capable of moving the cartridge between each of the aforementioned multiple devices, A work cell equipped with the following features.

117. The work cell according to claim 116, wherein the enclosure includes an air filtration inlet configured to maintain ISO 7 or higher air quality within the internal zone of the work cell.

118. The work cell according to any one of claims 116 to 117, wherein the work cell is automated.

119. The work cell according to any one of claims 116 to 118, wherein the instrument is linked to the cartridge in order to perform a cell processing operation on the cartridge.

120. The work cell according to any one of claims 116 to 119, wherein the work cell comprises a processor, and the processor is configured to control the robot and the plurality of devices.

121. The work cell according to any one of claims 116 to 120, wherein the work cell is configured to receive two or more cartridges.

122. The work cell according to any one of claims 116 to 121, wherein the work cell includes the cartridge.

123. The work cell according to any one of claims 116 to 121, wherein the cartridge includes a plurality of modules.

124. The work cell according to any one of claims 116 to 123, wherein the cartridge includes a bioreactor module.

125. The work cell according to any one of claims 116 to 124, wherein the cartridge includes a cell selection module.

126. The work cell according to claim 125, wherein the cell selection module is a magnetically activated cell selection module.

127. The work cell according to any one of claims 116 to 126, wherein the cartridge includes a classification module.

128. The work cell according to claim 127, wherein the classification module is a fluorescence-activated cell classification (FACS) module.

129. The work cell according to any one of claims 116 to 128, wherein the cartridge includes an electroporation module.

130. The work cell according to any one of claims 116 to 129, wherein the cartridge includes a countercurrent centrifugal elutriation (CCE) module.

131. The work cell according to any one of claims 116 to 130, wherein the cartridge includes one or more sterile fluid transfer ports.

132. The work cell according to any one of claims 116 to 131, wherein the cartridge includes a liquid transfer bus fluidly coupled to each module.

133. The work cell according to any one of claims 116 to 132, wherein the cartridge includes a pump fluidly coupled to the liquid transfer bus.

134. The work cell according to any one of claims 116 to 133, wherein the work cell includes a pump actuator configured to work in conjunction with the pump.

135. The work cell according to any one of claims 116 to 134, wherein the work cell includes a bioreactor device.

136. The work cell according to claim 135, wherein the bioreactor device includes a plurality of slots for a cartridge.

137. The work cell according to any one of claims 116 to 124, wherein the work cell includes a cell selection device.

138. The work cell according to claim 137, wherein the cell selection device is a magnetically activated cell selection device.

139. The work cell according to any one of claims 116 to 138, wherein the work cell includes a sorting device.

140. The work cell according to claim 139, wherein the classification instrument is a fluorescence-activated cell classification (FACS) instrument.

141. The work cell according to any one of claims 116 to 140, wherein the work cell includes an electroporation apparatus.

142. The work cell according to any one of claims 116 to 141, wherein the work cell includes a countercurrent centrifugal ertriation (CCE) apparatus.

143. The work cell according to any one of claims 116 to 142, wherein the work cell includes a reagent storage container.

144. The work cell according to any one of claims 116 to 143, wherein the cartridge includes a bioreactor module and a selection module.

145. The work cell according to any one of claims 116 to 144, wherein the cartridge includes a bioreactor module and a CCE module.

146. The cartridge comprises a bioreactor module, a selection module, and a CCE module, as described in any one of claims 116 to 145.

147. The cartridge comprises a bioreactor module, a selection module, and an electroporation module, as described in any one of claims 116 to 146.

148. The cartridge comprises a bioreactor module, a selection module, a CCE module, and an electroporation module, according to any one of claims 116 to 147.

149. The work cell according to any one of claims 116 to 148, wherein the cartridge includes a second bioreactor module having an internal volume that is two times, five times, or ten times larger than the internal volume of the first bioreactor.

150. The work cell according to any one of claims 116 to 149, wherein the enclosure includes a feedthrough.

151. The work cell according to any one of claims 116 to 149, wherein the work cell performs automated production of cell products.

152. The system according to any one of claims 1 to 40, wherein the system comprises a plurality of bioreactor devices, each bioreactor device configured to receive a single cartridge.

153. The first side includes the first fluid tube, Including a second fluid tube, the second side is opposite to the first side, A cone is coupled between the first fluid tube and the second fluid tube, A rotor equipped with a rotor.

154. The rotor according to claim 153, wherein the cone is biconical.

155. The rotor according to claim 153, wherein the cone comprises a first cone including a first base and a second cone including a second base, the first base facing the second base.

156. The rotor according to any one of claims 153 to 155, wherein the rotor includes a magnetic portion.

157. The rotor according to any one of claims 153 to 156, wherein the rotor defines the axis of rotation.

158. The rotor according to claim 157, wherein at least a portion of the first fluid tube and at least a portion of the second fluid tube extend parallel to the axis of rotation.

159. The rotor according to any one of claims 153 to 157, wherein at least a portion of the first fluid tube and at least a portion of the second fluid tube are coaxial.

160. The rotor according to any one of claims 153 to 158, wherein the cone contains a volume of approximately 10 mL to approximately 40 mL.

161. The rotor according to any one of claims 153 to 160, wherein the cone includes a cone angle of approximately 30 to approximately 60 degrees.

162. The rotor according to any one of claims 153 to 161, wherein at least a portion of the rotor is optically transparent.

163. The rotor according to any one of claims 153 to 162, wherein the rotor includes an asymmetrical shape.

164. The rotor according to any one of claims 153 to 163, further comprising a first portion including the cone and a second portion including the paddle shape.

165. A cartridge for cell processing comprising a liquid transfer bus and multiple modules, Each module is fluidly coupled to the liquid transfer bus. The cartridge comprises a countercurrent centrifugal ertriation (CCE) module including a rotor according to any one of claims 153 to 164.

166. The first fluid tube, A first cone containing a first volume is connected to the first fluid tube, A second fluid tube coupled to the first cone, A second cone is connected to the second fluid tube and contains a second volume that is larger than the first volume, A third fluid tube connected to the second cone, A rotor equipped with a rotor.

167. The rotor according to claim 166, wherein the first cone includes a first bicone and the second cone includes a second bicone.

168. The rotor according to claim 167, wherein the first biconical shape includes a third cone including a first base and a fourth cone including a second base, the first base facing the second base, and the second biconical shape includes a fifth cone including a third base and a sixth cone including a fourth base, the third base facing the fourth base.

169. The rotor according to any one of claims 166 to 168, further comprising a magnetic portion.

170. The rotor according to any one of claims 166 to 169, wherein at least a portion of the rotor is optically transparent.

171. The rotor according to any one of claims 166 to 170, wherein the first fluid tube includes an inlet and the third fluid tube includes an outlet.

172. A cartridge comprising a housing containing a rotor configured to separate cells from a fluid, A device including a magnet configured to magnetically rotate the rotor in conjunction with the cartridge, A system for cell processing that includes the following features.

173. The system according to claim 172, wherein the cartridge is configured to move between multiple devices.

174. The system according to claim 172 or claim 173, further comprising an air gap between the housing and the magnet.

175. The system according to any one of claims 172 to 174, wherein the housing houses the rotor.

176. The system according to any one of claims 172 to 175, wherein the housing includes consumable components and the magnet includes durable components.

177. The system according to any one of claims 172 to 176, wherein the magnet is releasably coupled to the housing.

178. The system according to any one of claims 172 to 177, wherein the magnet is configured to move relative to the housing.

179. The system according to any one of claims 172 to 178, wherein the separated cells include a first size and a first density, and the unseparated cells of the fluid include a second size and a second density different from the first size and the first density.

180. A cartridge for cell processing comprising a liquid transfer bus and multiple modules, Each module is fluidly coupled to the liquid transfer bus. The cartridge comprises a countercurrent centrifugal ertriation (CCE) module including a rotor according to any one of claims 166 to 171.

181. Moving the rotor that defines the axis of rotation toward the magnet, The fluid is to flow through the rotor, While the fluid flows through the rotor, the rotor is magnetically rotated around the axis of rotation using the magnet, A countercurrent centrifugal ertriation (CCE) method, including [specific method].

182. Using an optical sensor, generate image data of one or more of the fluid and particles in the rotor, Selecting one or more of the rotor's rotational speed and the fluid's flow rate, at least partially based on the image data, The method according to claim 181, further comprising:

183. The method according to claim 181 or claim 182, further comprising illuminating one or more of the fluid and the cells using an illumination source.

184. The method according to any one of claims 181 to 183, further comprising moving the rotor toward away from the magnet.

185. Moving the rotor toward the light source and optical sensor, Moving the rotor away from the illumination source and the optical sensor, The method according to any one of claims 181 to 184, further comprising:

186. The method according to any one of claims 181 to 185, wherein moving the rotor includes using a robot to move the magnet forward and backward relative to the rotor.

187. The method according to any one of claims 181 to 186, wherein the rotation of the rotor includes a rotational speed of up to 6,000 RPM.

188. The method according to any one of claims 181 to 187, wherein the flow of the fluid includes a maximum flow rate of about 150 ml / min during the rotation of the rotor.

189. The process involves flowing the fluid containing the input cells into a flow cell, wherein the set of cells is labeled with a magnetically activated cell selection (MACS) reagent. The cell set is magnetically attracted to the magnetic array only for the duration of its residence, After the aforementioned residence time, the cell set is flowed out of the flow cell, A method for selecting magnetically activated cells, including the method described above.

190. The method according to claim 189, further comprising culturing the MACS reagent using the input cells in order to label the cell set with the MACS reagent.

191. The method according to claim 190, wherein the cultivation of the MACS reagent includes a temperature of about 1°C to about 10°C.

192. The method according to any one of claims 189 to 191, wherein flowing the cell set out of the flow cell includes flowing gas through the flow cell.

193. The method according to any one of claims 189 to 192, further comprising flowing the fluid, which does not contain the cell set, out of the flow cell after the residence time.

194. The method according to any one of claims 189 to 193, wherein the residence time is at least about 1 minute.

195. The method according to any one of claims 189 to 194, wherein the magnet array is located outside the flow cell.

196. The method according to any one of claims 189 to 195, further comprising moving the magnet array relative to the flow cell.

197. The method according to claim 196, wherein moving the magnet array includes moving the magnet array away from the flow cell to facilitate the outflow of the cell set from the flow cell.

198. The method according to any one of claims 189 to 197, wherein the vertical axis of the flow cell is perpendicular to the ground.

199. The method according to any one of claims 189 to 198, wherein the flow cell does not contain beads.

200. A flow cell containing an elongated cavity with a cavity height, A magnet array comprising a plurality of magnets, each of which is separated by a certain distance, The ratio of the cavity height to the separation distance is approximately 20:1 to approximately 1:

20. Magnetically activated cell selection (MACS) module.

201. The MACS module according to claim 200, wherein the flow cell includes a set of linear channels, the linear channels including a first channel parallel to a second channel and a third channel in fluid communication with each of the first channel and the second channel.

202. The MACS module according to claim 201, wherein the first channel includes a first cavity height, the second channel includes a second cavity height, and the ratio of the first cavity height to the second cavity height is approximately 1:1 to approximately 3:

7.

203. The MACS module according to claim 201, wherein the ratio of the length of the third channel to the diameter of the third channel is about 2:1 to about 6:

1.

204. A first fluid tube connected to the inlet and outlet of the flow cell, configured to receive the cell set from the flow cell, A second fluid tube connected to the inlet and outlet of the flow cell, configured to receive fluid from the flow cell that does not contain the cell set, A MACS module according to any one of claims 200 to 203, further comprising:

205. A cartridge for cell processing comprising a liquid transfer bus and multiple modules, Each module is fluidly coupled to the liquid transfer bus. The cartridge comprises a magnetically activated cell selection (MACS) module according to any one of claims 200 to 204.

206. A cartridge including a rotor configured for countercurrent centrifugation of cells in a fluid, A first magnet configured to magnetically rotate the rotor and to separate the cells from the fluid in the rotor, The cartridge further includes a flow cell configured to be in fluid communication with the rotor and to receive the cells from the rotor, A second magnet configured to magnetically separate the cells in the flow cell, A system for cell processing that includes the following features.

207. A lighting source configured to illuminate the aforementioned cells, An optical sensor configured to generate image data corresponding to the aforementioned cells, The system according to claim 206, further comprising the following:

208. The system according to claim 206 or claim 207, further comprising one or more of the following: an oxygen deficiency sensor, a leak sensor, an inertia sensor, a pressure sensor, and a bubble sensor.

209. The system according to any one of claims 206 to 208, further comprising one or more valves and pumps.

210. The system according to any one of claims 206 to 208, wherein the separated cells have a first size and a first density, and the unseparated cells of the fluid have a second size and a second density different from the first size and the first density.

211. A fluid tube configured to receive a first fluid containing cells and a second fluid, An electrode set coupled to the fluid tube, A pump connected to the aforementioned fluid pipe, A controller including a processor and memory, A first signal is generated in order to introduce the first fluid into the fluid pipe using the pump. A second signal is generated to introduce the second fluid into the fluid tube such that the second fluid separates the first fluid from the third fluid. Using the electrode set, an electroporation signal is generated to perform electroporation of the cells in the fluid tube. A controller configured as follows, An electroporation module equipped with the following features.

212. The electroporation module according to claim 211, wherein the second fluid includes a gas or an oil.

213. The electroporation module according to claim 211 or 212, wherein the controller is configured to generate a third signal for introducing the third fluid into the fluid tube, and the third fluid is separated from the first fluid by the second fluid.

214. A cartridge for cell processing comprising a liquid transfer bus and a plurality of modules, wherein each module is fluidically coupled to the liquid transfer bus, and the cartridge comprises an electroporation module according to any one of claims 211 to 213.

215. The process involves receiving a first fluid containing cells into a fluid tube, The second fluid is received into the fluid tube in such a way that the first fluid is separated from the third fluid, Applying an electroporation signal to the first fluid to perform electroporation of the cells, A method for performing electroporation of cells, including the following.

216. The method according to claim 215, further comprising receiving the third fluid, separated from the first fluid by the second fluid, into the fluid tube.

217. The method according to claim 215 or claim 216, wherein the first fluid is substantially static when the electroporation signal is applied.

218. The process involves receiving a first fluid containing cells into a fluid tube, Applying a resistance measurement signal to the first fluid using an electrode set, Measuring the resistance between the first fluid and the electrode set, Applying an electroporation signal to the first fluid based on the measured resistance, A method for performing electroporation of cells, including the following.

219. The method according to claim 218, further comprising receiving a second fluid containing a gas into the fluid tube before applying the electroporation signal to the fluid, wherein the first fluid is separated from the third fluid by the second fluid.

220. An enclosure including a bottom, top, and at least one side wall, A gas-permeable membrane bonded to one or more of the bottom and side walls of the enclosure, A bioreactor equipped with [a specific feature / equipment].

221. The bioreactor according to claim 220, wherein the enclosure includes one or more nested surfaces curved about the longitudinal axis of the enclosure.

222. The bioreactor according to claim 220 or claim 221, wherein one or more nesting surfaces include a set of concentric annular bodies.

223. The bioreactor according to any one of claims 220 to 222, wherein the enclosure includes an annular shape.

224. The bioreactor according to any one of claims 220 to 223, wherein the enclosure includes a first chamber having a first volume and a second chamber having a second volume, the first chamber being separated from the second chamber, and the first volume being smaller than the second volume.

225. The bioreactor according to any one of claims 220 to 224, wherein the enclosure includes a column extending along the longitudinal axis of the enclosure.

226. The bioreactor according to any one of claims 220 to 225, further comprising a cavity between the enclosure and the gas-permeable membrane.

227. The bioreactor according to any one of claims 220 to 226, wherein the gas-permeable membrane extends along the bottom and side walls of the enclosure.

228. The bioreactor according to any one of claims 220 to 227, wherein the outer surface of the gas permeable membrane includes one or more protrusions.

229. The bioreactor according to any one of claims 220 to 228, wherein the bottom of the gas-permeable membrane includes an angle of about 3 to about 10 degrees with respect to the bottom of the enclosure.

230. The bioreactor according to any one of claims 220 to 229, wherein the gas permeable membrane includes a curved surface.

231. The bioreactor according to any one of claims 220 to 230, wherein the gas-permeable membrane includes a set of patterned curved surfaces.

232. The bioreactor according to claim 231, wherein the set of patterned curved surfaces has a radius of curvature of approximately 50 mm to approximately 500 mm.

233. A cartridge for cell processing comprising a liquid transfer bus and multiple modules, Each module is fluidly coupled to the liquid transfer bus. The cartridge comprises a bioreactor module, The bioreactor module is a cartridge comprising at least one bioreactor as described in any one of claims 220 to 232.

234. The bioreactor device further comprises a device configured to work in conjunction with the aforementioned cartridge, The bioreactor apparatus comprises the cartridge according to claim 233, which is configured to be coupled to the bioreactor and to include a stirrer configured to agitate a cell culture medium containing cells. A system for processing cells.

235. The system according to claim 234, wherein the bioreactor is configured to be coupled to a liquid transfer bus, and further comprises a fluid connector having a foldable side wall.

236. The system according to claim 234 or claim 235, further comprising a temperature controller coupled to the bioreactor.

237. The system according to any one of claims 234 to 236, further comprising a gas regulator coupled to the bioreactor.

238. A first connector including a first proximal end and a first distal end including a first port, configured to be coupled to a first fluid device, A second connector comprising a second proximal end configured to connect to a second fluid device and a second distal end including a second port configured to connect to the first port, The first distal end includes a first lumen, and the second distal end includes a second lumen. One of the first valve and the second valve is configured to move in parallel within the first lumen and the second lumen. Fluid connector.

239. The fluid connector according to claim 238, wherein the first valve and the second valve are configured to transition from a closed configuration to an open configuration only when the first valve is coupled with the second valve.

240. The fluid connector according to claim 238 or 239, wherein the first port and the second port are configured to transition between an open configuration and a closed configuration.

241. The fluid connector according to any one of claims 238 to 240, wherein the first connector includes a first port actuator, and / or the second connector includes a second port actuator.

242. The fluid connector according to any one of claims 238 to 241, wherein the second port coupled to the first port defines a chamber.

243. The fluid connector according to claim 242, wherein one or more of the first connector and the second connector include a sterilizer port configured to be coupled to a sterilizer source, and the sterilizer port is configured to be in fluid communication with the first distal end and the second distal end when the second port is coupled to the first port.

244. The fluid connector according to claim 243, wherein the chamber is configured to receive one or more of a fluid and a sterilizer from the sterilizer port.

245. The fluid connector according to any one of claims 238 to 244, wherein the sterilizer port is configured to receive the sterilizer so that the sterilizer sterilizes the first connector and the second connector.

246. The fluid connector according to any one of claims 238 to 245, wherein the first connector includes a first valve, and the second connector includes a second valve configured to be coupled to the first valve.

247. A fluid connector according to any one of claims 238 to 246, wherein the first seal includes the first port coupled to the second port, and the second seal includes the first valve coupled to the second valve.

248. The fluid connector according to any one of claims 244 to 247, wherein the sterilizer comprises one or more of vaporized hydrogen peroxide and ethylene oxide.

249. A fluid connector according to any one of claims 238 to 248, further comprising one or more robot engagement mechanisms.

250. The fluid connector according to any one of claims 238 to 249, wherein the first connector includes a first alignment mechanism, and the second connector includes a second alignment mechanism configured to be coupled to the first alignment mechanism in a predetermined axial rotation configuration.

251. A fluid connector according to any one of claims 238 to 250, wherein one or more of the first fluid device and the second fluid device include a fixture.

252. A robot configured to operate the aforementioned fluid connector, A controller comprising memory and a processor, coupled to the robot, configured to generate a first port signal for coupling the first port to the second port using the robot arm, The system according to any one of claims 234 to 237, further comprising:

253. The system according to claim 252, wherein the controller is configured to generate a first valve signal for moving the first valve in parallel with respect to the second valve using the robot arm, and further generates a second valve signal for moving the first valve and the second valve to the open configuration.

254. The system according to claim 252 or 253, wherein the controller is configured to generate a second port signal for uncoupling the first port from the second port, and the sterility of the fluid connector is maintained before coupling the first port to the second port and after uncoupling the first port from the second port.

255. The system according to any one of claims 252 to 254, further comprising a fluid pump coupled to the sterilizer source, wherein the controller is configured to generate a first fluid pump signal for circulating the fluid into the chamber via the sterilizer port.

256. The system according to claim 255, wherein the controller is configured to generate a second fluid pump signal for circulating the sterilizer into the chamber via the sterilizer port in order to sterilize at least the chamber.

257. The system according to claim 256, wherein the controller is configured to generate a third fluid pump signal for removing the sterilizer from the chamber.

258. The system according to any one of claims 255 to 257, wherein the controller is configured to generate a thermal sterilization signal for thermally sterilizing the fluid connector.

259. The system according to any one of claims 255 to 258, wherein the controller is configured to generate a radiation sterilization signal for sterilizing the fluid connector using radiation.

260. The system according to any one of claims 252 to 259, wherein the robot is configured to connect fluid connectors between at least two of the plurality of devices and the cartridge.

261. The fluid connector further comprises the fluid connector according to any one of claims 238 to 251. A controller comprising memory and a processor, coupled to the robot, Using the robot arm, a port signal is generated to connect the first port to the second port. Using the robot arm, a first valve signal is generated to move the first valve in parallel with respect to the second valve. A second valve signal is generated to move the first valve and the second valve to the open configuration. A controller configured in such a way, The system according to any one of claims 252 to 260, further comprising:

262. A non-temporary computer-readable medium for converting user-defined cell processing operations into cell processing steps executed by an automated cell processing system, which includes stored instructions. If the aforementioned instruction is executed on the processor, A step of receiving an ordered input list of cell processing operations, A step of running a transformation model on the ordered input list to generate an ordered output list of cell processing steps that can be performed by the system, A non-temporary computer-readable medium that performs [something].

263. The aforementioned ordered output list is: Each controls the robot to move one or more cartridges containing cell products between the instruments, and To control the instrument to perform a cell processing step for each cell product, A non-temporary computer-readable medium according to claim 262, which can be executed by the aforementioned system.

264. The above step further, A non-temporary computer-readable medium according to claim 262 or claim 263, comprising receiving one or more sets of cell processing parameters, each set of the cell processing parameters being associated with one of the cell processing operations, and each set of the cell processing parameters defining the characteristics of the cell processing step performed by the instrument in the cell processing step.

265. The non-temporary computer-readable medium according to any one of claims 262 to 264, wherein the conversion model includes constraints on the ordered output list determined by the configuration of the automated cell processing system.

266. The non-temporary computer-readable medium according to claim 264, wherein the constraints include information relating to the configuration of the automated cell processing system.

267. The aforementioned constraints Type and / or number of appliances, The type and / or number of modules on the cartridge, The type and number of storage compartments on the cartridge, The type and / or number of sterile fluid transfer ports on the cartridge, The number and location of fluid paths between the module, the storage unit, and the sterile liquid transfer port on the cartridge, A non-temporary computer-readable medium according to any one of claims 264 to 266, comprising one or more of the above.

268. The above step further, The automated cell processing system receives a set of two or more ordered input lists of cell processing operations, in order to be executed in two or more cartridges, This includes running the transformation model on the ordered input list to generate the ordered output list of the cell processing step, The aforementioned ordered output list is: To control the robot so that each of the two or more cartridges containing cell products moves between the instruments, To control the instrument to perform a cell processing step for each cell product in each cartridge, A non-temporary computer-readable medium according to any one of claims 262 to 267, which can be executed by the aforementioned system.

269. An automated cell processing system comprising a non-temporary computer-readable medium according to any one of claims 262 to 268.

270. A computer-based method for converting user-defined cell processing operations into cell processing steps executed by the processor of an automated cell processing system, Receiving an ordered list of cell processing operations, The transformation model is executed on the ordered input list to generate an ordered output list of cell processing steps that can be performed by the system, A method performed by a computer, including the above.

271. Each robot controls the robot to move one or more cartridges containing cell products between the instruments, Controlling the instrument to perform a cell processing step for each cell product, The method according to claim 270, further comprising:

272. The method according to claim 270 or claim 271, comprising receiving one or more sets of cell processing parameters, each set of the cell processing parameters being associated with one of the cell processing operations, and each set of the cell processing parameters defining the characteristics of the cell processing step performed by the instrument in the cell processing step.

273. The method according to any one of claims 270 to 272, wherein the conversion model includes constraints on the ordered output list determined by the configuration of the automated cell processing system.

274. The method according to claim 272, wherein the constraints include information relating to the configuration of the automated cell processing system.

275. The aforementioned constraints Type and / or number of appliances, The type and / or number of modules on the cartridge, The type and number of storage compartments on the cartridge, The type and / or number of sterile fluid transfer ports on the cartridge, The number and location of fluid paths between the module, the storage unit, and the sterile liquid transfer port on the cartridge, The method according to any one of claims 272 to 274, comprising one or more of the above.

276. The automated cell processing system receives a set of two or more ordered input lists of cell processing operations, in order to be executed in two or more cartridges, The transformation model is executed on the set of ordered input lists to generate the ordered output list of the cell processing step, Each controls the robot to move the two or more cartridges containing cell products between the instruments, Controlling the instrument to perform a cell processing step for each cell product in each cartridge, The method according to any one of claims 270 to 275, further comprising:

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