Point-of-care systems for automated cell processing

Abstract

The present disclosure generally relates to systems and methods for processing cells, and kits for use with such methods and processes. The systems include multiple subsystems, at least one of which is removably coupled to a frame of the system. In one embodiment, the systems are sterile and configured for use in non-sterile locations. In one embodiment, the systems produce processed cells in about 5 to about 7 hours.

Description

Technical Field

[0001] Cross - reference to related applications This application claims the priority and benefit of U.S. Provisional Patent Application No. 63 / 029,230, filed on May 22, 2020, the entire content of which is incorporated herein by reference in its entirety for all purposes.

[0002] The present disclosure generally relates to systems and methods for treating cells, and kits for use with such systems and methods for treating cells.

Background Art

[0003] Cell - based therapeutic agents, such as cell therapies, are designed to boost the immune response and are becoming an increasingly common treatment option for several diseases, such as cancer. In particular, cell therapies, such as cell - based therapies for cancer treatment, have several advantages over conventional treatment options such as chemotherapy and radiation, and as a result, cell therapies are becoming more widely used. These advantages can, in some cases, include a shorter treatment time compared to chemotherapy, since only a single injection may be required to achieve the desired treatment result. An additional advantage can be a more rapid recovery time compared to recovery from chemotherapy. A further advantage of cell - based therapeutic agents compared to chemotherapy is that cell - based therapeutic agents can be a targeted treatment of specific molecular targets associated with cancer, in contrast to standard chemotherapy, which acts on all rapidly dividing normal and cancerous cells.

[0004] However, despite the benefits of currently approved cell therapies, such as cell-based therapies, limitations in the manufacturing process for these cell therapies remain a significant bottleneck in the field. Furthermore, as will be discussed later, other manufacturing processes can take days to weeks to produce the desired cell therapy. Moreover, these other manufacturing processes generally involve the use of several different instruments, each requiring its own configuration and cleaning, significantly adding to the time required to produce the final product. Furthermore, other methods for producing cell therapies require several different instruments and sterile environments, thus necessitating the required sterile conditions, such as a large physical space with multiple cleanrooms, to carry out the manufacturing process, thereby adding significant costs to the manufacturing process.

[0005] Therefore, there is a need in the field for systems and methods for processing cells, such as for producing cell-based therapeutics, and such systems and methods represent an improvement over current systems and methods. [Overview of the Initiative] [Means for solving the problem]

[0006] This disclosure generally relates to a system for processing cells, the system comprising: a suspension preparation subsystem comprising a delivery medium inlet, a cell isolation device configured to isolate cells, and a cell suspension device configured to suspend the isolated cells in a delivery medium, thereby creating a cell suspension; a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, the cell deformation subsystem comprising a cell suspension inlet, and one or more cell deformation contractions configured to induce a perturbation in the cell membrane of cells to allow a payload to enter the cells, thereby creating a cell suspension containing the manipulated cells; a dilution subsystem fluidly communicating with the cell deformation subsystem, the dilution subsystem comprising a cell suspension inlet, a buffer inlet, and a container configured to receive a cell suspension containing the manipulated cells and a fluid or dry reagent to mix with the cell suspension containing the manipulated cells to create a diluted cell suspension; and an incubation subsystem fluidly communicating with the dilution subsystem. The system comprises an incubation subsystem comprising a stem comprising a diluted cell suspension inlet, a container configured to receive the diluted cell suspension, and a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension; a cell washing subsystem fluidly communicating with the incubation subsystem, wherein the cell washing subsystem comprises an incubated cell suspension inlet, a storage medium inlet, and a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension in the container, thereby suspending the cells in the storage medium; and a container filling subsystem fluidly communicating with the incubation subsystem, wherein the container filling subsystem comprises an inlet configured to receive cells suspended in the storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump the cells suspended in the storage medium into one or more containers, wherein at least one of the subsystems isIt is detachably coupled to the system frame.

[0007] In some embodiments, the system is sterile and configured for use in non-sterile locations. In some embodiments, the system produces processed cells in about 5 to 7 hours. In some embodiments, the cells include erythrocytes (RBCs). In some embodiments, the cells include RBC-derived vesicles, e.g., activated antigen carriers (AACs), tolerant antigen carriers (TACs). In some embodiments, the cells include peripheral blood mononuclear cells (PBMCs). In some embodiments, the cells include activated antigen carriers (APCs). In some embodiments, the cells include T cells, B cells, dendritic cells, monocytes, macrophages, eosinophils, basophils, natural killer (NK) cells, natural killer T (NKT) cells, mast cells, or neutrophils. In some embodiments, the cell suspension inlet and buffer inlet of the dilution subsystem are the same inlet. In some embodiments, the buffer inlet is used to deliver fluid to the dilution subsystem. In some embodiments, the buffer inlet is used to deliver dry reagents to the dilution subsystem. In some embodiments, the payload includes one or more reprogramming factors. In some embodiments, the payload includes one or more nucleic acids. In some embodiments, the payload includes one or more differentiation factors. In some embodiments, the payload includes one or more neuronal reprogramming factors. In some embodiments, one or more subsystems include one or more components configured to be removably bound to the system. In some embodiments, the suspension preparation subsystem further includes a wash medium inlet. In some embodiments, the suspension preparation subsystem further includes a dilution medium inlet. In some embodiments, the suspension preparation subsystem further includes an elutriation system configured to perform a cell isolation operation on cells. In some embodiments, the suspension preparation subsystem further includes a leukocyte removal filter system configured to perform a cell isolation operation on cells. In some embodiments, the suspension preparation subsystem further includes a tangential flow filtration system configured to perform a buffer exchange operation. In some embodiments, the suspension preparation subsystem further includes at least one outlet configured to be bound to at least one container for receiving cells suspended in a delivery medium.In some embodiments, the container is a bag having at least one inlet and at least one outlet. In some embodiments, the suspension preparation subsystem includes a weighing system configured to weigh the container. In some embodiments, the weighing system includes a tension load cell. In some embodiments, the weighing system includes a compression load cell. In some embodiments, the weighing system includes a straight bar load cell. In some embodiments, the suspension preparation subsystem further includes an agitation system including a platform that contacts the container and is configured to rock the container up and down. In some embodiments, the suspension preparation subsystem further includes an agitation system including a plate that contacts the container and is configured to move in and out of contact with the container. In some embodiments, the suspension preparation subsystem further includes one or more pumps configured to move fluid within the subsystem or between fluid-connected subsystems. In some embodiments, the suspension preparation subsystem can remove plasma. In some embodiments, the suspension preparation subsystem can remove serum.

[0008] In some embodiments, the cell deformation subsystem further includes a pressurizing system configured to generate pressure to pass a cell suspension through one or more cell deformation contractions. In some embodiments, the cell deformation subsystem further includes a preparation vessel configured to pass a cell suspension through one or more cell deformation contractions. In some embodiments, the preparation vessel is a rigid reservoir subassembly. In some embodiments, the cell deformation subsystem further includes a temperature control system comprising a heated plate configured to control the temperature of the cell suspension. In some embodiments, the cell deformation subsystem further includes at least one outlet configured to be coupled to at least one container for receiving a cell suspension or a cell suspension containing manipulated cells. In some embodiments, the container is a bag comprising at least one inlet and at least one outlet. In some embodiments, the cell deformation subsystem further includes an agitation system comprising a platform in contact with at least one container configured to oscillate the container up and down. In some embodiments, the agitation system comprising a platform in contact with at least one container is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension or a cell suspension containing manipulated cells. In some embodiments, the cell deformation subsystem further includes a stirring system comprising a plate configured to contact and move in and out of a container. In some embodiments, the dilution subsystem includes a weighing system for weighing the container. In some embodiments, the weighing system for weighing the container includes a tensile load cell. In some embodiments, the weighing system for weighing the container includes a compression load cell. In some embodiments, the weighing system for weighing the container includes a straight bar load cell. In some embodiments, the dilution subsystem includes a weighing system for measuring the amount of buffer added to cells in a cell suspension containing manipulated cells. In some embodiments, the weighing system for measuring the amount of buffer includes a tensile load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a compression load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a straight bar load cell.In some embodiments, the dilution subsystem includes at least one outlet configured to be coupled to a container configured to receive a cell suspension containing manipulated cells. In some embodiments, the container is a bag including at least one inlet and at least one outlet. In some embodiments, the dilution subsystem further includes a stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the first stirring system including the platform in contact with the container is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension containing manipulated cells or the diluted cell suspension. In some embodiments, the dilution subsystem further includes a first stirring system including a plate configured to be in contact with and move in and out of the container while in contact with the container. In some embodiments, the dilution subsystem further includes an illumination system configured to illuminate the cell suspension containing manipulated cells and the diluted cell suspension. In some embodiments, the dilution subsystem further includes a second stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, a second stirring system is configured to agitate the cell suspension containing the manipulated cells or the diluted cell suspension by oscillating the container up and down to promote homogeneity of the cell suspension containing the manipulated cells or the diluted cell suspension. In some embodiments, the dilution subsystem further includes a second stirring system comprising a plate configured to contact and move in and out of the container while in contact with the container. In some embodiments, the dilution subsystem comprises at least one pump configured to move fluid between fluid-connected subsystems.

[0009] In some embodiments, the incubation subsystem includes a temperature control device configured to regulate the temperature of a diluted cell suspension. In some embodiments, the temperature control device is a heated plate. In some embodiments, the incubation subsystem includes a weighing system configured to measure the amount of buffer added to the cells in the diluted cell suspension. In some embodiments, the weighing system includes a tensile load cell. In some embodiments, the weighing system includes a compression load cell. In some embodiments, the weighing system includes a straight bar load cell. In some embodiments, the incubation subsystem includes a first stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the first stirring system including the platform in contact with the container is configured to oscillate the container up and down to agitate the diluted cell suspension and promote homogeneity of the diluted or incubated cell suspension. In some embodiments, the incubation subsystem includes a second stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, a first agitation system including a platform in contact with a container is configured to agitate the cell suspension by oscillating the container up and down to promote homogeneity of the diluted or incubated cell suspension. In some embodiments, the incubation subsystem includes at least one pump configured to move fluid between fluid-connected subsystems. In some embodiments, the cell washing subsystem includes a tangential flow filtration system configured to perform a buffer exchange operation. In some embodiments, the cell washing subsystem includes a metering system configured to measure the amount of buffer added to the cells during the buffer exchange operation. In some embodiments, the metering system is a tensile load cell. In some embodiments, the metering system is a compression load cell. In some embodiments, the metering system is a straight bar load cell. In some embodiments, the cell washing subsystem includes at least one outlet configured to be coupled to a container.In some embodiments, the container is a bag having at least one inlet and at least one outlet. In some embodiments, the cell washing subsystem includes an agitation system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the agitation system including the platform in contact with the container is configured to oscillate the container up and down to agitate cells suspended in an incubated cell suspension or storage medium, thereby promoting homogeneity of cells suspended in a diluted cell suspension or storage medium. In some embodiments, the cell washing subsystem includes an illumination system configured to illuminate cells suspended in a diluted cell suspension or storage medium. In some embodiments, the storage medium is a cryoprotection medium. In some embodiments, the container filling subsystem includes a weighing system configured to measure the amount of cells suspended in a storage medium added to one or more containers. In some embodiments, the weighing system includes a tensile load cell. In some embodiments, the weighing system includes a compression load cell. In some embodiments, the weighing system includes a straight bar load cell. In some embodiments, the container filling subsystem includes an agitation system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, a stirring system including a platform in contact with a container is configured to agitate the container up and down to agitate the cells suspended in the storage medium, thereby promoting homogeneity of the cells suspended in the storage medium in one or more containers. In some embodiments, the container filling subsystem includes an illumination system configured to illuminate the cells suspended in the storage medium in one or more containers. In some embodiments, the container filling subsystem includes one or more outlets configured to be coupled to one or more containers. In some embodiments, one or more containers of the cell washing subsystem include one or more bags including at least one inlet and at least one outlet. In some embodiments, the container filling subsystem includes at least one pump configured to move fluid within the subsystem or between fluid-connected subsystems.In some embodiments, the system includes one or more pumps configured to pump fluid between two or more subsystems. In some embodiments, the system is used in a non-sterile environment. In some embodiments, the system is an automated system for processing cells. In some embodiments, the system processes cells for cell therapies such as cell-based therapeutics. In some embodiments, the suspension preparation subsystem further includes one or more of the following: cell aggregate filters, leukocyte removal filters, piping, pipe fittings, connectors, clamps, sampling bulbs, carboys, and air filters. In some embodiments, the cell deformation subsystem further includes one or more of the following: rigid sample containers, cell aggregate filters, rigid preparation containers, one or more microfluidic tip cartridges, one or more microfluidic tips, piping, pipe fittings, connectors, clamps, air filters, and barrel filters. In some embodiments, the dilution subsystem further includes one or more of the following: cell aggregate filters, piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys. In some embodiments, the incubation subsystem further includes one or more of the following: cell aggregate filters, piping, pipe fittings, connectors, and clamps. In some embodiments, the cell washing subsystem further includes one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, sampling spheres, air filters, and carboys. In some embodiments, the container filling subsystem further includes one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, and sampling spheres.

[0010] Furthermore, the present disclosure generally relates to a method for processing cells, wherein the method is carried out by a system comprising one or more of the following subsystems: a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method comprises: i. in the suspension preparation subsystem: (1) receiving cells from a container; (2) performing a cell isolation operation on the cells to thereby produce isolated cells; (3) receiving a delivery medium through a delivery medium inlet; and (4) producing a cell suspension by suspending the isolated cells in the delivery medium to thereby produce a cell suspension; and ii. in the cell deformation subsystem: (1) receiving a flow of cell suspension from the suspension preparation subsystem; and (2) flowing the cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membranes of the cells to allow the entry of a payload into the cells, thereby producing a cell suspension containing the manipulated cells; ii. Dilution subsystem: (1) receiving a flow of cell suspension containing manipulated cells from the cell deformation subsystem; (2) receiving a fluid or dry reagent via a buffer inlet; (3) producing a diluted cell suspension by mixing the cell suspension containing manipulated cells with the fluid or dry reagent; iv. Incubation subsystem: (1) receiving a flow of diluted cell suspension from the dilution subsystem; (2) adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension; v. Cell washing subsystem: (1) receiving a flow of incubated cell suspension from the incubation subsystem; (2) performing a buffer exchange operation on the cells to suspend them in a storage medium; vi. Container filling subsystem: (1) receiving a flow of cells suspended in a storage medium from the cell washing subsystem; (2) introducing the cells suspended in the storage medium into one or more containers.

[0011] In some embodiments, the cells include red blood cells (RBCs). In some embodiments, the cells include RBC-derived vesicles, e.g., activated antigen carriers (AACs), tolerant antigen carriers (TACs). In some embodiments, the cells include peripheral blood mononuclear cells (PBMCs). In some embodiments, the cells include activated antigen carriers (APCs). In some embodiments, the cells include T cells, B cells, dendritic cells, monocytes, macrophages, eosinophils, basophils, natural killer (NK) cells, natural killer T (NKT) cells, mast cells, or neutrophils. In some embodiments, the method removes plasma. In some embodiments, the method removes serum. In some embodiments, the method is carried out for about 5 to about 7 hours. In some embodiments, the method produces a cell therapy, e.g., a cell-based therapeutic agent. In some embodiments, the payload includes one or more reprogramming factors. In some embodiments, the payload includes one or more nucleic acids. In some embodiments, the payload includes one or more differentiation factors. In some embodiments, the payload comprises one or more neuronal reprogramming factors. In some embodiments, the method is an automated method for processing cells. In some embodiments, the method is carried out by a sterile system in a non-sterile environment.

[0012] Furthermore, the present disclosure relates generally to a kit for use in a system for processing cells, the kit comprising: i. a first kit comprising detachable components configured to be detachably bonded to a frame of a suspension preparation subsystem of the system, wherein a first set of detachable components comprises (1) a cell isolation device and / or (2) a cell suspension device; ii. a second kit comprising detachable components configured to be detachably bonded to a frame of a cell deformation subsystem of the system, wherein a second set of detachable components comprises one or more microfluidic chips comprising one or more cell deformation contractions that can compel cells to undergo perturbations of the cell membrane; and iii. a third kit comprising detachable components configured to be detachably bonded to a frame of a dilution subsystem of the system, wherein a The set of 3 comprises one or more of the following: a third kit comprising at least one cell aggregate filter; a fourth kit comprising detachably connectable components configured to be detachably connectable to the frame of the incubation subsystem of the system, wherein the fourth set of detachably connectable components comprises a fourth kit comprising at least one cell aggregate filter; a fifth kit comprising detachably connectable components configured to be detachably connectable to the frame of the cell washing subsystem of the system, wherein the fifth set of detachably connectable components comprises a second tangential flow filtration membrane assembly; and a sixth kit comprising detachably connectable components configured to be detachably connectable to the frame of the container filling subsystem of the system, wherein the sixth set of detachably connectable components comprises at least one cell aggregate filter.

[0013] In some embodiments, one or more components of the kit are configured to be fluidly connected to one or more components of the corresponding subsystem of the system. In some embodiments, the cell isolation device includes at least one elutriation device. In some embodiments, the cell isolation device includes at least one leukocyte removal filter. In some embodiments, the first kit includes the cell isolation device. In some embodiments, the cell suspension device is a tangential flow filtration membrane assembly. In some embodiments, i. one or more detachable components of a second kit of detachable components are configured to be fluidly connected to one or more detachable components of a first kit of detachable components; ii. one or more detachable components of a third kit of detachable components are configured to be fluidly connected to one or more detachable components of a second kit of detachable components; iii. one or more detachable components of a fourth kit of detachable components are configured to be fluidly connected to one or more detachable components of a third kit of detachable components; iv. one or more detachable components of a fifth kit of detachable components are configured to be fluidly connected to one or more detachable components of a fourth kit of detachable components; and / or v. one or more detachable components of a sixth kit of detachable components are configured to be fluidly connected to one or more detachable components of a fifth kit of detachable components. In some embodiments, the first kit of removablely connectable components includes a cell aggregate filter. In some embodiments, the first kit of removablely connectable components includes a leukocyte removal filter. In some embodiments, the first kit of removablely connectable components includes a container.In some embodiments, a first kit of detachably connectable components includes a tangential flow filtration filter assembly. In some embodiments, a first kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, carboys, air filters, and tangential flow filtration filter assemblies. In some embodiments, a second kit of detachably connectable components includes a rigid sample container. In some embodiments, a second kit of detachably connectable components includes a cell aggregate filter. In some embodiments, a second kit of detachably connectable components includes a preparation container. In some embodiments, a second kit of detachably connectable components includes one or more microfluidic chips. In some embodiments, a second kit of detachably connectable components includes one or more microfluidic chip cartridges. In some embodiments, a second kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, containers, bags, air filters, and barrel filters. In some embodiments, a third kit of detachably connectable components includes a container. In some embodiments, a third kit of detachably connectable components includes a cell aggregate filter. In some embodiments, a third kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys. In some embodiments, a fourth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, a fourth kit of detachably connectable components includes one or more containers. In some embodiments, a fourth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, and clamps. In some embodiments, a fifth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, a fifth kit of detachably connectable components includes containers. In some embodiments, a fifth kit of detachably connectable components includes a tangential flow filtration filter assembly.In some embodiments, a fifth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, air filters, and carboys. In some embodiments, a sixth kit of detachably connectable components includes a container. In some embodiments, the container is a cryopreservation bag. In some embodiments, a sixth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, a sixth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, and sampling bulbs. In some embodiments, the kit includes the first, second, third, fourth, fifth, and sixth kits. In some embodiments, the kit is packaged in an accordion tray package. In some embodiments, the kit is packaged as a rollable sheet. In some embodiments, each kit is packaged separately. In some embodiments, at least two kits are packaged together. In some embodiments, the kit is sterile.

[0014] Furthermore, the present disclosure generally relates to a system for processing cells, the system comprising: a suspension preparation subsystem comprising a cell isolation device, a tangential flow filtration membrane assembly, a cell aggregate filter, a leukocyte removal filter, and one or more containers; a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, wherein the cell deformation subsystem comprises one or more microfluidic chips, a rigid sample container, and a cell aggregate filter; and a dilution subsystem fluidly communicating with the cell deformation subsystem, wherein the dilution subsystem comprises a container and a cell aggregate filter. A fluid-communicating incubation subsystem comprising an incubation subsystem including a cell aggregate filter and a container; a cell washing subsystem fluid-communicating with the incubation subsystem, comprising a cell washing subsystem including a tangential flow filtration membrane assembly, a cell aggregate filter and a container; and a container filling subsystem fluid-communicating with the incubation subsystem, comprising a container and a cell aggregate filter, wherein subsystems i. to vi. are detachably coupled to the system frame.

[0015] Furthermore, the present disclosure relates in general to a system for processing cells, the system comprising: a suspension preparation subsystem comprising a delivery medium inlet, a cell isolation device configured to isolate cells, and a cell suspension device configured to suspend the isolated cells in a delivery medium, thereby creating a cell suspension; a payload entry subsystem fluidly communicating with the suspension preparation subsystem, the payload entry subsystem comprising a cell suspension inlet, a payload entry component that causes a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells; a dilution subsystem fluidly communicating with the cell deformation subsystem, the dilution subsystem comprising a cell suspension inlet, a buffer inlet, and a container configured to receive a cell suspension containing manipulated cells and a buffer that mixes with the cell suspension containing manipulated cells to create a diluted cell suspension; and an incubation subsystem fluidly communicating with the dilution subsystem, the incubation subsystem comprising a diluted cell suspension inlet, The system comprises an incubation subsystem including a container configured to receive a diluted cell suspension and a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension; a cell washing subsystem fluidly communicating with the incubation subsystem, wherein the cell washing subsystem includes an inlet for an incubated cell suspension, an inlet for a storage medium, and a container configured to receive an incubated cell suspension and a storage medium to be mixed with the incubated cell suspension in the container, thereby suspending the cells in the storage medium; and a container filling subsystem fluidly communicating with the incubation subsystem, wherein the container filling subsystem includes an inlet configured to receive cells suspended in the storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump the cells suspended in the storage medium into one or more containers, wherein subsystems i. to vi. are detachably connectable to a frame.In some embodiments, the payload entry component includes an electroporation device. In some embodiments, the payload entry component includes one or more cell deformation contractions through which a cell suspension flows, the cell deformation contractions configured to cause a perturbation at the cell membrane of the cell, thereby enabling the entry of the payload into the cell.

[0016] Furthermore, the present disclosure relates in general to a system for processing cells, the system comprising a suspension preparation subsystem, a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, a dilution subsystem fluidly communicating with the cell deformation subsystem, an incubation subsystem fluidly communicating with the dilution subsystem, a cell washing subsystem fluidly communicating with the incubation subsystem, and a container filling subsystem fluidly communicating with the incubation subsystem, wherein at least one of the subsystems is detachably coupled to the frame of the system.

[0017] Furthermore, the present disclosure relates in general to a system for processing cells, the system comprising a suspension preparation subsystem, a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, a temperature control subsystem fluidly communicating with the cell deformation subsystem, a cell washing subsystem fluidly communicating with the temperature control subsystem, and a container filling subsystem fluidly communicating with the incubation subsystem, wherein at least one of the subsystems is detachably coupled to the system frame.

[0018] Furthermore, the present disclosure relates in general to a system for processing cells, the system comprising: a suspension preparation subsystem comprising a delivery medium inlet, a cell suspension device configured to suspend cells in a delivery medium, thereby creating a cell suspension; a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, the cell deformation subsystem comprising a cell suspension inlet, one or more cell deformation contractions configured to induce a perturbation in the cell membrane of cells, thereby enabling the entry of a payload into the cells, thereby creating a cell suspension containing manipulated cells; a dilution subsystem fluidly communicating with the cell deformation subsystem, the dilution subsystem comprising a cell suspension inlet, a buffer inlet, a container configured to receive a cell suspension containing manipulated cells, and a fluid or dry reagent to be mixed with the cell suspension containing manipulated cells to create a diluted cell suspension; and an incubation subsystem fluidly communicating with the dilution subsystem, the incubation subsystem comprising a diluted cell suspension inlet, diluted cells The system comprises an incubation subsystem including a container configured to receive a suspension and a plate configured to regulate the temperature of a diluted cell suspension in the container to create an incubated cell suspension; a cell washing subsystem fluidly communicating with the incubation subsystem, wherein the cell washing subsystem includes an inlet for an incubated cell suspension, an inlet for a storage medium, and a container configured to receive an incubated cell suspension and a storage medium to be mixed with the incubated cell suspension in the container, thereby suspending the cells in the storage medium; and a container filling subsystem fluidly communicating with the incubation subsystem, wherein the container filling subsystem includes an inlet configured to receive cells suspended in the storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump cells suspended in the storage medium into one or more containers, wherein at least one of the subsystems is detachably coupled to the system frame.

[0019] Furthermore, the present disclosure relates in general to a system for processing cells, the system comprising: a suspension preparation subsystem comprising a delivery medium inlet, a cell isolation device configured to isolate cells, and a cell suspension device configured to suspend the isolated cells in the delivery medium, thereby creating a cell suspension; a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, the cell deformation subsystem comprising a cell suspension inlet, and one or more cell deformation contractions configured to induce a perturbation in the cell membrane of cells, thereby enabling the entry of a payload into the cells, thereby creating a cell suspension containing the manipulated cells; and a temperature control subsystem fluidly communicating with the cell deformation subsystem, the temperature control subsystem comprising a container configured to receive a cell suspension inlet, a buffer inlet, and a fluid or dry reagent to receive a cell suspension containing the manipulated cells and to mix with the cell suspension containing the manipulated cells to create a diluted cell suspension; and the diluted fine within the container. The system comprises a temperature control subsystem including a plate configured to regulate the temperature of a cell suspension to create an incubated cell suspension; a cell washing subsystem fluidly communicating with the temperature control subsystem, the cell washing subsystem comprising a container configured to receive an incubated cell suspension inlet, a storage medium inlet, and a storage medium that receives the incubated cell suspension and mixes it with the incubated cell suspension in the container, thereby suspending the cells in the storage medium; and a container filling subsystem fluidly communicating with the incubation subsystem, the container filling subsystem comprising an inlet configured to receive cells suspended in the storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump the cells suspended in the storage medium into one or more containers, wherein at least one of the subsystems is detachably coupled to the system frame. In some embodiments, the system is an automated system for processing cells.

[0020] Furthermore, the present disclosure relates generally to a system for automatically processing cells, the system comprising: a suspension preparation subsystem comprising a delivery medium inlet, a cell isolation device configured to isolate cells, and a cell suspension device configured to suspend the isolated cells in a delivery medium, thereby creating a cell suspension; a cell deformation subsystem fluidly communicating with the suspension preparation subsystem, the cell deformation subsystem comprising a cell suspension inlet, and one or more cell deformation contractions configured to induce a perturbation in the cell membrane of cells, thereby enabling the entry of a payload into the cells, thereby creating a cell suspension containing the manipulated cells; a dilution subsystem fluidly communicating with the cell deformation subsystem, the dilution subsystem comprising a cell suspension inlet, a buffer inlet, and a container configured to receive a cell suspension containing the manipulated cells and a fluid or dry reagent to be mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension; and an incubation subsystem fluidly communicating with the dilution subsystem. The system comprises: an incubation subsystem comprising a diluted cell suspension inlet, a container configured to receive the diluted cell suspension, and a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension; a cell washing subsystem fluidly communicating with the incubation subsystem, comprising a container configured to receive an incubated cell suspension inlet, a storage medium inlet, and a container configured to receive an incubated cell suspension and a storage medium to be mixed with the incubated cell suspension in the container, thereby suspending the cells in the storage medium; and a container filling subsystem fluidly communicating with the incubation subsystem, comprising a container filling subsystem comprising an inlet configured to receive cells suspended in the storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump the cells suspended in the storage medium into one or more containers.At least one of the subsystems is detachably coupled to the system frame, and furthermore, the system is configured to be sterile and used in non-sterile locations.

[0021] Furthermore, the disclosure generally relates to a method for processing cells, wherein the method is carried out by a system comprising one or more of the following subsystems: a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method comprises: i. in the suspension preparation subsystem: (1) receiving cells; (2) performing a cell isolation operation on the cells to thereby produce isolated cells; (3) receiving a delivery medium; and (4) producing a cell suspension by suspending the isolated cells in the delivery medium to thereby produce a cell suspension; and ii. in the cell deformation subsystem: (1) receiving a flow of cell suspension from the suspension preparation subsystem; and (2) flowing the cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membranes of the cells to allow the entry of a payload into the cells, thereby producing a cell suspension containing the manipulated cells. iii. Dilution subsystem: (1) receiving a flow of cell suspension containing manipulated cells from the cell deformation subsystem; (2) receiving a fluid or dry reagent; (3) producing a diluted cell suspension by mixing the cell suspension with the fluid or dry reagent; iv. Incubation subsystem: (1) receiving a flow of diluted cell suspension from the dilution subsystem; (2) adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension; v. Cell washing subsystem: (1) receiving a flow of incubated cell suspension from the incubation subsystem; (2) performing a buffer exchange operation on the cells to suspend them in a storage medium; vi. Container filling subsystem: (1) receiving a flow of cells suspended in a storage medium from the cell washing subsystem; (2) introducing the cells suspended in the storage medium into one or more containers.

[0022] Furthermore, this disclosure generally relates to a method for processing cells, wherein the method is carried out by a system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, and a temperature control subsystem, and the method comprises: i. in the suspension preparation subsystem: (1) receiving cells from a container; (2) performing a cell isolation operation on the cells to thereby produce isolated cells; (3) receiving a delivery medium through a delivery medium inlet; and (4) producing a cell suspension by suspending the isolated cells in the delivery medium to thereby produce a cell suspension; and ii. in the cell deformation subsystem: (1) receiving a flow of cell suspension from the suspension preparation subsystem; and (2) inducing a perturbation in the cell membrane of the cells to allow the entry of a payload into the cells, thereby producing one or more cell deformation contractions configured to produce a cell suspension containing the manipulated cells. The method includes: iii. flowing a cell suspension through a pharmacokinetic subsystem; (1) receiving a flow of cell suspension containing manipulated cells from a cell deformation subsystem; (2) receiving a fluid or dry reagent through an inlet; (3) producing a diluted cell suspension by mixing the cell suspension with the fluid or dry reagent; (4) adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension; iv. receiving a flow of incubated cell suspension from a cell washing subsystem; (1) performing a buffer exchange operation on the cells to suspend them in a storage medium; and v. introducing the cells suspended in the storage medium into one or more containers from a container filling subsystem. In some embodiments, the method is an automated method for processing cells. In some embodiments, the method is carried out by a sterile system in a non-sterile environment.

[0023] Furthermore, the present disclosure relates generally to a method for automatically processing cells, wherein the method is carried out by a sterile system in a non-sterile environment, and further comprises one or more of the following subsystems: a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method comprises i. in the suspension preparation subsystem: (1) receiving cells from a container, (2) performing a cell isolation operation on the cells to thereby produce isolated cells, (3) receiving a delivery medium through a delivery medium inlet, and (4) producing a cell suspension by suspending the isolated cells in the delivery medium to thereby produce a cell suspension; and ii. in the cell deformation subsystem: (1) receiving a flow of cell suspension from the suspension preparation subsystem, and (2) inducing a perturbation in the cell membrane of the cells to allow the entry of a payload into the cells, thereby producing a cell suspension containing the manipulated cells through one or more cell deformation contractions configured to produce a cell suspension. The system includes: iii. Streaming a cell suspension; in the dilution subsystem: (1) receiving a stream of cell suspension containing manipulated cells from the cell deformation subsystem; (2) receiving a fluid or dry reagent via a buffer inlet; and (3) producing a diluted cell suspension by mixing the cell suspension containing manipulated cells with the fluid or dry reagent; iv. In the incubation subsystem: (1) receiving a stream of diluted cell suspension from the dilution subsystem; and (2) adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension; v. In the cell washing subsystem: (1) receiving a stream of incubated cell suspension from the incubation subsystem; and (2) performing a buffer exchange operation on the cells to suspend them in a storage medium; and vi. In the container filling subsystem: (1) receiving a stream of cells suspended in a storage medium from the cell washing subsystem; and (2) introducing the cells suspended in the storage medium into one or more containers.

[0024] Furthermore, the present disclosure generally relates to a method for processing cells, wherein the method is carried out by a system comprising one or more of the following subsystems: a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, wherein the method comprises: i. in the suspension preparation subsystem: (1) receiving cells from a container, (2) receiving a delivery medium through a delivery medium inlet, and (3) producing a cell suspension by suspending the cells in the delivery medium; ii. in the cell deformation subsystem: (1) receiving a flow of cell suspension from the suspension preparation subsystem, and (2) flowing the cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membranes of the cells to allow the entry of a payload into the cells, thereby producing a cell suspension containing the manipulated cells; and iii. in the dilution subsystem: (1) the cell deformation subsystem The system includes: (1) receiving a flow of cell suspension containing manipulated cells from the system; (2) receiving a fluid or dry reagent via a buffer inlet; and (3) producing a diluted cell suspension by mixing the cell suspension containing manipulated cells with the fluid or dry reagent; iv. In the incubation subsystem: (1) receiving a flow of diluted cell suspension from the dilution subsystem; and (2) adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension; v. In the cell washing subsystem: (1) receiving a flow of incubated cell suspension from the incubation subsystem; and (2) performing a buffer exchange operation on the cells to suspend them in a storage medium; and vi. In the container filling subsystem: (1) receiving a flow of cells suspended in a storage medium from the cell washing subsystem; and (2) introducing the cells suspended in the storage medium into one or more containers.

[0025] Furthermore, the present disclosure generally relates to a kit for use in a system for processing cells, the kit comprising: i. a first kit comprising removably attachable components configured to be removably attachable to a frame of a suspension preparation subsystem of the system, wherein a first set of the removably attachable components comprises (1) a cell isolation device and / or (2) a cell suspension device; ii. a second kit comprising removably attachable components configured to be removably attachable to a frame of a cell deformation subsystem of the system, wherein a second set of the removably attachable components comprises one or more microfluidic chips comprising one or more cell deformation constrictions by which cells can be forced to cause perturbation of the cell membrane; iii. a third kit comprising removably attachable components configured to be removably attachable to a frame of a temperature control subsystem of the system, wherein a third set of the removably attachable components comprises at least one cell aggregate filter; iv. a fourth kit comprising removably attachable components configured to be removably attachable to a frame of a cell washing subsystem of the system, wherein a fifth set of the removably attachable components comprises a second tangential flow filtration membrane assembly; v. a fifth kit comprising removably attachable components configured to be removably attachable to a frame of a container filling subsystem of the system, wherein a sixth set of the removably attachable components comprises at least one cell aggregate filter, and comprising one or more of the above. The present invention provides, for example, the following: (Item 1) A system for processing cells, wherein the system is A suspension preparation subsystem, delivery medium inlet, A cell isolation device configured to isolate cells, A suspension preparation subsystem comprising a cell suspension device configured to suspend isolated cells in a delivery medium, thereby creating a cell suspension, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, wherein the cell deformation subsystem is Cell suspension inlet, A cell deformation subsystem comprising one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and creating a cell suspension containing the manipulated cells, A dilution subsystem that is in fluid communication with the cell deformation subsystem, wherein the dilution subsystem is Cell suspension inlet, buffer inlet, A dilution subsystem comprising a container configured to receive the cell suspension containing the manipulated cells and a fluid or dry reagent to be mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension, An incubation subsystem that is in fluid communication with the dilution subsystem, wherein the incubation subsystem is Diluted cell suspension inlet, A container configured to receive the diluted cell suspension, and An incubation subsystem comprising a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, A cell washing subsystem that is in fluid communication with the incubation subsystem, wherein the cell washing subsystem is Incubated cell suspension entry point, storage medium entrance, A cell washing subsystem includes a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension within the container, thereby suspending the cells in the storage medium. A container filling subsystem that is in fluid communication with the incubation subsystem, wherein the container filling subsystem is An entry point configured to receive cells suspended in a storage medium, One or more containers configured to receive cells suspended in a preservation medium, and A container filling subsystem comprising one or more pumps configured to pump the cells suspended in a storage medium into one or more containers, A system in which at least one of the subsystems is detachably coupled to the frame of the system. (Item 2) The system according to item 1, wherein the system is sterilized and configured for use in a non-sterile location. (Item 3) The system according to item 1 or 2, wherein the system produces processed cells in approximately 5 to 7 hours. (Item 4) The system described in any one of items 1 to 3, wherein the cells include red blood cells (RBCs). (Item 5) The method according to any one of items 1 to 4, wherein the cells include erythrocyte-derived vesicles. (Item 6) The method according to item 5, wherein the erythrocyte-derived vesicles contain an activated antigen carrier (AAC). (Item 7) The method according to item 5, wherein the erythrocyte-derived vesicles contain a tolerance antigen carrier (TAC). (Item 8) The system described in any one of items 1 to 3, wherein the cells include peripheral blood mononuclear cells (PBMCs). (Item 9) The method according to any one of items 1 to 3, wherein the cells include T cells, B cells, dendritic cells, monocytes, macrophages, eosinophils, basophils, natural killer (NK) cells, natural killer T (NKT) cells, mast cells, or neutrophils. (Item 10) The system according to any one of items 1 to 4 or 8, wherein the cells include antigen-presenting cells (APCs). (Item 11) The method according to any one of items 1 to 10, wherein the cell suspension inlet and the buffer inlet of the dilution subsystem are the same inlet. (Item 12) The method according to any one of items 1 to 11, wherein the buffer inlet is used to deliver fluid to the dilution subsystem. (Item 13) The method according to any one of items 1 to 12, wherein the buffer inlet is used to deliver the dry reagent to the dilution subsystem. (Item 14) The system according to any one of items 1 to 13, wherein the payload includes one or more initialization factors. (Item 15) The system according to any one of items 1 to 14, wherein the payload comprises one or more nucleic acids. (Item 16) The system according to any one of items 1 to 15, wherein the payload includes one or more differentiation factors. (Item 17) The system according to any one of items 1 to 16, wherein the payload includes one or more neuronal reprogramming factors. (Item 18) The system according to any one of items 1 to 17, wherein one or more of the subsystems include one or more components configured to be detachably coupled to the system. (Item 19) The system according to any one of items 1 to 18, wherein the suspension preparation subsystem further includes a washing medium inlet. (Item 20) The system according to any one of items 1 to 19, wherein the suspension preparation subsystem further includes a dilution medium inlet. (Item 21) The system according to any one of items 1 to 20, wherein the suspension preparation subsystem further comprises an elutriation system configured to perform the cell isolation operation on the cells. (Item 22) The system according to any one of items 1 to 21, further comprising a leukocyte removal filter system configured to perform the cell isolation operation on the cells, wherein the suspension preparation subsystem further comprises the leukocyte removal filter system. (Item 23) The system according to any one of items 1 to 22, further comprising a suspension preparation subsystem, a tangential flow filtration system configured to perform a buffer exchange operation. (Item 24) The system according to any one of items 1 to 23, wherein the suspension preparation subsystem further comprises at least one outlet configured to be coupled to at least one container for receiving cells suspended in a delivery medium. (Item 25) The system according to item 24, wherein the container is a bag having at least one inlet and at least one outlet. (Item 26) The system according to any one of items 1 to 25, wherein the suspension preparation subsystem includes a weighing system configured to weigh a container. (Item 27) The weighing system is the system described in item 26, wherein the weighing system includes a tensile load cell. (Item 28) The weighing system is the system described in item 26, wherein the weighing system includes a compression load cell. (Item 29) The weighing system described in item 26, wherein the weighing system includes a straight bar load cell. (Item 30) The system according to any one of items 1 to 29, wherein the suspension preparation subsystem further includes a stirring system comprising a platform configured to contact a container and to oscillate the container up and down. (Item 31) The system according to any one of items 1 to 30, wherein the suspension preparation subsystem further includes a stirring system comprising a plate configured to be in contact with and moving in and out of a container while in contact with the container. (Item 32) The system according to any one of items 1 to 31, wherein the suspension preparation subsystem further comprises one or more pumps configured to move fluid within the subsystem or between fluid-connected subsystems. (Item 33) The system according to any one of items 1 to 32, wherein the suspension preparation subsystem is capable of removing plasma. (Item 34) The system according to any one of items 1 to 33, wherein the suspension preparation subsystem is capable of removing serum. (Item 35) The system according to any one of items 1 to 34, wherein the cell deformation subsystem further comprises a pressurizing system configured to generate pressure to pass the cell suspension through the one or more cell deformation contractions. (Item 36) The system according to any one of items 1 to 35, wherein the cell deformation subsystem further comprises a preparation vessel configured to flow the cell suspension through one or more cell deformation contractions. (Item 37) The system according to item 36, wherein the preparation container is a rigid reservoir subassembly. (Item 38) The system according to any one of items 1 to 37, wherein the cell deformation subsystem further comprises a temperature control system including a heated plate configured to control the temperature of the cell suspension. (Item 39) The system according to any one of items 1 to 38, wherein the cell deformation subsystem further comprises at least one outlet configured to be coupled to at least one container for receiving a cell suspension or a cell suspension containing manipulated cells. (Item 40) The system according to item 39, wherein the container is a bag having at least one inlet and at least one outlet. (Item 41) The system according to item 39 or 40, wherein the cell deformation subsystem further comprises a stirring system including a platform that contacts the at least one container and is configured to oscillate the container up and down. (Item 42) The system according to item 41, wherein the stirring system, which includes a platform in contact with at least one container, is configured to shake the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension or the cell suspension containing manipulated cells. (Item 43) The system according to item 41 or 42, further comprising a stirring system including a plate configured to contact and move in and out of the container while in contact with the container, wherein the cell deformation subsystem further comprises a stirring system. (Item 44) The system according to any one of items 1 to 43, wherein the dilution subsystem includes a weighing system for weighing the container. (Item 45) The weighing system for determining the weight of the container, the system according to item 44, wherein the weighing system includes a tensile load cell. (Item 46) The weighing system for determining the weight of the container, the system according to item 44, wherein the weighing system includes a compression load cell. (Item 47) The weighing system for measuring the weight of the container, the system according to item 44, wherein the weighing system includes a straight bar load cell. (Item 48) The system according to any one of items 1 to 47, wherein the dilution subsystem includes a metering system for measuring the amount of buffer added to the cells in the cell suspension containing the manipulated cells. (Item 49) The system according to item 46, wherein the weighing system for measuring the amount of the buffer solution includes a tensile load cell. (Item 50) The metering system for measuring the amount of the buffer solution, the system according to item 46, comprising a compressed load cell. (Item 51) The weighing system for measuring the amount of the buffer solution, the system according to item 46, wherein the weighing system includes a straight bar load cell. (Item 52) The system according to any one of items 1 to 51, wherein the dilution subsystem includes at least one outlet configured to be coupled to the container configured to receive the cell suspension containing the manipulated cells. (Item 53) The system according to item 52, wherein the container is a bag having at least one inlet and at least one outlet. (Item 54) The system according to any one of items 1 to 53, wherein the dilution subsystem further includes a stirring system comprising a platform in contact with the container, configured to oscillate the container up and down. (Item 55) The system according to item 54, wherein the first stirring system, which includes a platform in contact with the container, is configured to shake the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension containing the manipulated cells or the diluted cell suspension. (Item 56) The system according to any one of items 1 to 55, further comprising a first stirring system including a plate configured to be in contact with and moving in and out of a container. (Item 57) The system according to any one of items 1 to 56, wherein the dilution subsystem further comprises the cell suspension containing the manipulated cells and an illumination system configured to illuminate the diluted cell suspension. (Item 58) The system according to any one of items 1 to 57, wherein the dilution subsystem further comprises a second stirring system including a platform in contact with the container, configured to oscillate the container up and down. (Item 59) The system according to item 58, wherein the second stirring system is configured to shake the container up and down to agitate the cell suspension containing the manipulated cells or the diluted cell suspension, thereby promoting homogeneity of the cell suspension containing the manipulated cells or the diluted cell suspension. (Item 60) The system according to any one of items 1 to 59, further comprising a second stirring system including a plate configured to contact and move in and out of the container. (Item 61) The system according to any one of items 1 to 60, wherein the dilution subsystem includes at least one pump configured to move fluid between fluid-connected subsystems. (Item 62) The system according to any one of items 1 to 61, wherein the incubation subsystem includes a temperature control device configured to regulate the temperature of the diluted cell suspension. (Item 63) The system according to item 62, wherein the temperature control device is a heated plate. (Item 64) The system according to any one of items 1 to 63, wherein the incubation subsystem includes a weighing system configured to measure the amount of the buffer added to the cells in the diluted cell suspension. (Item 65) The weighing system is the system described in item 64, wherein the weighing system includes a tensile load cell. (Item 66) The weighing system is the system described in item 64, wherein the weighing system includes a compression load cell. (Item 67) The weighing system described in item 64, wherein the weighing system includes a straight bar load cell. (Item 68) The system according to any one of items 1 to 67, wherein the incubation subsystem includes a first stirring system including a platform in contact with the container, configured to oscillate the container up and down. (Item 69) The system according to item 68, wherein the first stirring system, which includes a platform in contact with the container, is configured to shake the container up and down to agitate the diluted cell suspension and promote homogeneity of the diluted cell suspension or the incubated cell suspension. (Item 70) The system according to any one of items 1 to 69, wherein the incubation subsystem includes a second stirring system which includes a platform in contact with the container and is configured to oscillate the container up and down. (Item 71) The system according to item 70, wherein the first stirring system, which includes a platform in contact with the container, is configured to shake the container up and down to agitate the cell suspension and promote homogeneity of the diluted cell suspension or the incubated cell suspension. (Item 72) The system according to any one of items 1 to 71, wherein the incubation subsystem includes at least one pump configured to move fluid between fluid-connected subsystems. (Item 73) The system according to any one of items 1 to 72, wherein the cell washing subsystem includes a tangential flow filtration system configured to perform a buffer exchange operation. (Item 74) The system according to any one of items 1 to 73, wherein the cell washing subsystem includes a metering system configured to measure the amount of buffer added to the cells during the buffer exchange operation. (Item 75) The weighing system is a tensile load cell, as described in item 74. (Item 76) The system described in item 74, wherein the weighing system is a compression load cell. (Item 77) The weighing system is a straight bar load cell, as described in item 74. (Item 78) The system according to any one of items 1 to 77, wherein the cell washing subsystem includes at least one outlet configured to be coupled to the container. (Item 79) The system according to item 78, wherein the container is a bag having at least one inlet and at least one outlet. (Item 80) The system according to any one of items 1 to 79, wherein the cell washing subsystem includes a stirring system that includes a platform in contact with the container, configured to oscillate the container up and down. (Item 81) The system according to item 80, wherein the agitation system, which includes a platform in contact with the container, is configured to agitate the container up and down to agitate the cells suspended in the incubated cell suspension or storage medium, thereby promoting homogeneity of the cells suspended in the diluted cell suspension or storage medium. (Item 82) The system according to any one of items 1 to 81, wherein the cell washing subsystem includes an illumination system configured to illuminate the cells suspended in the diluted cell suspension or the storage medium. (Item 83) The system according to any one of items 1 to 82, wherein the storage medium is a freeze-protected medium. (Item 84) The system according to any one of items 1 to 83, wherein the container filling subsystem includes a weighing system configured to measure the amount of cells suspended in the storage medium added to one or more containers. (Item 85) The weighing system is the system described in item 84, wherein the weighing system includes a tensile load cell. (Item 86) The weighing system is the system described in item 84, wherein the weighing system includes a compression load cell. (Item 87) The weighing system is the system described in item 84, which includes a straight bar load cell. (Item 88) The system according to any one of items 1 to 87, wherein the container filling subsystem includes a stirring system that includes a platform in contact with the container, configured to oscillate the container up and down. (Item 89) The system according to item 88, wherein the stirring system, which includes a platform in contact with the container, is configured to shake the container up and down to agitate the cells suspended in the storage medium, thereby promoting homogeneity of the cells suspended in the storage medium in one or more containers. (Item 90) The system according to any one of items 1 to 89, wherein the container filling subsystem includes an illumination system configured to illuminate the cells suspended in the storage medium in one or more containers. (Item 91) The system according to any one of items 1 to 90, wherein the container filling subsystem includes one or more outlets configured to be coupled to one or more containers. (Item 92) The system according to any one of items 1 to 91, wherein one or more containers of the cell washing subsystem include one or more bags having at least one inlet and at least one outlet. (Item 93) The system according to any one of items 1 to 92, wherein the container filling subsystem includes at least one pump configured to move fluid within the subsystem or between fluid-connected subsystems. (Item 94) The system according to any one of items 1 to 93, wherein the system includes one or more pumps configured to pump fluid between two or more of the subsystems. (Item 95) The system described above is used in a non-sterile environment and is one of the systems described in any one of items 1 to 94. (Item 96) The system described in any one of items 1 to 95 is an automated system for processing cells. (Item 97) The system described above is a system for processing cells for cell therapy, as described in any one of items 1 to 96. (Item 98) The system according to any one of items 1 to 97, wherein the suspension preparation subsystem further comprises one or more of the following: a cell aggregate filter, a leukocyte removal filter, piping, pipe fittings, connectors, clamps, sampling bulbs, a carboy, and an air filter. (Item 99) The system according to any one of items 1 to 98, wherein the cell deformation subsystem further comprises one or more of the following: a rigid sample container, a cell aggregate filter, a rigid preparation container, one or more microfluidic chip cartridges, one or more microfluidic chips, piping, pipe fittings, connectors, clamps, air filters, and barrel filters. (Item 100) The system according to any one of items 1 to 99, wherein the dilution subsystem further comprises one or more of a cell aggregate filter, piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys. (Item 101) The system according to any one of items 1 to 100, wherein the incubation subsystem further comprises one or more of the following: cell aggregate filters, piping, pipe fittings, connectors, and clamps. (Item 102) The system according to any one of items 1 to 101, wherein the cell washing subsystem further comprises one or more of a cell aggregate filter, piping, pipe fittings, connectors, clamps, sampling bulbs, air filters, and carboys. (Item 103) The system according to any one of items 1 to 102, wherein the container filling subsystem further comprises one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, and sampling spheres. (Item 104) A method for processing cells, wherein the method is carried out by a system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method is i. In the suspension preparation subsystem: (1) Receiving cells from a container, (2) Performing a cell isolation procedure on the aforementioned cells to produce isolated cells, (3) Receiving the delivery medium through the delivery medium inlet, (4) To produce a cell suspension by suspending the isolated cells in the delivery medium, ii. In the cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby allowing the payload to enter the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the dilution subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) Receiving a fluid or dry reagent through a buffer inlet, (3) To produce a diluted cell suspension by mixing the cell suspension containing the manipulated cells with the fluid or dry reagent, iv. In the incubation subsystem: (1) Receiving the flow of the diluted cell suspension from the dilution subsystem, (2) Adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension, v. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the incubation subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, vi. In the container filling subsystem: (1) The cell washing subsystem receives a stream of cells suspended in the storage medium, (2) A method comprising introducing the cells suspended in a storage medium into one or more containers. (Item 105) The method according to item 104, wherein the cells include red blood cells (RBCs). (Item 106) The method according to item 104 or 105, wherein the cells include erythrocyte-derived vesicles. (Item 107) The method according to item 106, wherein the erythrocyte-derived vesicles contain an activated antigen carrier (AAC). (Item 108) The method according to item 106, wherein the erythrocyte-derived vesicles contain a tolerance antigen carrier (TAC). (Item 109) The method according to item 104, wherein the cells include peripheral blood mononuclear cells (PBMCs). (Item 110) The method according to item 104, wherein the cells include T cells, B cells, dendritic cells, monocytes, macrophages, eosinophils, basophils, natural killer (NK) cells, natural killer T (NKT) cells, mast cells, or neutrophils. (Item 111) The method according to item 104, 105, or 109, wherein the cells include antigen-presenting cells (APCs). (Item 112) The method according to any one of items 104 to 111, wherein the method removes plasma. (Item 113) The method described above, wherein the method is the method described in any one of items 104 to 112, wherein the serum is removed. (Item 114) The method described above is the method described in any one of items 104 to 113, which is carried out in approximately 5 to 7 hours. (Item 115) The method according to any one of items 104 to 114, wherein the method produces a cell therapy drug. (Item 116) The method according to any one of items 104 to 115, wherein the payload includes one or more initialization factors. (Item 117) The method according to any one of items 104 to 116, wherein the payload comprises one or more nucleic acids. (Item 118) The method according to any one of items 104 to 117, wherein the payload comprises one or more differentiation factors. (Item 119) The method according to any one of items 104 to 118, wherein the payload comprises one or more neuronal reprogramming factors. (Item 120) The method according to any one of items 104 to 119, wherein the method is an automated method for processing cells. (Item 121) The method according to any one of items 104 to 120, wherein the method is carried out by a sterilization system in a non-sterile environment. (Item 122) A kit for use in a system for processing cells, wherein the kit is i. A first kit comprising removable connectable components configured to be removablely connectable to a frame of the suspension preparation subsystem of the system, wherein the first set of removable connectable components is (1) Cell isolation devices, and / or (2) The first kit, including a cell suspension device, ii. A second kit comprising removable connectable components configured to be removablely connectable to a frame of the cell deformation subsystem of the system, wherein the second set of removable connectable components comprises one or more microfluidic chips comprising one or more cell deformation contractions that can compel cells to cause perturbations of the cell membrane, iii. A third kit comprising removablely connectable components configured to be removablely connectable to a frame of the dilution subsystem of the system, wherein the third set of removablely connectable components comprises at least one cell aggregate filter, iv. A fourth kit comprising removablely connectable components configured to be removablely connectable to the frame of the incubation subsystem of the system, wherein the fourth set of removablely connectable components comprises at least one cell aggregate filter, v. A fifth kit comprising removable connectable components configured to be removablely connectable to the frame of the cell washing subsystem of the system, wherein the fifth set of removable connectable components comprises a second tangential flow filtration membrane assembly, vi. A sixth kit comprising a detachable connectable component configured to be detachably connectable to a frame of a container-filling subsystem of the system, wherein the sixth set of detachable connectable components comprises at least one cell aggregate filter, and one or more of the following kits. (Item 123) The kit according to item 122, wherein one or more of the components of the kit are configured to be fluidly connected to one or more components of the corresponding subsystem of the system. (Item 124) The kit according to item 122, wherein the cell isolation device comprises at least one elutriation device. (Item 125) The kit according to item 122, wherein the cell isolation device includes at least one leukocyte removal filter. (Item 126) The first kit described above is a kit according to any one of items 122 to 125, comprising a cell isolation device. (Item 127) The kit according to any one of items 122 to 126, wherein the cell suspension device is a tangential flow filtration membrane assembly. (Item 128) i. One or more of the detachably connectable components of the second kit of detachably connectable components are configured to be fluidly connected to one or more of the detachably connectable components of the first kit of detachably connectable components, ii. One or more of the removablely connectable components of the third kit of removablely connectable components are configured to be fluidly connected to one or more of the removablely connectable components of the second kit of removablely connectable components. iii. One or more of the detachably connectable components of the fourth kit of detachably connectable components are configured to be fluidly connected to one or more of the detachably connectable components of the third kit of detachably connectable components. iv. One or more of the removable connectable components of the fifth kit of removable connectable components are configured to be fluidly connected to one or more of the removable connectable components of the fourth kit of removable connectable components, and / or v. The kit according to any one of items 122 to 127, wherein one or more of the detachably connectable components of the sixth kit of detachably connectable components are configured to be fluidly connected to one or more of the detachably connectable components of the fifth kit of detachably connectable components. (Item 129) The first kit of removable and connectable components comprises a cell aggregate filter, as described in any one of items 122 to 128. (Item 130) The kit according to any one of items 122 to 129, wherein the first kit of removablely connectable components includes a leukocyte removal filter. (Item 131) The first kit of removable and connectable components includes a container, as described in any one of items 122 to 130. (Item 132) The first kit of removable and connectable components includes a tangential flow filtration filter assembly, as described in any one of items 122 to 131. (Item 133) The first kit of removable and connectable components includes one or more of the following: pipes, pipe fittings, connectors, clamps, sampling bulbs, carboys, air filters, and tangential flow filtration filter assemblies, as described in any one of items 122 to 132. (Item 134) The second kit of removable and connectable components includes a rigid sample container, as described in any one of items 122 to 133. (Item 135) The kit according to any one of items 122 to 134, wherein the second kit of removablely connectable components includes a cell aggregate filter. (Item 136) The kit according to any one of items 122 to 135, wherein the second kit of removablely connectable components includes a preparation container. (Item 137) The second kit of removable and connectable components is the kit described in any one of items 122 to 136, comprising one or more microfluidic chips. (Item 138) The second kit of removable and connectable components is the kit described in any one of items 122 to 137, comprising one or more microfluidic chip cartridges. (Item 139) The second kit of removable and connectable components is one or more of the following kits, as described in any one of items 122 to 138: pipes, pipe fittings, connectors, clamps, containers, bags, air filters, and barrel filters. (Item 140) The third kit of removable and connectable components is the kit according to any one of items 122 to 139, including a container. (Item 141) The third kit of removablely connectable components includes a cell aggregate filter, as described in any one of items 122 to 140. (Item 142) The third kit of removablely connectable components is the kit described in any one of items 122 to 141, wherein the third kit includes one or more of pipes, pipe fittings, connectors, clamps, sampling bulbs, and carboys. (Item 143) The fourth kit of removablely connectable components includes a cell aggregate filter, as described in any one of items 122 to 142. (Item 144) The fourth kit of removable and connectable components is the kit according to any one of items 122 to 143, comprising one or more containers. (Item 145) The fourth kit of removablely connectable components is the kit described in any one of items 122 to 144, wherein the fourth kit includes one or more of pipes, pipe fittings, connectors, and clamps. (Item 146) The fifth kit of removablely connectable components includes a cell aggregate filter, as described in any one of items 122 to 145. (Item 147) The fifth kit of removable and connectable components is the kit according to any one of items 122 to 146, including a container. (Item 148) The fifth kit of removable and connectable components includes a tangential flow filtration filter assembly, as described in any one of items 122 to 147. (Item 149) The fifth kit of removablely connectable components is the kit described in any one of items 122 to 148, wherein the kit includes one or more of the following: pipes, pipe fittings, connectors, clamps, sampling bulbs, air filters, and carboys. (Item 150) The sixth kit of removable and connectable components includes a container, as described in any one of items 122 to 149. (Item 151) The kit described in item 150, wherein the container is a freezer bag. (Item 152) The sixth kit of removablely connectable components comprises a cell aggregate filter, as described in any one of items 122 to 151. (Item 153) The sixth kit of removablely connectable components is the kit described in any one of items 122 to 152, wherein the sixth kit includes one or more of pipes, pipe fittings, connectors, clamps, and sampling spheres. (Item 154) The kit described in any one of items 122 to 153, wherein the kit includes the first, second, third, fourth, fifth, and sixth kits. (Item 155) The kit described in item 154, which is packaged in an accordion tray package. (Item 156) The kit described in item 154, wherein the kit is packaged as a rollable sheet. (Item 157) Each kit is individually packaged, as described in item 154. (Item 158) The kit described in item 154, which contains at least two kits packaged together. (Item 159) The kit is a sterilized kit as described in any one of items 122 to 158. (Item 160) A system for processing cells, wherein the system is A suspension preparation subsystem comprising a cell isolation device, a tangential flow filtration membrane assembly, a cell aggregate filter, a leukocyte removal filter, and one or more containers, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, wherein the cell deformation subsystem includes one or more microfluidic chips, a rigid sample container, and a cell aggregate filter, A dilution subsystem that is in fluid communication with the cell deformation subsystem, wherein the dilution subsystem includes a container, a cell aggregate filter, and An incubation subsystem that is in fluid communication with the dilution subsystem, wherein the incubation subsystem includes a cell aggregate filter, a container, and A cell washing subsystem that is in fluid communication with the incubation subsystem, wherein the cell washing subsystem includes a tangential flow filtration membrane assembly, a cell aggregate filter, and a container. A container filling subsystem that is in fluid communication with the incubation subsystem, the container filling subsystem comprising a container, a cell aggregate filter, and A system in which subsystems i. to vi. are detachably coupled to the frame of the system. (Item 161) A system for processing cells, wherein the system is A suspension preparation subsystem comprising a delivery medium inlet, a cell isolation device configured to isolate cells, and a cell suspension device configured to suspend the isolated cells in the delivery medium, thereby creating a cell suspension; A payload entry subsystem that is in fluid communication with the suspension preparation subsystem, wherein the payload entry subsystem includes a cell suspension inlet, a payload entry component that induces perturbations in the cell membrane of the cell to enable the entry of the payload into the cell, A dilution subsystem that is in fluid communication with the cell deformation subsystem, wherein the dilution subsystem includes a cell suspension inlet, a buffer inlet, and a container configured to receive the cell suspension containing the manipulated cells and to receive a buffer that mixes with the cell suspension containing the manipulated cells to create a diluted cell suspension, An incubation subsystem having fluid communication with the dilution subsystem, wherein the incubation subsystem includes a diluted cell suspension inlet, a container configured to receive the diluted cell suspension, and a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, A cell washing subsystem that is in fluid communication with the incubation subsystem, the cell washing subsystem comprising: an inlet for incubated cell suspension; a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension within the container, thereby suspending the cells in the storage medium; A container filling subsystem that is in fluid communication with the incubation subsystem, the container filling subsystem comprising: an inlet configured to receive cells suspended in a storage medium; one or more containers configured to receive cells suspended in a storage medium; and one or more pumps configured to pump the cells suspended in the storage medium into the one or more containers, A system in which subsystems i. to vi. can be detachably coupled to the frame. (Item 162) The system according to item 161, wherein the payload entry component includes an electroporation device. (Item 163) The system according to item 161, wherein the payload entry component includes one or more cell deformation contractions through which the cell suspension flows, and the cell deformation contractions are configured to cause a perturbation in the cell membrane of the cell, thereby enabling the entry of the payload into the cell. (Item 164) A system for processing cells, wherein the system is Suspension preparation subsystem, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, A dilution subsystem that is in fluid communication with the aforementioned cell deformation subsystem, An incubation subsystem that is in fluid communication with the aforementioned dilution subsystem, A cell washing subsystem that is in fluid communication with the incubation subsystem, The system comprises a container filling subsystem that is in fluid communication with the incubation subsystem, A system in which at least one of the subsystems is detachably coupled to the frame of the system. (Item 165) A system for processing cells, wherein the system is Suspension preparation subsystem, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, A temperature control subsystem that is in fluid communication with the cell deformation subsystem, A cell washing subsystem that is in fluid communication with the temperature control subsystem, The system comprises a container filling subsystem that is in fluid communication with the incubation subsystem, A system in which at least one of the subsystems is detachably coupled to the frame of the system. (Item 166) A system for processing cells, wherein the system is A suspension preparation subsystem, delivery medium inlet, A suspension preparation subsystem includes a cell suspension device configured to suspend cells in a delivery medium, thereby creating a cell suspension, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, wherein the cell deformation subsystem is Cell suspension inlet, A cell deformation subsystem comprising one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and creating a cell suspension containing the manipulated cells, A dilution subsystem that is in fluid communication with the cell deformation subsystem, wherein the dilution subsystem is Cell suspension inlet, buffer inlet, A dilution subsystem comprising a container configured to receive the cell suspension containing the manipulated cells and a fluid or dry reagent to be mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension, An incubation subsystem that is in fluid communication with the dilution subsystem, wherein the incubation subsystem is Diluted cell suspension inlet, A container configured to receive the diluted cell suspension, and An incubation subsystem comprising a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, A cell washing subsystem that is in fluid communication with the incubation subsystem, wherein the cell washing subsystem is Incubated cell suspension entry point, storage medium entrance, A cell washing subsystem includes a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension within the container, thereby suspending the cells in the storage medium. A container filling subsystem that is in fluid communication with the incubation subsystem, wherein the container filling subsystem is An entry point configured to receive cells suspended in a storage medium, One or more containers configured to receive cells suspended in a preservation medium, and A container filling subsystem comprising one or more pumps configured to pump the cells suspended in a storage medium into one or more containers, A system in which at least one of the subsystems is detachably coupled to the frame of the system. (Item 167) A system for processing cells, wherein the system is A suspension preparation subsystem, delivery medium inlet, A cell isolation device configured to isolate cells, A suspension preparation subsystem comprising a cell suspension device configured to suspend isolated cells in a delivery medium, thereby creating a cell suspension, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, wherein the cell deformation subsystem is Cell suspension inlet, A cell deformation subsystem comprising one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and creating a cell suspension containing the manipulated cells, A temperature control subsystem that is in fluid communication with the cell deformation subsystem, wherein the temperature control subsystem is Cell suspension inlet, buffer inlet, A container configured to receive the cell suspension containing the manipulated cells and to receive a fluid or dry reagent to be mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension, A temperature control subsystem including a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, A cell washing subsystem that is in fluid communication with the temperature control subsystem, wherein the cell washing subsystem is Incubated cell suspension entry point, storage medium entrance, A cell washing subsystem includes a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension within the container, thereby suspending the cells in the storage medium. A container filling subsystem that is in fluid communication with the incubation subsystem, wherein the container filling subsystem is An entry point configured to receive cells suspended in a storage medium, One or more containers configured to receive cells suspended in a preservation medium, and A container filling subsystem comprising one or more pumps configured to pump the cells suspended in a storage medium into one or more containers, A system in which at least one of the subsystems is detachably coupled to the frame of the system. (Item 168) The system described in item 163 is an automated system for processing cells. (Item 169) A system for automatically processing cells, wherein the system A suspension preparation subsystem, delivery medium inlet, A cell isolation device configured to isolate cells, A suspension preparation subsystem comprising a cell suspension device configured to suspend isolated cells in a delivery medium, thereby creating a cell suspension, A cell deformation subsystem that is in fluid communication with the suspension preparation subsystem, wherein the cell deformation subsystem is Cell suspension inlet, A cell deformation subsystem comprising one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and creating a cell suspension containing the manipulated cells, A dilution subsystem that is in fluid communication with the cell deformation subsystem, wherein the dilution subsystem is Cell suspension inlet, buffer inlet, A dilution subsystem comprising a container configured to receive the cell suspension containing the manipulated cells and a fluid or dry reagent to be mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension, An incubation subsystem that is in fluid communication with the dilution subsystem, wherein the incubation subsystem is Diluted cell suspension inlet, A container configured to receive the diluted cell suspension, and An incubation subsystem comprising a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, A cell washing subsystem that is in fluid communication with the incubation subsystem, wherein the cell washing subsystem is Incubated cell suspension entry point, storage medium entrance, A cell washing subsystem includes a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension within the container, thereby suspending the cells in the storage medium. A container filling subsystem that is in fluid communication with the incubation subsystem, wherein the container filling subsystem is An entry point configured to receive cells suspended in a storage medium, One or more containers configured to receive cells suspended in a preservation medium, and A container filling subsystem comprising one or more pumps configured to pump the cells suspended in a storage medium into one or more containers, A system wherein at least one of the subsystems is detachably coupled to the frame of the system, and the system is sterilized and configured for use in a non-sterile location. (Item 170) A method for processing cells, wherein the method is carried out by a system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method is i. In the suspension preparation subsystem: (1) Receiving cells, (2) Performing a cell isolation procedure on the aforementioned cells to produce isolated cells, (3) Receiving the delivery medium, (4) To produce a cell suspension by suspending the isolated cells in the delivery medium, ii. In the cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby allowing the payload to enter the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the dilution subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) To accept a fluid or dry reagent, (3) Mixing the cell suspension with the fluid or dry reagent to produce a diluted cell suspension, iv. In the incubation subsystem: (1) Receiving the flow of the diluted cell suspension from the dilution subsystem, (2) Adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension, v. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the incubation subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, vi. In the container filling subsystem: (1) The cell washing subsystem receives a stream of cells suspended in the storage medium, (2) A method comprising introducing the cells suspended in a storage medium into one or more containers. (Item 171) A method for processing cells, wherein the method is carried out by a system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, and a temperature control subsystem, i. In the suspension preparation subsystem: (1) Receiving cells from a container, (2) Performing a cell isolation procedure on the aforementioned cells to produce isolated cells, (3) Receiving the delivery medium through the delivery medium inlet, (4) To produce a cell suspension by suspending the isolated cells in the delivery medium, ii. In the cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby allowing the payload to enter the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the temperature control subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) Receiving a fluid or dry reagent through the inlet, (3) Mixing the cell suspension with the fluid or dry reagent to produce a diluted cell suspension, (4) Adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension, iv. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the temperature control subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, v. In the container filling subsystem: (1) The cell washing subsystem receives a stream of cells suspended in the storage medium, (2) A method comprising introducing the cells suspended in a storage medium into one or more containers. (Item 172) The method according to item 167, wherein the method is an automated method for processing cells. (Item 173) The method described above is carried out by a sterilization system in a non-sterile environment, as described in item 167 or 168. (Item 174) A method for automatically processing cells, wherein the method is carried out by a sterilization system in a non-sterile environment, and the system further comprises one or more of a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method is i. In the suspension preparation subsystem: (1) Receiving cells from a container, (2) Performing a cell isolation procedure on the aforementioned cells to produce isolated cells, (3) Receiving the delivery medium through the delivery medium inlet, (4) To produce a cell suspension by suspending the isolated cells in the delivery medium, ii. In the cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby allowing the payload to enter the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the dilution subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) Receiving a fluid or dry reagent through a buffer inlet, (3) To produce a diluted cell suspension by mixing the cell suspension containing the manipulated cells with the fluid or dry reagent, iv. In the incubation subsystem: (1) Receiving the flow of the diluted cell suspension from the dilution subsystem, (2) Adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension, v. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the incubation subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, vi. In the container filling subsystem: (1) The cell washing subsystem receives a stream of cells suspended in the storage medium, (2) A method comprising introducing the cells suspended in a storage medium into one or more containers. (Item 175) A method for processing cells, wherein the method is carried out by a system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the method is i. In the suspension preparation subsystem: (1) Receiving cells from a container, (2) Receiving the delivery medium through the delivery medium inlet, (3) To produce a cell suspension by suspending the cells in the delivery medium, ii. In the cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contractions configured to induce a perturbation in the cell membrane of the cells, thereby allowing the payload to enter the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the dilution subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) Receiving a fluid or dry reagent through a buffer inlet, (3) To produce a diluted cell suspension by mixing the cell suspension containing the manipulated cells with the fluid or dry reagent, iv. In the incubation subsystem: (1) Receiving the flow of the diluted cell suspension from the dilution subsystem, (2) Adjusting the temperature of the diluted cell suspension to thereby produce an incubated cell suspension, v. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the incubation subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, vi. In the container filling subsystem: (1) The cell washing subsystem receives a stream of cells suspended in the storage medium, (2) A method comprising introducing the cells suspended in a storage medium into one or more containers. (Item 176) A kit for use in a system for processing cells, wherein the kit is i. A first kit comprising removable connectable components configured to be removablely connectable to a frame of the suspension preparation subsystem of the system, wherein the first set of removable connectable components is (1) Cell isolation devices, and / or (2) The first kit, including a cell suspension device, ii. A second kit comprising removable connectable components configured to be removablely connectable to a frame of the cell deformation subsystem of the system, wherein the second set of removable connectable components comprises one or more microfluidic chips comprising one or more cell deformation contractions that can compel cells to cause perturbations of the cell membrane, iii. A third kit comprising removable connectable components configured to be removablely connectable to the frame of the temperature control subsystem of the system, wherein the third set of removable connectable components comprises at least one cell aggregate filter, iv. A fourth kit comprising removable connectable components configured to be removablely connectable to the frame of the cell washing subsystem of the system, wherein a fifth set of removable connectable components comprises a second tangential flow filtration membrane assembly, v. A fifth kit comprising a detachable connectable component configured to be detachably connectable to a frame of a container-filling subsystem of the system, wherein a sixth set of detachable connectable components comprises at least one cell aggregate filter, the fifth kit, and one or more of the above. [Brief explanation of the drawing]

[0026] [Figure 1] A flowchart is presented comparing a peripheral blood mononuclear cell ("PBMC") manufacturing process that does not include the use of a point-of-care system (the current PBMC manufacturing process) with a PBMC manufacturing process that includes the use of a point-of-care system (the POC manufacturing process) in the embodiment. [Figure 2] A schematic diagram of a PBMC treatment in an embodiment that does not involve the use of a point-of-care system for treating cells is presented. [Figure 3] A schematic diagram of PBMC manufacturing, excluding the use of a point-of-care system, in an embodiment is presented. [Figure 4] A table is presented comparing the unit operation time of a cell processing method that does not include the use of a point-of-care system with that of a cell processing method that includes the use of a point-of-care system in the embodiment. [Figure 5A] A schematic diagram of a reservoir used in a cell processing system that does not include a point-of-care system, as in the embodiment, is presented. [Figure 5B] A schematic diagram of the reservoir of the point-of-care cell processing system in the embodiment is shown. [Figure 6] Two schematic diagrams of the point-of-care system in the embodiment are presented. [Figure 7] A schematic diagram of the point-of-care system in the embodiment is presented. [Figure 8] A schematic diagram of the point-of-care system in the embodiment is presented. [Figure 9]A schematic diagram of a disposable kit for use with a point-of-care system in an embodiment is presented. [Figure 10] A schematic diagram of the point-of-care system in the embodiment is presented. [Figure 11] A schematic diagram of a disposable waste liquid tank kit in an embodiment is shown. [Figure 12] A schematic diagram of a disposable kit for the suspension preparation subsystem (zone 1) for cell processing in an embodiment is presented. [Figure 13] A schematic diagram of a disposable kit for the suspension preparation subsystem (zone 1) for cell processing in an embodiment is presented. [Figure 14] A schematic diagram of the cell deformation subsystem (zone 2) disposable kit in the embodiment is shown. [Figure 15] A schematic diagram of the disposable dilution subsystem (zone 3) kit in the embodiment is shown. [Figure 16] A schematic diagram of the incubation subsystem (zone 4) disposable kit in the embodiment is presented. [Figure 17] A schematic diagram of the disposable cell washing subsystem (zone 5) kit in the embodiment is shown. [Figure 18] A schematic diagram of the disposable kit of the container filling subsystem (zone 6) in the embodiment is shown. [Figure 19] An image of a heated plate for use with a point-of-care system in an embodiment is shown. [Figure 20] A schematic diagram of the bag in the embodiment is shown. [Figure 21] Images of a bag filling and weighing station for use with a point-of-care system in an embodiment are shown. [Figure 22] A schematic diagram of an accordion tray embodiment of a disposable kit for a point-of-care system is presented. [Figure 23]A schematic diagram of an embodiment of a shower curtain for a disposable kit for a point-of-care system is presented. [Figure 24A] A schematic diagram of a direct thermoelectric cooler (TEC) liquid cooling system for use with a point-of-care system in an embodiment is presented. [Figure 24B] A schematic diagram of a direct TEC liquid cooling system for use with a point-of-care system in an embodiment is presented. [Figure 25] A schematic diagram of a stirring system for use with a point-of-care system in an embodiment is presented. [Figure 26-1] A schematic diagram of the point-of-care system (PBMC) process flow in the embodiment is presented. [Figure 26-2] A schematic diagram of the point-of-care system (PBMC) process flow in the embodiment is presented. [Figure 26-3] A schematic diagram of the point-of-care system (PBMC) process flow in the embodiment is presented. [Figure 27] A schematic diagram of the process flow of the suspension preparation subsystem (zone 1) for PBMC in the embodiment is presented. [Figure 28] A schematic diagram of the process flow of the suspension preparation subsystem (zone 1) for RBC in the embodiment is presented. [Figure 29] A schematic diagram of the cell deformation (zone 2) and dilution subsystem (zone 3) process flow for PBMCs in the embodiment is presented. [Figure 30] A schematic diagram of the cell deformation (zone 2) and dilution subsystem (zone 3) process flow for RBCs in the embodiment is presented. [Figure 31] A schematic diagram of the process flow for the incubation subsystem (zone 4), cell washing subsystem (zone 5), and container filling subsystem (zone 6) for PBMCs and RBCs in the embodiment is presented. [Figure 32]A schematic diagram of a pressure integrity test for use with a point-of-care system in an embodiment is presented. [Figure 33] A schematic diagram of the initial buffer injection sector of the point-of-care system in the embodiment is presented. [Figure 34] A schematic diagram of the cell deformation process in the embodiment is presented. [Figure 35-1] A schematic diagram of the point-of-care system RBC process flow in the embodiment is presented. [Figure 35-2] A schematic diagram of the point-of-care system RBC process flow in the embodiment is presented. [Figure 35-3] A schematic diagram of the point-of-care system RBC process flow in the embodiment is presented. [Modes for carrying out the invention]

[0027] Detailed description of the invention 1. A system for processing cells a. Background As discussed above, other systems and methods for cell processing may take several days to several weeks to produce the desired cell therapy. Furthermore, other manufacturing processes generally involve the use of several different instruments, each requiring its own configuration and cleaning, significantly adding to the time required to produce the final product. Moreover, other methods for producing cell therapy require several different instruments and sterile environments, thus requiring a large amount of physical space, i.e., multiple cleanrooms, to carry out the manufacturing process. For example, Figure 1 presents a flowchart of a cell processing manufacturing process in which PBMCs are processed, without the use of a point-of-care system. The PBMC process without the use of a point-of-care system comprises 14 separate steps (see Figure 1, Current PBMC Manufacturing Process, 1-14), each requiring dedicated equipment and materials. By comparison, and as will be discussed further herein, the point-of-care system for processing cells described herein may be used as part of a point-of-care manufacturing process, which comprises four steps (see Figure 1, POC Manufacturing Process, 1-4), and furthermore, these steps are carried out using a single point-of-care system described herein.

[0028] As discussed above, the overall process efficiency is limited by the number of off-the-shelf technologies currently available and the number of off-the-shelf technologies used to process the cells. For example, referring to Figures 2 and 3, a method for processing cells is shown that does not involve the use of a point-of-care system, and this process involves the use of off-the-shelf components. As presented in Figure 2, numerous instruments are used to carry out the operation of the process, and these include (1) peristaltic pumps for pumping fluids for a given step, (2) devices such as LOVO devices for carrying out cell washing and buffer exchange operations, (3) cell isolation devices such as elutriation devices for carrying out cell isolation, (4) microfluidic chips and cartridges for carrying out cell deformation processes, and (5) devices for temperature control such as oscillating incubators and refrigerators. For example, as shown in Figure 2, the following components are used during each of the 14 steps of a system and method for processing cells that does not involve the use of a point-of-care system: in step 1, a container containing the sample such as a LeukoPak receiver; in step 2, a device for hemodilution; in step 3, a PBMC purification device such as a water elutriation device; in step 4, a LOVO device for premechanoporation; in step 5, a cell deformation system such as a mechanoporation system; in step 6, a device for CpG addition; in step 7, a device for product filtration; in step 8, a device for agitating the incubation; in step 9, a second product filtration device; in step 10, a LOVO device for pre-freezing and storage operations; in step 11, a third product filtration device; in step 12, a vial or container filling device such as a cryovial filler; in step 13, a speed-controlled refrigerator; and in step 14, a storage unit such as a storage unit in a cryopreservation tank.

[0029] Furthermore, as shown in Figure 3, a system for processing cells that does not involve the use of a point-of-care system includes several hidden operations within the 14-step process, including in-process sterile seals / junctions, represented by stars in Figure 3, in-process cell counts, represented by squares in Figure 3, and in-process weighing, represented by circles in Figure 3. For example, cell counts and weights are required as input parameters for certain parts of the instrument, and each cell count may take up to 30 minutes to perform. Moreover, each step in the 14-step process presented in Figures 2 and 3 must be documented, signed, and verified, representing a considerable investment of time. Furthermore, some instruments require the use of dedicated single-use kits that can only be assembled during each manufacturing operation. Furthermore, as shown in Figure 5A, processing cells using the process of Figure 2 or Figure 3 may require 12 or more reservoirs compared to just 6 or fewer reservoirs for processing cells using the systems for cell processing described herein (see Figure 5B).

[0030] Furthermore, processing cells using the process in Figure 2 or Figure 3 can take 15–18 hours in manufacturing time, if not in a hurry. For example, Figure 4 presents a table detailing the average time to complete each step of a cell manufacturing process that does not include the use of a point-of-care system, as presented in Figures 2 and 3. For comparison, the time for each corresponding step when using the cell processing system described herein is also presented in Figure 4. As presented in Figure 4, the use of a point-of-care system for cell processing described herein can be used to process cells in less time compared to a cell manufacturing process that does not include the use of a point-of-care system. For example, a manufacturing process that does not include the use of a point-of-care system may take about 15 hours and 20 minutes, while manufacturing cells using the cell processing system described herein can save a significant amount of time, about 7 hours, compared to the current process for manufacturing cells.

[0031] b. Point of care system Therefore, this disclosure generally relates to systems and methods for processing cells, and kits for use with such systems and methods. In some embodiments, a suspension preparation subsystem (Zone 1) may include a delivery medium inlet, a cell isolation device configured to isolate cells, also called a cell isolation device, and a cell suspension device configured to suspend the isolated cells in a delivery medium, thereby creating a cell suspension. In some embodiments, the delivery medium inlet of Zone 1 may be used to introduce any type of fluid, such as a delivery medium, into the system. In some embodiments, the delivery medium may be any buffer or fluid compatible with the cells to be suspended in the delivery medium. In some embodiments, Zone 1 may omit the cell isolation device. In some embodiments, Zone 1 may include one or more additional inlets, such as an additional inlet for introducing a sample into Zone 1, an inlet for introducing a buffer into Zone 1, an inlet for introducing a washing medium such as RPMI or PBS into Zone 1, an inlet for introducing a diluent into Zone 1, and / or an inlet for introducing an antigen into Zone 1. In some embodiments, an inlet that is in fluid communication with Zone 1 may also be in fluid communication with Zone 2. In some embodiments, the fluid introduced into the system through one of the inlets of Zone 1 may be temperature-controlled so that the fluid is introduced into the system at a desired temperature. In some embodiments, the cells introduced into Zone 1 may be isolated from undesirable cell types and aggregates, for example, by using a cell isolation device in Zone 1. In some embodiments, the cells introduced into Zone 1 may be buffers exchanged in a delivery medium, for example, by using a cell suspension device in Zone 1. In some embodiments, the cell suspension device includes one or more tangential flow filtration (TFF) assemblies. In some embodiments, the cell suspension device includes one or more ultrasonic standing wave devices having cross-flow tips. In some embodiments, the cell suspension device includes one or more gel filtration chromatography devices. In some embodiments, the cell suspension device includes one or more centrifugal washing devices. In some embodiments, the cell suspension device includes one or more deterministic lateral displacement (DLD) tips.In some embodiments, the cell suspension device includes one or more centrifuge and flexible diaphragm devices. In some embodiments, the cell isolation device includes one or more leukocyte removal filters. In some embodiments, the cell isolation device includes one or more helical inertia separation (SIS) devices. In some embodiments, the cell isolation device includes one or more microfluidic chips. In some embodiments, the cell isolation device includes one or more deterministic lateral displacement (DLD) chips. In some embodiments, the cell isolation device includes one or more elutriation devices. In some embodiments, the cell isolation device includes one or more hydrodynamic microfluidic separation chips. In some embodiments, the cell isolation device includes one or more immunomagnetocellular isolation devices. In some embodiments, the cell isolation device includes one or more acoustic cell processing devices. In some embodiments, the cell isolation device includes one or more fluorescence-activated cell sorting (FACS) devices. In some embodiments, the cell isolation device includes one or more microfluidic centrifuge combination chips. In some embodiments, the cell isolation device includes one or more TFF filter assemblies. In some embodiments, the cell isolation device includes one or more dielectrophoresis (DEP) chips. In some embodiments, the cell isolation device includes one or more microfiltration tips. In some embodiments, the cell isolation device includes one or more buoyancy-activated cell sorting devices. In some embodiments, the cell isolation device includes one or more precipitation devices.

[0032] In some embodiments, the system for processing cells includes a cell deformation subsystem (zone 2) in fluid communication with a suspension preparation subsystem, the cell deformation subsystem including a cell suspension inlet, one or more cell deformation contractions configured to induce perturbations in the cell membrane of the cells, thereby allowing the payload to enter the cells and creating a cell suspension containing the manipulated cells, and a preparation vessel configured to flow the cell suspension through one or more cell deformation contractions. In some embodiments, the preparation vessel may be a rigid reservoir subassembly. In some embodiments, the payload includes cargo to be delivered to the cells. For example, in some embodiments, the payload includes one or more reprogramming factors. In some embodiments, the reprogramming factors may include differentiation factors, i.e., any drugs that can induce the differentiation of cells into different types of cells. In some embodiments, the payload includes one or more neuronal reprogramming factors. In some embodiments, the payload includes polypeptides, lipids, carbohydrates, small molecules, metal-containing compounds, antibodies, transcription factors, nanoparticles, liposomes, fluorescently labeled molecules, or combinations thereof. In some embodiments, the payload includes one or more nucleic acids. In some embodiments, the nucleic acids include DNA, RNA, or both. In certain embodiments, DNA includes recombinant DNA, cDNA, genomic DNA, or a combination thereof. In certain embodiments, RNA includes siRNA, mRNA, miRNA, lncRNA, tRNA, shRNA, auto-amplified mRNA, or a combination thereof. In some embodiments, the preparation vessel may be a rigid reservoir subassembly. In some embodiments, zone 2 may include one or more inlets, such one or more inlets for introducing buffer into zone 2. In some embodiments, a cell suspension from zone 1 may enter zone 2 through the cell suspension inlet. In some embodiments, zone 2 may include a container into which the cell suspension may flow and be stored. In some embodiments, the container is temperature-controlled, such as being cooled using a TEC as described herein. In some embodiments, the cell suspension contained in the container may be agitated, such as by using an agitation system as described herein. Such agitation may prevent cell sedimentation.In some embodiments, the cell deformation process may be influenced using an electroporation device rather than one or more cell deformation contractions configured to induce perturbations in the cell membrane of the cell to allow the entry of a payload into the cell. The electroporation device may be used to allow the entry of a payload into the cell, thereby creating a cell suspension containing the manipulated cells.

[0033] In some embodiments, the system for processing cells includes a dilution subsystem (zone 3) that is in fluid communication with a cell deformation subsystem, the dilution subsystem including a cell suspension inlet, an inlet for introducing fluids or dry reagents, a container configured to receive a cell suspension containing the manipulated cells and a buffer to be mixed with the cell suspension to create a diluted cell suspension. In some embodiments, zone 3 may include one or more additional inlets, the one or more of which are for introducing a fluid, such as a buffer, for example, CpG buffer, into zone 3. In some embodiments, zone 3 includes a container in which a cell suspension containing the manipulated cells may flow and be stored. In some embodiments, the cell suspension containing the manipulated cells in the container of zone 3 may be diluted with the buffer. In some embodiments, zone 3 further includes one or more metering systems that can be used to measure the amount of buffer to be added to the cell suspension containing the manipulated cells. Such metering systems may be tensile load cells, compression load cells, or straight bar load cells. In some embodiments, the weighing system may come into contact with a container, such as a container in Zone 3 containing a cell suspension containing manipulated cells, and may be used to measure the weight of the container. In some embodiments, Zone 3 may further include a stirring system, such as those described herein. The stirring system may be used, for example, to continuously mix the cell suspension containing the manipulated cells and buffer in the container in Zone 3. In some embodiments, Zone 3 may further include one or more aggregate filters, such as a 40 μm aggregate filter, to remove cell debris and / or aggregates.

[0034] In some embodiments, the system for processing cells includes an incubation subsystem (zone 4) in fluid communication with a dilution subsystem, the incubation subsystem including a diluted cell suspension inlet, a container configured to receive the diluted cell suspension, and a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension. In some embodiments, zone 4 may include a container into which the diluted cell suspension can flow and be stored. In some embodiments, the container of zone 4 is in contact with a plate, which may be a temperature-controlled plate. In some cases, the temperature-controlled plate may be used to control the temperature of the diluted cell suspension. In some embodiments, zone 4 may further include a stirring system, such as those described herein. The stirring system may be used, for example, to continuously mix the diluted cell suspension in the container of zone 4. In some embodiments, zone 4 may further include one or more aggregate filters, such as a 40 μm aggregate filter, to remove cell debris and / or aggregates.

[0035] In some embodiments, a system for processing cells includes a cell washing subsystem (zone 5) in fluid communication with an incubation subsystem, the cell washing subsystem including an inlet for an incubated cell suspension, a container configured to receive the incubated cell suspension and a storage medium to be mixed with the incubated cell suspension in the container, thereby suspending the cells in the storage medium. In some embodiments, zone 5 includes one or more additional inlets, such one or more inlets for introducing a buffer, e.g., dimethyl sulfoxide ("DMSO"), into zone 5. In some cases, zone 5 further includes one or more weighing systems that can be used to measure the amount of buffer added to the incubated cell suspension. Such weighing systems may be tensile load cells, compression load cells, or straight bar load cells. In some cases, the weighing system may come into contact with a container, such as the container in zone 5 containing the incubated cell suspension, and may be used to measure the weight of the container. In some embodiments, zone 5 may further include one or more cell suspension devices, such as one or more TFF assemblies. Such cell suspension devices may be used in buffer exchange operations such that the incubated cell suspension is a modified buffer in a cryoprotective storage medium. In some embodiments, the cell suspension device includes one or more tangential flow filtration (TFF) assemblies. In some embodiments, the cell suspension device includes one or more ultrasonic standing wave devices having cross-flow tips. In some embodiments, the cell suspension device includes one or more gel filtration chromatography devices. In some embodiments, the cell suspension device includes one or more centrifugal washing devices. In some embodiments, the cell suspension device includes one or more deterministic lateral displacement (DLD) tips. In some embodiments, the cell suspension device includes one or more centrifugation and flexible diaphragm devices. In some embodiments, zone 4 may further include a stirring system as described herein. The stirring system may be used, for example, to continuously mix the contents of the container in zone 5.

[0036] In some embodiments, the system for processing cells includes a container-filling subsystem (zone 6) in fluid communication with an incubation subsystem, the container-filling subsystem including an inlet configured to receive cells suspended in a storage medium, one or more containers configured to receive cells suspended in the storage medium, and one or more pumps configured to pump cells suspended in the storage medium into one or more containers. In some embodiments, zone 6 comprises additional containers into which a cell suspension storage medium flows and is stored before filling one or more containers. In some embodiments, zone 6 further includes one or more weighing systems, such as those described herein. The weighing systems may be in contact with each of the one or more containers and may be used to measure the weight of each of the one or more containers. In some embodiments, zone 6 further includes an aggregate filter, such as a 40 μm aggregate filter, which may be used to remove cell debris and / or aggregates before filling one or more containers. In some embodiments, zone 6 may further include a stirring system, such as those described herein. The stirring system may be used, for example, to continuously mix the contents of the additional containers in zone 6. In some embodiments, each of the containers in zone 6 may be in contact with its respective temperature-controlled plate to control the temperature of the contents of a given container.

[0037] In some embodiments, the system is sterile and configured for use in non-sterile locations. Such features are advantageous and present significant advantages over manufacturing processes that do not involve the use of point-of-care systems. For example, manufacturing processes for cell processing that do not involve the use of point-of-care systems generally occur in at least one cleanroom facility, which represents significant costs to configure and maintain and requires a significant amount of space to carry out the cell processing. In contrast, the cell processing systems described herein may, in some embodiments, be sterile systems that can be used in non-sterile environments, i.e., environments that are not cleanroom environments. Such systems can be used in many locations that do not have the capacity to provide a cleanroom for cell processing. Therefore, the cell processing systems described herein may be used, for example, in hospitals or other point-of-care locations.

[0038] Referring to Figure 6, in some embodiments, the point-of-care system includes the point-of-care system 4000. In some cases, the frame 4052 may include the frame 6000 in Figure 8. For example, the frame 4052 may include a plate on which a container rests, such as a temperature-controlled plate; a stirring system for mixing the contents of the container throughout the system, as further described below; a weighing system, such as a tensile load cell, compression load cell, or straight bar load cell, for measuring the weight of the container throughout the system; fastening elements, such as hooks, which can be used to removably connect the container or various other components to the frame; filter clips, which can be used to removably connect components to the frame; valves for controlling fluid flow throughout the system; pumps, such as peristaltic pumps, for pumping fluid within and between subsystems; one or more casters for moving the frame and / or the assembled system; a graphical user interface (GUI) which can be used by an operator to control the system; and a backlight for illuminating the container throughout the system. In some embodiments, the fastening elements throughout the system may be any type of connector. For example, fastening elements may be hooks, hook-and-loop fasteners, temporary adhesives, ties, or pins. In some embodiments, temperature-controlled plates may be used to control the temperature of the contents of each container in contact with a given plate. In some cases, the system frame 4052 is designed to be divided into at least two modules, as represented by the dashed lines in the rotated diagram of system 4000. Such division of the system frame may provide ease of transport of the point-of-care system. In some embodiments, the overall dimensions of the assembled system frame 4052 are approximately 8.0 feet long, approximately 6.5 feet high, and approximately 2.3 feet deep. In some embodiments, the point-of-care system 4000 includes a container 4040, which in some cases may contain a fluid for introduction into the system, such as an antigen in a buffer.The container 4040 may be detachably connected to the system by fastening element 4041, which may be a hook and may also function as part of a mechanism for weighing the container 4040, for example, the fastening element 4041 may be part of a weighing system. In some embodiments, the container 4040 is fluidly connected to zone 1 by piping such as pipe 4010, and the fluid contained in the container 4040 is pumped from the bag through the piping by a pump such as pump 4008. The piping 4010 may be PVC piping such as di-2-ethylhexyl phthalate (DEHP) free PVC piping, and the fluid may flow through the piping 4010. The point-of-care system 4000 may further include a container 4042, which may contain a buffer. Container 4042 may be detachably connected to the system, for example by coupling container 4042 to a fastening element 4043, the fastening element may also function as part of a weighing system for weighing container 4042. In some embodiments, container 4042 is fluidly connected to zone 1 by piping, and the fluid contained in container 4042 is pumped from the container through the piping by a pump. The point-of-care system 4000 may further include container 4044, which may contain a sample for processing, such as an input blood sample. Container 4044 may be detachably connected to the system, for example by coupling container 4044 to a fastening element 4045, the fastening element 4045 may be a hook, and may also function as part of a weighing system for weighing container 4044. In some embodiments, the container 4044 is fluidly connected to zone 1 by piping such as piping 4010, and the fluid contained within the container 4044 flows by gravity through an aggregate filter such as aggregate filter 4016 before entering zone 1. In some cases, the aggregate filter 4016 may be a 40 μm aggregate filter capable of filtering particles larger than 40 μm, such as cell aggregates or debris. In some cases, the filter diameter may be varied for a given process. For example, the filter size may be about 10 μm, about 20 μm, about 30 μm, about 40 μm, or about 50 μm. In some embodiments, the filter material of the aggregate filter 4016 is selected for a given process.In some cases, the filter material of the aggregate filter 4016 may be a mixture of cellulose esters, cellulose acetate, coated cellulose acetate, hydrophilic polytetrafluoroethylene (PTFE), hydrophobic PTFE, nylon, or polycarbonate.

[0039] In some embodiments, the sample flowing from container 4044 through aggregate filter 4016 flows into container 4024. In some embodiments, container 4024 is in contact with a plate, such as plate 4025. The plate, such as plate 4025, may be a temperature-controlled plate for heating or cooling the sample in the container. Mechanisms for heating and cooling plates are shown in Figures 19 and 23A-23B, respectively, and are discussed in further detail below. In some embodiments, a stirring system 4023 is in contact with container 4024. In some embodiments, the stirring system 4023 may be used to homogenize the sample in container 4024. The stirring system 4023 may be the stirring system 2000 in Figure 25, an oscillating plate, or a shaking plate. In some embodiments, container 4024 is fluidly connected via piping to a cell suspension device 4012, such as a tangential flow filtration (TFF) filter assembly, and the TFF filter assembly is detachably coupled to the system frame. A cell suspension device 4012, such as a TFF filter assembly, is fluid-connected to a valve, such as a valve 4018, to regulate the inflow and outflow into the cell suspension device 4012, such as a TFF filter assembly. The cell suspension device 4012, such as a TFF filter assembly, can be used to suspend cells in a desired fluid, such as a delivery medium. Furthermore, before entering the cell suspension device 4012, the fluid passes through one or more air filters, such as an air filter 4014, to remove air from the fluid, such as in the form of bubbles. In some embodiments, the cell suspension device 4012 is fluid-connected to one or more cell isolation devices, such as a cell isolation device 4020. In some embodiments, the cell isolation device 4020 includes one or more microfluidic chips. In some embodiments, the cell isolation device 4020 includes one or more SIS devices. In some embodiments, the cell isolation device 4020 includes one or more leukocyte removal filters. In some embodiments, the cell isolation device 5020 includes one or more elutriation devices. In some embodiments, the cell isolation device 4020 is in fluid communication with the container 4024 by piping. In some embodiments, the cell isolation device 4020 may be omitted from zone 1.

[0040] In some embodiments, container 4024 is fluidly connected to container 4047 in zone 2 by piping. The fluid may be pumped from container 4024 by a pump, and the fluid may pass through an aggregate filter to remove any additional cell debris or aggregates that may have formed during processing in zone 1 or during the cell isolation process, for example, by pumping the fluid throughout zone 1 before it enters container 4047. In some embodiments, container 4047 is removably attached to a frame by fastening elements such as hooks, hook-and-loop fasteners, temporary adhesives, ties, or pins. In some embodiments, container 4047 is in contact with a plate, such as plate 4046. Plate 4046 may be temperature-controlled, such as by heating, cooling, or being held at a relatively constant temperature, to control the temperature of the sample in container 4047. Container 4047 is fluidly connected to an aggregate filter by piping, through which the sample may pass before entering the cell deformation device 4026. In some embodiments, the cell deformation device 4026 is in fluid communication with the container 4030 in zone 3 through piping 4010. In some embodiments, the container 4030 is detachably coupled to the system by fastening elements such as hooks. In some embodiments, the container 4030 may be in contact with a plate 4031, which may be temperature-controlled, such as by heating, cooling, or being held at a relatively constant temperature. In some embodiments, the temperature-controlled plate may be a plate as shown in Figure 19. In some embodiments, the temperature-controlled plate may be used to control the temperature of the contents of the container. In some embodiments, the container 4030 is in contact with a stirring system 4029. In some embodiments, the container 4048 is in fluid communication with the container 4030 by piping, and the container 4048 may contain a buffer. The container 4048 may be detachably coupled to the frame 4042 by fastening elements such as 4049.

[0041] In some embodiments, container 4038 is fluidly connected to container 4050 by piping, and the container may contain a buffer. Container 4050 may be removably coupled to frame 4042 by fastening elements 4051, etc. In some embodiments, container 4030 is fluidly connected to aggregate filter 4016. Aggregate filter 4016 may be used to filter out cell debris and aggregates that may accumulate during cell processing using the system. In some embodiments, fluid may flow from container 4030 through the aggregate filter and then pumped to container 4038 in zone 4, which is fluidly connected to the aggregate filter and container 4030 via piping.

[0042] In some embodiments, the container 4038 is detachably coupled to the frame 4052 by fastening elements 4039, which may be hooks in some cases. In some embodiments, the fastening elements 4039 may also be part of a weighing system for weighing the container 4038. In some embodiments, the weighing system may be a tensile load cell, a compression load cell, or a straight bar load cell. In some embodiments, the container 4038 is in contact with an agitation system 4037. The agitation system 4037 may be, for example, the agitation system 2000 in Figure 25, an oscillating plate, or a shaking plate. In some embodiments, the container 4038 is in fluid communication with the container 4050 by piping. In some embodiments, the container 4038 is in fluid communication with an aggregate filter, such as a 40 μm aggregate filter, which may be used to remove cell debris and / or cell aggregates. In some embodiments, the fluid may flow from the container 4038 through the cell aggregate filter and then into the container 4034 in zone 5 through piping.

[0043] In some embodiments, the container 4034 is in fluid communication with a second cell suspension device 4012, such as a second TFF filter assembly. In some embodiments, the cell suspension device 4012, such as a second TFF filter assembly, may be used to suspend cells in a cryopreservation medium. In some embodiments, the second cell suspension device 4012 is detachably coupled to the frame of the system. In some embodiments, the container 4034 is in fluid communication with a container 4056 in zone 6 by piping. In some embodiments, the container 4034 is in contact with a stirring system 4033. The stirring system 4033 may be used to mix the contents of the container. In some cases, the container 4034 may be further placed on a plate 4035, which may be a temperature-controlled plate.

[0044] In some embodiments, container 4056 is in contact with a stirring system 4057. In some embodiments, container 4056 is removably coupled to the system frame by fastening elements such as hooks. In some embodiments, container 4056 is in contact with a plate, such as a temperature-controlled plate. In some embodiments, container 4056 is in contact with a stirring system 4056'. Container 4056 is further in fluid communication with container 4054, which may function as a container for processed cells. In some embodiments, container 4054 may be placed on a weighing system 4055 to measure the weight of bags containing processed cells. In some embodiments, the point-of-care system 4000 further includes waste containers for buffer 4002, general system waste 4004, and DMSO buffer 4006. Each of these containers is in fluid communication with subsystems of the system, e.g., zones 1-6, via piping. In some embodiments, each of the waste liquid containers is in contact with a metering system, which may include a tensile load cell or a compression load cell.

[0045] In some embodiments, one or more of zones 1-6 may be omitted from the system. For example, zone 4 may be omitted from the system, and zone 3 may be used to perform the function otherwise performed by zone 4. For example, the plate 4031 in zone 3 may be a temperature-controlled plate so that the container 4030 can be incubated by using the plate 4031. For example, zone 5 may be omitted from the system, and zone 1 may be used to perform the function otherwise performed by zone 5. For example, a cell suspension device in zone 1, such as a TFF filter assembly, may be used to suspend cells in a cryopreservation medium so that the function can be performed by the components of zone 5. In some embodiments, the piping of each zone is fabricated independently of a second different zone so that the first zone can be connected to a second different zone by connecting the piping of the first and second zones. Such fabrication allows the zones to be assembled in different orientations and use different numbers of zones as desired for a given process.

[0046] In some embodiments, the point-of-care system may include a temperature control subsystem. The temperature control subsystem may be used to perform functions otherwise performed by the dilution subsystem (zone 3) and the incubation subsystem (zone 4). For example, the temperature control subsystem may include a container for receiving a cell suspension containing manipulated cells from the cell deformation subsystem (zone 2), and one or more inlets for introducing fluids and / or dry reagents into the container containing the cell suspension. The fluids and / or dry reagents may be mixed with the cell suspension, for example, by the use of a stirring system, to create a diluted cell suspension. The temperature control system may further include a plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension. In some embodiments, the temperature control system may be fluid-connected to the cell deformation subsystem (zone 2) and also to the cell washing subsystem (zone 5).

[0047] In some embodiments, zones 3 and 4 may be merged. For example, a filtration-recirculation loop may be used when merging zones 3 and 4. In some embodiments, a first container in zone 2 that contacts a temperature-controlled plate may be merged with zone 1. For example, a temperature-controlled plate may be added to zone 1 to contact a container, and the first container in zone 2 may be eliminated. In some embodiments, a cell suspension device in zone 1, such as a TFF filter assembly, may be used to perform cell suspension operations performed by a cell suspension device in zone 5. Therefore, the cell suspension device in zone 5 may be eliminated. For example, the cell suspension device in zone 1 may be rinsed before reuse for cell washing operations. A branch from zone 5 to zone 1 may be added, such as by piping, to directly connect zones 5 and 1. In some cases, the cell suspension device in zone 1 may include more than one different filter. In some cases, zone 5 may be eliminated from the system when the cell suspension device in zone 1 is used for cell washing operations. In some embodiments, zone 4 may be merged with zone 5. For example, heating elements and recirculation filter loops may be added to Zone 5 to carry out incubation and cell washing within Zone 5.

[0048] In some embodiments, the point-of-care system 4000 may further include a graphical user interface (GUI) system 4060. The user may interface with the GUI to control the point-of-care system, for example, to plan and execute cell processing operations using the point-of-care system.

[0049] In some embodiments, the housing may be constructed on the work area of ​​the point-of-care system. For example, a disposable kit may be placed on the frame of the point-of-care system, and then a cover such as a band may be used to close the front of the point-of-care system. Closing such a system may, in some cases, help maintain sterile conditions. In some embodiments, the point-of-care system may further include an environmental monitoring system.

[0050] In some embodiments, the point-of-care system includes a point-of-care system 5000 as shown in Figure 7. In some embodiments, the frame 5052 of the point-of-care system 5000 may include a plate on which a container rests, such as a temperature-controlled plate; a stirring system for mixing the contents of the container throughout the system, as further described below; a weighing system, such as a tensile load cell, compression load cell, or straight bar load cell, for measuring the weight of the container throughout the system; fastening elements, such as hooks, which can be used to removably connect the container or various other components to the frame; filter clips, which can be used to removably connect components to the frame; valves for controlling fluid flow throughout the system; pumps, such as peristaltic pumps, for pumping fluid within and between subsystems; one or more casters for moving the frame and / or the assembled system; a graphical user interface (GUI) which can be used by an operator to control the system; and a backlight for illuminating the container throughout the system. In some embodiments, the fastening elements throughout the system may be any type of connector. For example, fastening elements may be hooks, hook-and-loop fasteners, temporary adhesives, ties, or pins. In some embodiments, temperature-controlled plates may be used to control the temperature of the contents of each container in contact with a given plate. In some embodiments, the frame 5052 of the point-of-care system 5000 may include the frame 6000 shown in Figure 8.

[0051] In some embodiments, the point-of-care system 5000 includes a container 5040, which in some cases may contain an antigen. The container 5040 may be detachably connected to the system by, for example, connecting the container 5040 to a fastening element 5041, the fastening element may be a hook, and may also function as part of a weighing system for weighing the container 5040. In some embodiments, the container 4040 is connected to zone 1 by piping such as a pipe 5010, and the fluid contained in the container 5040 is pumped from the bag through the pipe by a pump such as a pump 5008. The pipe 5010 may be a PVC pipe, and the fluid may flow through the pipe 5010.

[0052] The point-of-care system 5000 may further include a container 5042 which may contain a buffer solution. The container 5042 may be removably connected to the system, for example, by coupling the container 5042 to a fastening element 5043, the fastening element may be a hook and may also function as part of a mechanism for weighing the container 5042. In some embodiments, the container 5042 is connected to zone 1 by piping such as a pipe 5010, and the fluid contained in the container 5042 is pumped from the container through the piping by a pump such as a pump 5008.

[0053] The point-of-care system 5000 may further include a container 5044, which may contain a sample for processing, such as an input blood sample. The container 5044 may be detachably connected to the system, for example, by coupling the container 5044 to a fastening element 5045, which may be a hook and may also function as part of a weighing system for weighing the container 5044. In some embodiments, the container 5044 is fluidly connected to zone 1 by piping such as a pipe 5010, and the fluid contained in the container 5044 flows by gravity through an aggregate filter 5016 before entering zone 1. In some cases, the aggregate filter 5016 may be a 40 μm aggregate filter capable of filtering particles larger than 40 μm in size, such as cell aggregates or debris. In some cases, the filter diameter may be modified for a given process. For example, the filter size may be approximately 10 μm, 20 μm, 30 μm, 40 μm, or 50 μm. In some embodiments, the filter material of the aggregate filter 5016 is selected for a given process. In some cases, the filter material of the aggregate filter 5016 may be a mixed cellulose ester, cellulose acetate, coated cellulose acetate, hydrophilic polytetrafluoroethylene (PTFE), hydrophobic PTFE, nylon, or polycarbonate.

[0054] In some embodiments, the sample flowing from container 5044 through aggregate filter 5016 flows into container 5024. In some embodiments, container 5024 is in contact with a plate, such as plate 5025. The plate, such as plate 5025, may be temperature-controlled for purposes such as heating or cooling the sample in the container. In some embodiments, a stirring system 5023 is in contact with container 5024. The stirring system 5023 may be the stirring system 2000 in Figure 25, an oscillating plate, or a shaking plate. In some embodiments, the stirring system 5023 may be used to homogenize the sample in container 5024. In some embodiments, container 5024 is fluidly connected via piping to a cell suspension device 5012, such as a tangential flow filtration (TFF) filter assembly, which is detachably coupled to the system frame. The cell suspension device 5012 is fluidly connected to a valve, such as valve 5018, to regulate the inflow and outflow into the cell suspension device. Cell suspension devices, such as TFF filter assemblies, can be used to suspend cells in a desired fluid, such as a delivery medium. Furthermore, before entering the cell suspension device 5012, the fluid passes through one or more filters, such as an air filter 5014, to remove air from the fluid, such as in the form of bubbles. In some embodiments, the cell suspension device 5012 is in fluid communication with one or more cell isolation devices, such as a cell isolation device 5020. In some embodiments, the cell isolation device 5020 includes one or more SIS devices. In some embodiments, the cell isolation device 5020 is an SIS device. In some embodiments, the cell isolation device is used to separate PBMC cells from other cell types. In some embodiments, the cell isolation device 5020 includes one or more leukocyte removal filters. In some embodiments, the cell isolation device 5020 includes one or more elutriation devices. In some embodiments, the cell isolation device 5020 is in fluid communication with a container 5024 by piping. In some embodiments, zone 1 may omit the cell isolation device 5020.

[0055] In some embodiments, container 5024 is fluidly connected to container 5047 in zone 2 by piping for the cell deformation process to be performed. The fluid may be pumped from container 5024 by a pump, and the fluid may pass through an aggregate filter before entering container 5047. In some embodiments, container 5047 is detachably coupled to a frame by fastening elements, such as hooks. In some embodiments, container 5047 is in contact with a plate 5046. The plate 5046 may be temperature-controlled, such as by heating, cooling, or being held at a relatively constant temperature, to control the temperature of the sample in container 5047. In some embodiments, the temperature-controlled plate may be a plate as shown in Figure 19. In some embodiments, the temperature-controlled plate may be used to control the temperature of the contents of the container. Container 5047 is fluidly connected to an aggregate filter by piping, through which the sample may pass before entering the cell deformation device 5026. In some embodiments, the cell deformation device 5026 is in fluid communication with the container 5030 in zone 3 through piping 5010. In some embodiments, the container 5030 is removably coupled to the system by fastening elements such as hooks, hook fasteners, temporary adhesives, ties, or pins. In some embodiments, the container 5030 may be placed on a plate such as a plate 5031, which may be temperature-controlled. Furthermore, the container 5030 may be in contact with the plate, which may be temperature-controlled, such as by heating, cooling, or being held at a relatively constant temperature. In some embodiments, the container 5030 is in contact with a stirring system 5029. In some embodiments, the container 5048 is in fluid communication with the container 5030 by piping, and the container 5048 may contain a buffer solution. The container 5048 may be removably coupled to the frame 5042 by fastening elements such as 5049.

[0056] In some embodiments, container 5038 is in fluid communication with container 5050 by piping, and the container may contain a buffer solution. Container 5050 may be removably coupled to frame 5042 by fastening elements 5051, etc. In some embodiments, container 5030 is in fluid communication with aggregate filter 5016. In some embodiments, fluid may flow from container 5030 through the aggregate filter and then pumped to container 5038 in zone 4, which is in fluid communication with the aggregate filter and container 5030 via piping.

[0057] In some embodiments, the container 5038 is detachably coupled to the frame 5052 by fastening elements 5039, etc. In some cases, the fastening elements 5039 may be hooks and may also be part of a weighing system for weighing the container 5038. In some embodiments, the container 5038 is in contact with a stirring system 5037. In some embodiments, the container 5038 is in fluid communication with the container 5050 by piping. In some embodiments, the container 5038 is in fluid communication with an aggregate filter 5016. The aggregate filter 5016 may be used to filter out cell debris and aggregates that may accumulate during cell processing using the system. In some embodiments, the fluid may flow from the container 5038 through the cell aggregate filter and then into the container 5034 in zone 5 through piping.

[0058] In some embodiments, the vessel 5034 is in fluid communication with a second cell isolation device 5012, such as a second TFF filter assembly, which is detachably coupled to the frame of the system. In some embodiments, the vessel 5034 is in fluid communication with a vessel 5056 in zone 6 by piping. In some embodiments, the vessel 5034 is in contact with a stirring system 5033. In some cases, the vessel 5034 may further be in contact with a plate 5035, which may be a temperature-controlled plate.

[0059] In some embodiments, the container 5056 is in contact with a stirring system 5057. In some embodiments, the container 5056 is removably coupled to the system frame by fastening elements such as hooks. In some embodiments, the container 5056 is in contact with a plate, such as a temperature-controlled plate. In some embodiments, the container 5056 is in contact with a stirring system 4056. The container 5056 is further in fluid communication with a container 5054, which may function as a container for processed cells. In some embodiments, the container 5054 may be in contact with a weighing system 5055 to measure the weight of bags containing processed cells.

[0060] In some embodiments, the point-of-care system 5000 further includes waste containers for buffer 5002, general system waste 5004, and DMSO buffer 5006. Each of these containers is in fluid communication with subsystems of the system, i.e., zones 1-6, via piping. In some embodiments, each of the waste containers is in contact with a metering system, which may include a tensile load cell or a compression load cell.

[0061] In some embodiments, one or more of zones 1-6 may be omitted from the system. For example, zone 4 may be omitted from the system, and zone 3 may be used to perform the function otherwise performed by zone 4. For example, plate 5031 in zone 3 may be a temperature-controlled plate so that container 5030 can be incubated by using plate 5031. For example, zone 5 may be omitted from the system, and zone 1 may be used to perform the function otherwise performed by zone 5. For example, a cell suspension device in zone 1, such as a TFF filter assembly, may be used to suspend cells in a cryopreservation medium so that the function can be performed by the components of zone 5. In some embodiments, the piping of each zone is fabricated independently of a second different zone so that the first zone can be connected to a second different zone by connecting the piping of the first and second zones. Such fabrication allows the zones to be assembled in different orientations and use different numbers of zones as desired for a given process.

[0062] In some embodiments, the system frame for processing cells may include various different components incorporated into the system frame, such as those presented in Figure 8. As presented in Figure 8, the system frame may, in some cases, include heating stations 6022 and 6027, where the heating stations may be heated plates (see, for example, Figure 22). System 6000 may further include cooling stations 6023 and 6026, which may be an inline TEC liquid cooling system (see, for example, Figures 24A-24B). In some embodiments, system 6000 may include stirring systems 6014, 6016, 6018, 6024, and 6046, such as the stirring system presented in Figure 25. In some embodiments, system 6000 may include waste liquid container holders such as 6002, 6004, and 6006. In some embodiments, each of the waste liquid container holders 6002, 6004, and 6006 may include a weighing system for weighing a given waste liquid container. In some embodiments, the system 6000 includes a filter clip which can be used to removably connect components to the frame 6000. In some embodiments, the frame 6000 includes a pump 6008 for pumping fluid through the system. In some embodiments, the frame 6000 includes a valve 6012 for controlling fluid flow throughout the system. In some embodiments, the frame 6000 includes fastening elements 6020, 6028, 6030, 6032, 6034, 6036, 6038, 6040, 6042, and 6048 which, in some cases, may be hooks. Such fastening elements may be used to removably connect containers to the frame 6000. Furthermore, such fastening elements, such as hooks, may be used as part of a weighing system for measuring the weight of a container detachably coupled to a given fastening element. In some embodiments, the frame 6000 includes a backlight 6044 that can be used to illuminate a sample inside the container, and the container may be detachably coupled to a fastening element 6048. In some embodiments, the fastening elements throughout the system may be any type of connector. For example, the fastening elements may be hooks, hook-and-loop fasteners, temporary adhesives, ties, or pins.

[0063] In some embodiments, the point-of-care system may include one or more TEC cooling loops, as shown in Figures 24A and 24B. Referring here to Figure 24A, the TEC cooling loop 1000 may include a vessel 1002, which includes inlets 1004 and 1006. In some embodiments, the vessel 1002 is in contact with plate 1008, which is in contact with plate 1010. In some embodiments, plate 1010 is in contact with cooling loop piping 1012, which is connected to cooling station 1014 via piping 1012. Referring here to Figure 24B, the TEC cooling loop 1000' may include a vessel 1002', which includes inlets 1006' and 1004', which may be connected to piping 1010'. In some embodiments, piping 1010' is further connected to 1012', and piping 1010' is in contact with plate 1014'. Plate 1014' is in contact with cooling unit 1018', and plate 1016' is in contact with it. In some embodiments, a temperature-controlled plate may be in contact with the TEC cooling loop.

[0064] In some embodiments, a point-of-care system for processing cells may include system 8000 as shown in Figure 10. In some embodiments, the frame 8001 of the point-of-care system 8000 may include a plate on which a container rests, such as a temperature-controlled plate; a stirring system for mixing the contents of the container throughout the system, as further described below; a weighing system, such as a tensile load cell, compression load cell, or straight bar load cell, for weighing the container throughout the system; fastening elements, such as hooks, which can be used to removably connect the container or various other components to the frame; filter clips, which can be used to removably connect components to the frame; valves for controlling fluid flow throughout the system; pumps, such as peristaltic pumps, for pumping fluid within and between subsystems; one or more casters for moving the frame and / or the assembled system; a graphical user interface (GUI) which can be used by an operator to control the system; and backlights for illuminating the container throughout the system. In some embodiments, the fastening elements throughout the system may be any type of connector. For example, the fastening elements may be hooks, hook-and-loop fasteners, temporary adhesives, ties, or pins. In some embodiments, a temperature-controlled plate may be used to control the temperature of the contents of each container in contact with the given plate.

[0065] In some embodiments, system 8000 includes a caster 8002, which may allow an operator to move system 8000. In some embodiments, system 8000 includes inlets 8049, 8051, and 8056, which may be used to introduce a fluid, e.g., a sample, e.g., a buffer, e.g., a delivery medium, into zone 1 of system 8000. In some embodiments, the inlets may have an air filter 8050, which is fluid-connected to the inlets. The air filter may be used to remove air, such as in the form of bubbles, from the fluid introduced into the system. In some embodiments, zone 1 of system 8000 further includes a cell isolation device 8036, which may be a leukocyte removal filter. In some embodiments, the cell isolation device 8036 includes one or more microfluidic chips. In some embodiments, the cell isolation device 8036 includes one or more SIS devices. In some embodiments, the cell isolation device 8036 includes one or more leukocyte removal filters. In some embodiments, the cell isolation device 8036 includes one or more elutriation devices. In some embodiments, zone 1 may omit the cell isolation device. In some embodiments, zone 1 further includes one or more cell suspension devices, such as a TFF filter assembly 8020 that can come into contact with plate 8022. The cell suspension device 8020 may be used to suspend cells in a desired buffer, such as a delivery medium. The cell suspension device 8020 is further connected to a pressure gauge 8038 to monitor the pressure. The cell suspension device 8020 is detachably coupled to the frame of the system. In some embodiments, zone 1 further includes a waste outlet 8010.

[0066] In some embodiments, Zone 1 further includes a container 8052 that is in contact with a plate 8054 and can be removably coupled to the system frame by fastening elements 8088, which may be hooks in some examples. In some examples, the plate 8054 may be a temperature-controlled plate, such as being heated, cooled, or maintained at a relatively constant temperature. In some embodiments, Zone 1 further includes a valve, such as a valve 8018, to control the movement of fluid within Zone 1. In some embodiments, the container 8052 is in fluid communication with 8048. In some embodiments, the container 8052 is in fluid communication with a cell suspension device via piping 8012. In some embodiments, the container 8052 is in fluid communication with a container 8058 in Zone 2 via piping.

[0067] In some embodiments, the vessel 8058 in zone 2 may be in contact with a plate 8059, which may be a temperature-controlled plate, such as being heated, cooled, or maintained at a relatively constant temperature. In some embodiments, the vessel 8058 is removably coupled to the frame 8001 via fastening elements 8090, which may be hooks. In some embodiments, the vessel 8058 is in fluid communication with the vessel 8060 via piping. In some embodiments, the vessel 8060 is in contact with a plate 8061, which may be a temperature-controlled plate, such as being heated, cooled, or maintained at a relatively constant temperature. In some embodiments, the vessel 8060 is further in fluid communication with an inlet 8064, which is connected to an air filter 8066. Furthermore, the vessel 8060 is fluidly connected to an aggregate filter 8062 via piping, which is further fluidly connected to a cell deformation device 8040. In some embodiments, the cell deformation device 8040 includes an electroporation device. In some embodiments, the cell deformation device 8040 includes one or more microfluidic chips, each containing one or more cell deformation contractions configured to induce a perturbation in the cell membrane of a cell, thereby allowing a payload to enter the cell. In some embodiments, the cell deformation device 8040 is fluidly connected to a vessel 8028 in zone 3 via piping. In some embodiments, a valve, such as valve 8024, may direct the fluid flow into a waste outlet, such as waste outlet 8010.

[0068] In some embodiments, the container 8028 is in contact with a plate 8026, which may be a temperature-controlled plate, such as being heated, cooled, or maintained at a relatively constant temperature. In some embodiments, the container 8028 is in further fluid communication with an aggregate filter 8030 via piping. The aggregate filter 8030 may be used to filter out cell debris and aggregates that may accumulate during cell processing using the system. In some embodiments, the aggregate filter is fluidly connected to a container 8070 in zone 4 via piping. In some embodiments, the container 8070 is removably coupled to the frame 8001 by fastening elements 8092, which may be hooks in some cases. In some embodiments, the container 8070 is in contact with a plate 8072, which may be a temperature-controlled plate, such as being heated, cooled, or maintained at a relatively constant temperature. In some embodiments, the container 8070 is in fluid communication with an inlet 8068 via piping. In some embodiments, the inlet 8068 may be used to introduce a buffer into zone 4. In some embodiments, container 8070 is in fluid communication with aggregate filter 8074 via piping. In some embodiments, aggregate filter 8074 is in fluid communication with container 8078 in zone 5 via piping. In some embodiments, container 8078 in zone 5 is in contact with plate 8080, which may be a temperature-controlled plate. In some embodiments, container 8078 is removably coupled to frame 8001 by fastening elements 8094, which may be hooks in some cases. In some embodiments, container 8078 is in fluid communication with inlet 8042 via piping. In some embodiments, container 8078 is in fluid communication with a cell suspension device, such as a TFF filter assembly 8032, via piping. Such a cell suspension device may be used in a buffer exchange operation, such as when an incubated cell suspension is a modified buffer in a cryoprotective storage medium. In some embodiments, inlet 8076 is fluidly connected to a cell suspension device, such as a TFF filter assembly 8032, via piping. In some embodiments, a cell suspension device, such as a TFF filter assembly 8032, is detachably coupled to the system frame 8001.In some embodiments, waste liquid from zone 5 may flow through piping to the waste liquid outlet 8014.

[0069] In some embodiments, container 8078 is fluidly connected to container 8084 in zone 6 via piping. In some embodiments, container 8084 is in contact with plate 8086, which may be a temperature-controlled plate. In some embodiments, container 8084 is removably coupled to frame 8001 by fastening elements 8096, which may be hooks in some cases. In some embodiments, container 8084 is in fluid communication with inlets 8082 and 8046 via piping. In some embodiments, container 8084 is in fluid communication with aggregate filter 8044 via piping. In some embodiments, container 8084 is fluidly connected to a valve, such as valve 8034, which regulates the inflow of fluid into sample container 8016. In some embodiments, waste liquid may flow from a given zone via piping into waste liquid containers, such as waste liquid containers 8004 and 8006. In some embodiments, waste liquid from two or more different zones flows into the same waste liquid container. In some embodiments, waste liquids from two or more zones flow into their respective waste liquid containers. In some embodiments, waste liquids are grouped by type and flow into containers for a given type of waste liquid. For example, system 8000 may include dedicated waste liquid containers for buffers, general system waste liquids 8004, and DMSO buffers 8006. In some embodiments, each waste liquid container is in contact with a metering system, which may include a tensile load cell or a compression load cell.

[0070] In some embodiments, the point-of-care system includes one or more heated plates, as shown in Figure 19. Referring here to Figure 19, the heated plate 70 includes a plate 700 which can be coupled to a mounting bar 704 by fasteners 702 such as screws, bolts, nuts, washers, and retaining rings. The plate 700 is further connected to a heating unit 706 which can be used to control the temperature of the plate by connectors 708 such as wire connectors and temperature ring connectors.

[0071] In some embodiments, the system for processing cells may be an automated system for processing cells. For example, the system may be operated with minimal operator interaction with the system, for example, in one or more zones. In some embodiments, the operator may load cells into the system, for example, by detachably coupling a LeukoPak to the system frame, and then the operator may start cell processing by using the system's GUI. In some embodiments, the operator may use the GUI to program cell processing operations, to visualize the progress of operations, to visualize errors in the system, and / or to visualize the results of cell processing operations.

[0072] In some embodiments, the system for processing cells includes piping for fluid connection of the system's components. For example, the piping may be PVC piping, such as DEHP-free PVC piping. In some embodiments, the piping may be flexible plastic piping. In some embodiments, the piping may be rigid plastic piping. In some embodiments, the piping may be metal piping.

[0073] As discussed above, the cell processing system described herein can produce a container containing processed cells in about 5 to 7 hours, which is a significant time saving compared to a manufacturing process that does not involve the use of a point-of-care system. Such time savings can enable cells to be collected from a patient, processed using the system, and administered to the patient in less than a day, resulting in a significant time improvement and significant benefit to the patient receiving the processed cells. Furthermore, as discussed above, in some embodiments, the system is configured to be used in a sterile and non-sterile location. This feature is particularly advantageous when such a system may be used in many locations that do not have the ability to provide a cleanroom for cell processing. Therefore, the cell processing system described herein may be used, for example, in a hospital or another point-of-care location that does not have an alternative sufficient sterile environment for carrying out cell processing that does not involve the use of a cleanroom or a point-of-care system.

[0074] i. subsystem As discussed above, a system for processing cells may include one or more subsystems, which include a suspension preparation subsystem (zone 1), a cell deformation subsystem (zone 2), a dilution subsystem (zone 3), an incubation subsystem (zone 4), a cell washing subsystem (zone 5), and a container filling subsystem (zone 6). In some cases, a system for processing cells may include a temperature control subsystem, which in some cases may be used instead of the dilution subsystem and the incubation subsystem, as further described below.

[0075] 1. Suspension preparation subsystem (Zone 1) In some embodiments, the systems for processing cells described herein include a suspension preparation subsystem. In some embodiments, the suspension preparation subsystem is designed for processing PBMCs. In some embodiments, the suspension preparation subsystem is designed for processing RBCs. In some embodiments, the suspension preparation subsystem may be used to further suspend the cells in a desired delivery medium, such as by isolating the desired cells from cell debris, aggregates, and undesirable cell types and by using a cell suspension device to perform a buffer exchange operation on the cell suspension. In some embodiments, the suspension preparation subsystem includes the subsystem shown in Figure 12. In some embodiments, the suspension preparation subsystem includes the subsystem shown in Figure 13.

[0076] Referring here to Figure 12, in some embodiments, the suspension preparation subsystem 10 includes a cell suspension inlet 130. The cell suspension inlet 130 may be fluidly connected via piping 102 to a cell isolation device 128, such as one or more leukocyte removal filters. The cell isolation device 128, such as a leukocyte removal filter, may be fluidly connected via piping 102 to a pump piping subassembly 104. In some embodiments, zone 1 may omit the cell isolation device. In some embodiments, the suspension preparation subsystem includes a delivery medium inlet 126, which may be used to introduce a delivery medium into the suspension preparation subsystem. The delivery medium inlet 126 may be fluidly connected via piping 102 to a cell suspension device 124, such as a tangential flow filtration (TFF) filter assembly. In some embodiments, the suspension preparation subsystem further includes a washing medium inlet 122, which is coupled to one or more filters 110, such as a 0.2 μm air filter. Such filters may be used to remove particles larger than 0.2 μm and to remove air, such as air in the form of bubbles, from the fluid. In some embodiments, the suspension preparation subsystem further includes a container 118, such as a bag, which is in contact with a plate, such as a plate 114, which in some cases is a temperature-controlled plate. The container 118 may have an inlet and an outlet, such as an inlet 115 and an outlet 114, which are connected to piping. The plate 114 is further fluid-connected to a sample sphere 116. In some embodiments, a fluid, such as a sample, may flow from the container 118 into the sample sphere 116. In some cases, the sample sphere may be welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, the suspension preparation subsystem further includes an outlet, such as outlets 120 and 112, which are fluid-connected to a waste liquid container via piping 102. In some embodiments, the suspension preparation subsystem further includes a diluent inlet 108, which is coupled to one or more filters 110, such as one or more 0.2 μm air filters. In some embodiments, the suspension preparation subsystem further includes an aggregate filter 106, which is located at the end of the suspension preparation subsystem and connected via piping 102 to a pump piping subassembly 104, which is fluidly connected to the cell deformation subsystem 20 via an outlet 100.In some embodiments, the aggregate filter 106 may be a 40 μm aggregate filter capable of filtering particles larger than 40 μm in size, such as cell aggregates or fragments. In some cases, the filter diameter may be varied for a given process. For example, the filter size may be about 10 μm, about 20 μm, about 30 μm, about 40 μm, or about 50 μm. In some embodiments, the filter material of the aggregate filter 4016 is selected for a given process. In some cases, the filter material of the aggregate filter 4016 may be a mixed cellulose ester, cellulose acetate, coated cellulose acetate, hydrophilic polytetrafluoroethylene (PTFE), hydrophobic PTFE, nylon, or polycarbonate. Further referring to Figure 12, TW represents pipe welding, CH represents cyclohexanone, LT represents adhesive, and TS represents seal.

[0077] Referring here to Figure 13, in some embodiments, the suspension preparation subsystem 10' includes a cell suspension inlet 138'. The cell suspension inlet 138' may be fluidly connected via piping 103' to a cell isolation device 136', such as one or more leukocyte removal filters. In some embodiments, zone 1 may omit the cell isolation device. The cell isolation device 136', such as a leukocyte removal filter, may be fluidly connected via piping 103' to a pump piping subassembly 104'. In some embodiments, the suspension preparation subsystem includes a washing medium inlet 110', which is coupled to a filter 108', such as an air filter, and piping 103'. In some embodiments, the suspension preparation subsystem includes a delivery medium inlet 126', which is coupled to a filter 124', such as an air filter, and piping 103'. The delivery medium inlet 126' may be fluidly connected via piping 103' to a cell suspension device 134', such as a tangential flow filtration (TFF) filter assembly. In some embodiments, the cell suspension device 134' is coupled to a pressure gauge such as a pressure gauge 132'. In some embodiments, the suspension preparation subsystem 10' further includes a container 116' such as a bag, which is in contact with a plate such as a plate 114', which in some embodiments is a temperature-controlled plate. The container 116' may have inlets and outlets such as an inlet 122' and an outlet 113', which are connected to piping. The plate 114' is further connected to a sample sphere 120'. In some embodiments, a fluid such as a sample fluid may flow from the container 116' into the sample sphere 120'. In some embodiments, the sample sphere may be welded open so that the fluid inside the sample sphere can be analyzed. In some embodiments, the suspension preparation subsystem further includes outlets such as outlets 130', 128', 118', and 102', which are fluid-connected to a waste liquid container via piping 102. In some embodiments, the container 116 is further fluid-connected via piping to an aggregate filter 112', which is further fluid-connected via piping to an outlet 106', which is an outlet to zone 2. In some embodiments, the aggregate filter 106' may be a 40 μm aggregate filter, which can remove cell debris and / or aggregates.Referring further to Figure 13, TW represents pipe welding, SB represents solvent bonding, and TS represents sealing.

[0078] In some embodiments, the suspension preparation subsystem (zone 1) components shown in Figures 12 and 13 may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10. In some embodiments, the suspension preparation subsystem (zone 1) components may be further used to carry out cell washing performed by the cell washing subsystem (zone 5), since zone 1 contains the components necessary to carry out such a process.

[0079] In some embodiments, the suspension preparation subsystem further includes a washing medium inlet. In some embodiments, the suspension preparation subsystem further includes a dilution medium inlet. In some embodiments, the suspension preparation subsystem further includes an elutriation system configured to perform a cell isolation operation on cells. In some embodiments, the suspension preparation subsystem further includes a leukocyte removal filter system configured to perform a cell isolation operation on cells. In some embodiments, the suspension preparation subsystem further includes a tangential flow filtration system configured to perform a buffer exchange operation. In some embodiments, the suspension preparation subsystem further includes at least one outlet configured to be coupled to at least one container for receiving cells suspended in a delivery medium. In some embodiments, the container is a bag including at least one inlet and at least one outlet.

[0080] In some embodiments, the suspension preparation subsystem includes a weighing system configured to weigh the containers. The weighing system may be part of the system frame. In some embodiments, the weighing system includes a tensile load cell. In some embodiments, the weighing system includes a compression load cell. In some embodiments, the weighing system includes a straight bar load cell. In some embodiments, the weighing system may be used to measure the volume of a fluid, such as a buffer, added to the containers. Such measurements may be performed, for example, when diluting cells in zone 3 or when filling output containers in zone 6 to ensure that each container holds the same amount of product.

[0081] In some embodiments, the suspension preparation subsystem further includes an agitation system comprising a platform configured to contact a container and to oscillate the container up and down. In some embodiments, the suspension preparation subsystem further includes an agitation system comprising a plate configured to contact a container and to move in and out while in contact with the container. For example, an example of such an agitation system is shown in Figure 25. Referring to Figure 25, a container 2004 having an inlet 2008, such as a bag, is placed on the plate 2002. The agitation system further includes a plate 2006, which is movable and in contact with the container 2004. While in contact with the container 2004, the plate 2006 may move in and out, thereby agitating the bag. In some embodiments, the agitation system is part of the frame of the system.

[0082] In some embodiments, the suspension preparation subsystem further includes one or more pumps configured to move fluid within the subsystem. In some embodiments, the suspension preparation subsystem further includes one or more pumps configured to move fluid between fluid-connected subsystems. The pumps may be part of the system frame.

[0083] In some embodiments, the suspension preparation subsystem can remove serum from the sample introduced into the subsystem. In some embodiments, the suspension preparation subsystem can remove plasma from the sample introduced into the subsystem. For example, a cell suspension device of the suspension preparation subsystem, such as a TFF filter assembly, can remove plasma and / or serum during the cell suspension process using the TFF filter assembly. In some embodiments, a cell suspension using a Zone 1 cell suspension device, such as a TFF filter assembly, can remove about 2, 3, 4, 5, 6, 7, 8, 9, or 10 times more serum and / or plasma compared to a sample that was not subjected to the cell suspension.

[0084] In some embodiments, the suspension preparation subsystem further includes one or more of the following components: pipe fittings, connectors, clamps, sampling bulbs, carboys, and air filters.

[0085] 2. Cell deformation subsystem (Zone 2) In some embodiments, the systems for processing cells described herein include a cell deformation subsystem. In some embodiments, the cell deformation subsystem may be used to introduce a payload into the cell. In some embodiments, the cell deformation subsystem further includes a pressurizing system configured to generate pressure to pass a cell suspension through one or more cell deformation contractions. For example, a schematic diagram of a cell being forced to pass through a cell deformation contraction is shown in Figure 34. Referring here to Figure 34, cell 17000 may pass through cell deformation contraction 17002, thereby allowing a payload such as payload 17004 to be placed in the center of the cell.

[0086] In some embodiments, the cell deformation subsystem further includes an electroporation device for carrying out the cell deformation process. In some embodiments, the cell deformation subsystem further includes a temperature control system including a heated plate configured to control the temperature of the cell suspension. In some embodiments, the cell deformation subsystem further includes at least one outlet configured to be coupled to at least one container for receiving a cell suspension or a cell suspension containing manipulated cells.

[0087] In some embodiments, the container is a bag as shown in Figure 20. Referring here to bag 80 in Figure 20, bag 80 may include an outlet pipe 800. In some embodiments, bag 80 may further include an inlet pipe 806 and a grommet 804. In some embodiments, the bag film 802 may consist of ultra-low density polyethylene / ethylene vinyl alcohol (ULDPE / EVOH). In some embodiments, the container is a plastic container, a screw-top cryovial, a bag, a sealed vial, a glass container, a plastic bottle, or a glass container.

[0088] In some embodiments, the cell deformation subsystem further includes a stirring system comprising a platform in contact with at least one container, configured to oscillate the container up and down. In some embodiments, the stirring system comprising a platform in contact with at least one container is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension or a cell suspension containing manipulated cells. In some embodiments, the cell deformation subsystem further includes a stirring system comprising a plate configured to contact and move in and out of the container, for example, as shown in Figure 25. In some embodiments, the stirring system is part of the system frame.

[0089] In some embodiments, the cell deformation subsystem includes a cell deformation subsystem 20 as shown in Figure 14. In some embodiments, the cell deformation subsystem includes a cell suspension inlet 208 which is fluidly connected to a suspension preparation subsystem 10 via piping 202. In some embodiments, the cell suspension inlet 208 may be further fluidly connected to an outlet 218 via piping 202 configured to deliver the cell suspension to a container 214 that contacts a plate 219. In some embodiments, the plate 219 may be temperature-controlled, such as by heating, cooling, or being maintained at a relatively constant temperature. In some embodiments, the plate 219 may be cooled, such as by using a TEC liquid cooling system. In some embodiments, the TEC liquid cooling system may include a TEC liquid cooling system 1000 as shown in Figures 24A-24B. In some embodiments, the container 218 may include an inlet 218 and an outlet 211 which are coupled to the piping and the container. In some embodiments, the plate 219 further includes a sample sphere section 210. In some embodiments, a fluid such as a sample may flow from the container 218 into the sample sphere 210. In some cases, the sample sphere may be welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, the cell deformation subsystem may include a buffer inlet 216, which can be fluidly connected to a pump piping subassembly 212 via piping 202. In some embodiments, the cell deformation subsystem may include an end cap 215, which is removably coupled to the inlet 215' and can be removed after coupling the system to the frame. The inlet 215' can be fluidly connected to a filter 217, which may be a 0.2 μm air filter. The buffer inlet 215' can be further fluidly connected via piping 202 to a preparation container 220, which may be a preparation container 220 such as a rigid reservoir assembly. The preparation container 220 can be further fluidly connected via piping 202 to a barrel filter subassembly 206, which may be a 40 μm barrel filter subassembly. In some embodiments, the barrel filter subassembly 206 may be fluidly connected to the cell deformation device 204 via piping 202. In some embodiments, the cell deformation device 204 is fluidly connected to a dilution subsystem, such as a dilution subsystem 30, via piping 202 and outlet 200.

[0090] The cell deformation device 204 may include a microfluidic device and, in some examples, a cartridge for housing the microfluidic device. In some embodiments, the cell deformation device includes one or more cell deformation contractions configured to induce a perturbation in the cell membrane of a cell, thereby allowing a payload to enter the cell. In some embodiments, the perturbation is induced by flowing a cell suspension through a cell deformation contraction under high pressure, such as by using the cell deformation subsystem 20. In some embodiments, the cell deformation device, such as the microfluidic device and cartridge, may be any of the cell deformation devices described in PCT / US2018 / 066295 and PCT / US2020 / 026891, each of which is incorporated herein by reference in whole.

[0091] In some embodiments, a cell suspension may pass through an electric field generated by at least one electrode after passing through the contraction of a microfluidic chip of a cell deformation device. In some embodiments, the electric field assists in the delivery of a payload from the cell suspension to the cells. For example, the combination of cell deformation contraction and an electric field may be used to deliver a payload, such as a plasmid, into a cell, e.g., the cell nucleus. In some embodiments, one or more electrodes generate the electric field in proximity to the cell deformation contraction of the microfluidic chip. In some embodiments, the electric field is in the range of about 0.1 kV / m to about 100 MV / m, or any number or number in between. In some embodiments, an integrated circuit is used to provide electrical signals to drive the electrodes. In some embodiments, the cells of the cell suspension are exposed to the electric field for a pulse width of about 1 ns to about 1 s, a period of about 100 ns to about 10 s, or any time or time range in between.

[0092] In some embodiments, the cell deformation device 204 may further include an electroporation device, such as one or more electrodes positioned so that cells in a cell suspension are exposed to an electric field generated by one or more electrodes. In some embodiments, cells in a cell suspension pass through an electric field generated by at least one electrode. In some embodiments, the electric field is in the range of about 0.1 kV / m to about 100 MV / m, or any number or number in between. In some embodiments, an integrated circuit is used to provide electrical signals for driving the electrodes. In some embodiments, cells in a cell suspension are exposed to the electroporation for a pulse width of about 1 ns to about 1 s, a period of about 100 ns to about 10 s, or any time or time range in between.

[0093] In some embodiments, the cell deformation device 204 may include one or more contractions, one or more of which are pores or contained within pores. In some embodiments, the pores are contained within a surface. In some embodiments, the surface is a filter. In some embodiments, the surface is a membrane. In some embodiments, the contraction size is a function of the cell diameter. In some embodiments, the contraction size is about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, or about 99% of the cell diameter. Examples of cell deformation devices containing pores for use with the systems, methods, and kits described herein are described in PCT / US2016 / 050287, which is incorporated herein by reference in its entirety.

[0094] In some embodiments, the suspension preparation subsystem components of Figure 14 may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10.

[0095] In some embodiments, the cell deformation subsystem further includes one or more of the following components: a rigid sample container, a cell aggregate filter, a preparation container, a rigid reservoir assembly, one or more microfluidic chip cartridges, one or more microfluidic chips, piping, pipe fittings, connectors, clamps, an air filter, and a barrel filter.

[0096] 3. Dilution subsystem (Zone 3) In some embodiments, the systems for processing cells described herein include a dilution subsystem. In some embodiments, a cell suspension containing the manipulated cells flows from the cell deformation subsystem to the dilution subsystem, and the cell suspension containing the manipulated cells may remain in the container of the dilution subsystem. In some embodiments, the dilution subsystem may be used to dilute the cell suspension containing the manipulated cells in a desired buffer, for example, to achieve a desired concentration of cells or desired buffer conditions before incubation. In some cases, a dilution step may not be required following cell deformation, in which case the dilution subsystem is not included in the system. In some embodiments, the dilution subsystem includes a dilution subsystem 30 as shown in Figure 15. In some embodiments, the dilution subsystem 30 includes a cell suspension inlet 311, which is fluidly connected to the cell deformation subsystem via piping 312. In some embodiments, the cell suspension inlet 311 is further fluidly connected to a container 306, such as a bag, and the connection to the container may be made via an outlet 310. In some embodiments, the container 306 may be in contact with a plate 309. In some embodiments, the plate 309 is temperature-controlled, such as by heating, cooling, or being held at a relatively constant temperature. In some embodiments, the container 306 is further connected to the sample sphere 302. In some embodiments, a fluid such as a sample may flow from the container 306 into the sample sphere 302. In some embodiments, the sample sphere is welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, the container 306 is fluid-connected to the pump piping subassembly 308 and to the waste liquid outlet 300 via the inlet 310 and piping 312. The container 306 may be further fluid-connected to the aggregate filter 304 via the outlet 305 and piping 312. The aggregate filter 304 is connected to the pump piping subassembly 308 via piping 312 and may be further fluid-connected to an incubation subsystem such as the incubation subsystem 40 via the outlet 314. Referring further to Figure 15, TW represents pipe welding, SB represents solvent bonding, and TS represents seal.

[0097] In some embodiments, the dilution subsystem components of Figure 15 may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10. In some embodiments discussed above, the dilution subsystem components may be omitted from the point-of-care system in cases where dilution is not required. In some embodiments, the dilution subsystem may also function as an incubation subsystem (zone 4) when the container 306 is in contact with a temperature-controlled plate.

[0098] In some embodiments, the dilution subsystem includes a weighing system for weighing the container. In some embodiments, the weighing system for weighing the container includes a tensile load cell. In some embodiments, the weighing system for weighing the container includes a compression load cell. In some embodiments, the weighing system for weighing the container includes a straight bar load cell. In some embodiments, the dilution subsystem includes a weighing system for measuring the amount of buffer added to the cells in a cell suspension containing the manipulated cells. In some embodiments, the weighing system for measuring the amount of buffer includes a tensile load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a compression load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a straight bar load cell. In some embodiments, the dilution subsystem includes at least one outlet configured to be coupled to a container configured to receive a cell suspension containing the manipulated cells. In some embodiments, the container is a bag including at least one inlet and at least one outlet. In some embodiments, the container is a bag such as bag 80 in Figure 20.

[0099] In some embodiments, the dilution subsystem further includes a stirring system comprising a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the first stirring system comprising a platform in contact with the container is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension containing the manipulated cells or the diluted cell suspension. In some embodiments, the dilution subsystem further includes a first stirring system comprising a plate configured to be in contact with and moving in and out of the container, such as the stirring system presented in Figure 25. In some embodiments, the stirring system is part of the system frame.

[0100] In some embodiments, the dilution subsystem further includes an illumination system configured to illuminate a cell suspension containing manipulated cells and a diluted cell suspension. Such an illumination system may allow an operator to visually inspect the container for undesirable material, such as cell aggregates or debris. In some embodiments, the dilution subsystem further includes a second stirring system, which includes a platform in contact with the container and configured to oscillate the container up and down. In some embodiments, the second stirring system is configured to oscillate the container up and down to agitate the cell suspension containing manipulated cells or the diluted cell suspension, thereby promoting homogeneity of the cell suspension containing manipulated cells or the diluted cell suspension. In some embodiments, the dilution subsystem further includes a second stirring system, which includes a plate configured to be in contact with and move in and out of the container, such as the stirring system presented in Figure 25. In some embodiments, the stirring system is part of the system frame. In some embodiments, the dilution subsystem includes at least one pump configured to move fluid between fluid-connected subsystems.

[0101] In some embodiments, the dilution subsystem further includes one or more of the following components: a cell aggregate filter, piping, pipe fittings, connectors, clamps, a sampling sphere, and a carboy.

[0102] 4. Incubation Subsystem (Zone 4) In some embodiments, the systems for processing cells described herein include an incubation subsystem. In some embodiments, a diluted cell suspension may flow from the dilution subsystem to the incubation subsystem, and the diluted cell suspension may be incubated in a vessel of the incubation subsystem. In some embodiments, the incubation temperature is in the range of about 10°C to about 40°C, about 15°C to about 40°C, about 20°C to about 40°C, about 25°C to about 40°C, or about 30°C to 40°C. In some cases, a dilution step may not be required following cell deformation, in which case the dilution subsystem is not included in the system, and the incubation subsystem is fluidly connected to the cell deformation subsystem. In some embodiments, the incubation subsystem may be used to carry out the dilution of a cell suspension containing the manipulated cells to form a diluted cell suspension. In some embodiments, the incubation subsystem may include one or more additional inlets, such as an inlet for delivering a buffer to zone 4, to allow zone 4 to dilute the cell suspension containing the manipulated cells.

[0103] In some embodiments, the incubation subsystem includes a dilution subsystem 40 as shown in Figure 16. In some embodiments, the incubation subsystem 40 includes a diluted cell suspension inlet 411, which is fluidly connected to the dilution subsystem via a pipe 412. In some embodiments, the cell suspension inlet 411 is further fluidly connected to a container 406, such as a bag, and the connection to the container may be made via an outlet 410. In some embodiments, the container 406 may be in contact with a plate 409. The plate 409 is temperature-controlled, such as being heated, cooled, or held at a relatively constant temperature in some cases. In some embodiments, the container 406 is further fluidly connected to a sample sphere 402. In some embodiments, a fluid, such as a sample, may flow from the container 406 into the sample sphere 402. In some cases, the sample sphere may be welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, the container 406 is fluidly connected to a pump piping subassembly 408 and a wastewater outlet 400 via an inlet 410 and piping 412. The container 406 may be further fluidly connected to an aggregate filter 404 via an outlet 405 and piping 412. The aggregate filter 404 is connected to the pump piping subassembly 408 via piping 412 and may be further fluidly connected to a cell washing subsystem, such as a cell washing subsystem 50, via an outlet 414. Referring further to Figure 16, TW represents pipe welding, SB represents solvent bonding, and TS represents seal.

[0104] In some embodiments, the incubation subsystem components of Figure 16 may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10. In some embodiments discussed above, the dilution subsystem components may be omitted from the point-of-care system in cases where dilution is not required, and thus Zone 2 is fluidly connected to Zone 4. In some embodiments, the incubation subsystem may also function as a dilution subsystem (Zone 3), if the incubation subsystem includes one or more inlets for introducing fluids such as buffers to dilute the cell suspension containing the manipulated cells.

[0105] In some embodiments, the incubation subsystem includes a temperature control device configured to regulate the temperature of the diluted cell suspension. In some embodiments, the temperature control device is a heated plate. In some embodiments, the heated plate is a heated plate 70 as shown in Figure 19. In some embodiments, the plate is part of the system frame. In some embodiments, the incubation subsystem includes a metering system configured to measure the amount of buffer added to the cells in the diluted cell suspension. In some embodiments, the metering system includes a tensile load cell. In some embodiments, the metering system includes a compression load cell. In some embodiments, the metering system includes a straight bar load cell. In some embodiments, the incubation subsystem includes a first stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the first stirring system including the platform in contact with the container is configured to oscillate the container up and down to agitate the diluted cell suspension and promote homogeneity of the diluted or incubated cell suspension. In some embodiments, the incubation subsystem includes a second stirring system, which includes a platform in contact with the container and is configured to oscillate the container up and down. In some embodiments, the second stirring system, which includes a platform in contact with the container, is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the diluted or incubated cell suspension. In some embodiments, the first and second stirring systems are part of the system frame. In some embodiments, the incubation subsystem includes at least one pump configured to move fluid between fluid-connected subsystems. In some embodiments, the incubation subsystem further includes one or more of the following components: cell aggregate filters, piping, pipe fittings, connectors, and clamps.

[0106] 5. Cell washing subsystem In some embodiments, the systems for processing cells described herein include a cell washing subsystem. In some embodiments, an incubated cell suspension flows from the incubation subsystem to the cell washing subsystem, where the incubated cell suspension is washed, such as through a buffer exchange, and flows into a desired buffer, such as a cryoprotective medium. The buffer exchange may occur using one or more cell suspension devices in Zone 5, such as a TFF filter assembly. In some embodiments, the cell washing subsystem may be omitted from the point-of-care system, and instead, components of Zone 1 may be used to perform the cell washing that would otherwise be carried out by the cell washing subsystem (Zone 5).

[0107] In some embodiments, the cell washing subsystem includes a cell deformation subsystem 50, as shown in Figure 17. In some embodiments, the cell washing subsystem 50 includes an incubated cell suspension inlet 528, which is fluidly connected via a tube 500 to an incubation subsystem, such as an incubation subsystem 40. In some embodiments, the incubated cell suspension inlet 528 is further fluidly connected via piping 500 to a cell suspension device 526, such as a tangential flow filtration (TFF) filter assembly. The cell suspension device 526, such as a TFF filter assembly, may include pressure sensors 524 connected to the inlet and outlet of the cell suspension device 526. In some embodiments, the cell suspension device 526 is further fluidly connected to a container 512 via piping 500 and a pump piping subassembly 506. In some embodiments, the container 512 is a bag. In some embodiments, the container 512 may be in contact with a plate 514, which is a temperature-controlled plate, such as being heated, cooled, or held at a relatively constant temperature in some cases. In some embodiments, the container 512 is fluidly connected to the sample sphere 518. In some embodiments, a fluid such as a sample may flow from the container 512 into the sample sphere 518. In some cases, the sample sphere may be welded open so that the fluid inside the sample sphere can be analyzed. In some embodiments, the container 512 may include an inlet 515 and an outlet 511, the inlet and outlet being connected to piping. In some embodiments, the cell washing subsystem includes a buffer inlet 520 connected to a filter 510, which may be a 0.2 μm air filter. In some embodiments, the cell washing subsystem includes a buffer inlet 523, which is fluidly connected to the container 512 by piping 500. In some embodiments, the cell washing subsystem 50 includes a storage medium inlet 504, which is connected to a filter 510, which may be a 0.2 μm air filter, and is further fluidly connected to the container 512 via piping 500. In some embodiments, the cell washing subsystem includes waste outlets 522 and 516. In some embodiments, the container 512 is fluidly connected via piping 500 to an aggregate filter 508, which may be a 40 μm aggregate filter.In some embodiments, the aggregate filter 508 is fluidly connected to an outlet 502 via piping 500 and a pump piping subassembly 506, and the outlet 502 is fluidly connected to a container filling subsystem such as a container filling subsystem 60. Referring further to Figure 17, TW represents pipe welding, SB represents solvent bonding, and TS represents sealing.

[0108] In some embodiments, the cell washing subsystem components of Figure 17 may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10. In some embodiments, the cell washing subsystem may be omitted from the point-of-care system, and the suspension preparation subsystem (zone 1) components may be used to perform cell washing, such as by exchanging the incubated cell suspension in a storage medium, such as a cryopreservation medium.

[0109] In some embodiments, the cell washing subsystem includes a device configured to perform a buffer exchange operation. In some embodiments, the device includes a tangential flow filtration system configured to perform a buffer exchange operation. In some embodiments, the cell washing subsystem includes a metering system configured to measure the amount of buffer added to the cells during the buffer exchange operation. In some embodiments, the metering system is a tensile load cell. In some embodiments, the metering system is a compression load cell. In some embodiments, the metering system is a straight bar load cell. In some embodiments, the cell washing subsystem includes at least one outlet configured to be coupled to a container. In some embodiments, the container is a bag including at least one inlet and at least one outlet. In some embodiments, the cell washing subsystem includes an agitation system including a platform in contact with the container, configured to rock the container up and down. In some embodiments, the agitation system including a platform in contact with the container is configured to rock the container up and down to agitate cells suspended in an incubated cell suspension or storage medium, thereby promoting homogeneity of cells suspended in a diluted cell suspension or storage medium. In some embodiments, the cell washing subsystem includes an illumination system configured to illuminate cells suspended in a diluted cell suspension or storage medium. Such an illumination system may allow an operator to visually inspect a container for undesirable substances, such as cell aggregates or debris. In some embodiments, the cell washing subsystem further includes one or more of the following components: a cell aggregate filter, piping, pipe fittings, connectors, clamps, a sampling bulb, an air filter, and a carboy.

[0110] In some embodiments, the storage medium is a cryoprotective medium. The cryoprotective medium may be any cryoprotective medium known in the art, but is not limited to, DMSO, glycerol, ethylene glycol, propylene glycol, sucrose, trehalose, 2-methyl-2,4-pentanediol (MPD), sorbitol, proline, glycerol 3-phosphate, and formamide.

[0111] 6. Container filling subsystem In some embodiments, the systems for processing cells described herein include a container-filling subsystem. In some embodiments, the container-filling subsystem may be used to remove aggregates and debris from cells suspended in a storage medium by using one or more aggregate filters of the container-filling subsystem, and then to fill one or more containers of the subsystem with the processed cells. In some embodiments, the container-filling subsystem may include a weighing system to ensure that each of the one or more containers containing the processed cells is filled to a desired volume.

[0112] In some embodiments, the system for processing cells includes a container-filling subsystem 60 as shown in Figure 18. In some embodiments, the container-filling subsystem 60 includes an inlet for receiving cells suspended in a storage medium 614, the inlet being fluidly connected to a container 605 via a pipe 613. In some embodiments, the container 605 may be a bag. In some embodiments, the container 605 may be in contact with a plate 606, the plate may be temperature-controlled, such as being heated, cooled, or held at a relatively constant temperature. In some embodiments, the container 605 is fluidly connected to a sample sphere 608. In some embodiments, a fluid, such as a sample, may flow from the container 406 into the sample sphere 608. In some embodiments, the sample sphere may be welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, the container 605 includes an inlet 609 which is connected to a pipe. In some embodiments, the container 605 is further fluid-connected to an outlet 600 via an outlet 607, piping 613, and a pump piping subassembly 604, the outlet being configured to connect to a container for the received processed cells. In some embodiments, the container filling subsystem 60 further includes a waste liquid outlet 612. In some embodiments, the container filling subsystem 60 further includes a buffer inlet 610. In some embodiments, the container filling subsystem 60 further includes an aggregate filter 602 which is fluid-connected to the container 605 via piping 613. Referring further to Figure 18, TW represents pipe welding, SB represents solvent bonding, and TS represents seal.

[0113] In some embodiments, the container-filling subsystem components of Figure 18 can be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10. In some embodiments, the number of containers for receiving the treated cells may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 14, 16, 18, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 100, 105, 110, 115, 120, 125, 130, 135, 140, 145, 150, 155, 160, 165, 170, 175, 180, 185, 190, 195, or 200 individual containers such as bags or vials. In some embodiments, the number of containers for receiving the treated cells may be 5 to 20 containers.

[0114] In some embodiments, the container filling subsystem 60 further includes a container filling station, such as the container filling station 90 shown in Figure 21. In some embodiments, the container filling station 90 includes a housing 900. In some embodiments, the filling station further includes a container 904 including an inlet 906, the container being in contact with a metering system 902. The container 904 may be further coupled to a valve 908. The metering system 902 may include, for example, a compression load cell, a tension load cell, or a straight bar load cell. In some embodiments, the container filling station includes a container 918 including an inlet 922 coupled to a valve 920. The container 918 also includes an outlet 916 coupled to piping 910. Furthermore, the container 918 is in contact with a stirring system 914. The container filling station 90 may further include valves, such as 912, to control the fluid flow within the station.

[0115] In some embodiments, the container filling subsystem includes a weighing system configured to measure the amount of cells suspended in a storage medium added to one or more containers. In some embodiments, the weighing system includes a tensile load cell. In some embodiments, the weighing system includes a compression load cell. In some embodiments, the weighing system includes a straight bar load cell. In some embodiments, the container filling subsystem includes an agitation system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the agitation system including the platform in contact with the container is configured to oscillate the container up and down to agitate the cells suspended in the storage medium, thereby promoting homogeneity of the cells suspended in the storage medium in one or more containers. In some embodiments, the container filling subsystem includes an illumination system configured to illuminate the cells suspended in the storage medium in one or more containers. In some embodiments, the container filling subsystem includes one or more outlets configured to be coupled to one or more containers. In some embodiments, one or more containers of the cell washing subsystem include one or more bags including at least one inlet and at least one outlet. In some embodiments, the container-filling subsystem includes at least one pump configured to move fluid within the subsystem or between fluid-connected subsystems. In some embodiments, the container-filling subsystem further includes one or more of the following components: a cell aggregate filter, piping, pipe fittings, connectors, clamps, and a sampling sphere.

[0116] 7. Temperature control subsystem In some embodiments, the cell processing system described herein includes a temperature control subsystem. In some embodiments, the temperature control subsystem may be used to perform functions otherwise performed by the dilution subsystem (zone 3) and the culture subsystem (zone 4). For example, in some embodiments, a cell suspension containing the manipulated cells may flow from the cell deformation subsystem to the temperature control subsystem, and the cell suspension containing the manipulated cells may remain in the container of the temperature control subsystem. In some embodiments, the temperature control subsystem may be used to dilute the cell suspension containing the manipulated cells in a desired fluid or buffer, for example, to achieve a desired concentration of cells or desired buffer conditions before incubation, and / or to add dry reagents to the cell suspension. In some embodiments, the diluted cell suspension may be incubated in the container of the temperature control subsystem. In some embodiments, the incubation temperature is in the range of about 10°C to about 40°C, about 15°C to about 40°C, about 20°C to about 40°C, about 25°C to about 40°C, or about 30°C to 40°C. In some embodiments, the container is further fluid-connected to the sample sphere. In some embodiments, a fluid, such as a sample, may flow from the container into the sample sphere. In some cases, the sample sphere may be welded open so that the fluid within the sample sphere can be analyzed. In some embodiments, a temperature control subsystem includes a temperature control device configured to regulate the temperature of a diluted cell suspension. In some embodiments, the temperature control device is a heated plate. In some embodiments, the heated plate is a heated plate 70 as shown in Figure 19. In some embodiments, the plate is part of the system frame.

[0117] In some embodiments, the temperature control subsystem components may be detachably coupled to the frame of the POC system, as shown in Figures 6 to 10.

[0118] In some embodiments, the temperature control subsystem includes a weighing system for weighing the container. In some embodiments, the weighing system for weighing the container includes a tensile load cell. In some embodiments, the weighing system for weighing the container includes a compression load cell. In some embodiments, the weighing system for weighing the container includes a straight bar load cell. In some embodiments, the temperature control subsystem includes a weighing system for measuring the amount of buffer added to cells in a cell suspension containing manipulated cells. In some embodiments, the weighing system for measuring the amount of buffer includes a tensile load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a compression load cell. In some embodiments, the weighing system for measuring the amount of buffer includes a straight bar load cell. In some embodiments, the temperature control subsystem includes at least one outlet configured to be coupled to a container configured to receive a cell suspension containing manipulated cells. In some embodiments, the container is a bag including at least one inlet and at least one outlet. In some embodiments, the container is a bag such as bag 80 in Figure 20.

[0119] In some embodiments, the temperature control subsystem further includes a stirring system including a platform in contact with the container, configured to oscillate the container up and down. In some embodiments, the first stirring system including the platform in contact with the container is configured to oscillate the container up and down to agitate the cell suspension and promote homogeneity of the cell suspension containing manipulated cells or a diluted cell suspension. In some embodiments, the dilution subsystem further includes a first stirring system including a plate configured to be in contact with and moving in and out of the container, such as the stirring system shown in Figure 25. In some embodiments, the stirring system is part of the system frame.

[0120] In some embodiments, the temperature control subsystem further includes an illumination system configured to illuminate the cell suspension containing the manipulated cells and the diluted cell suspension. Such an illumination system may allow an operator to visually inspect the container for undesirable material, such as cell aggregates or debris. In some embodiments, the dilution subsystem further includes a second stirring system, which includes a platform in contact with the container and configured to oscillate the container up and down. In some embodiments, the second stirring system is configured to oscillate the container up and down to agitate the cell suspension containing the manipulated cells or the diluted cell suspension, thereby promoting homogeneity of the cell suspension containing the manipulated cells or the diluted cell suspension. In some embodiments, the dilution subsystem further includes a second stirring system, which includes a plate configured to be in contact with and move in and out of the container, such as the stirring system presented in Figure 25. In some embodiments, the stirring system is part of the system frame. In some embodiments, the dilution subsystem includes at least one pump configured to move fluid between fluid-connected subsystems.

[0121] In some embodiments, the temperature control subsystem further includes one or more of the following components: a cell aggregate filter, piping, pipe fittings, connectors, clamps, a sampling sphere, and a carboy.

[0122] ii. Reservoir In some embodiments, the systems for processing cells described herein include one or more containers in each zone of the system. In some cases, these containers may be called “reservoirs,” and the reservoirs (or containers) may have a variety of different volumes, as shown in Figures 33 and 34.

[0123] Referring to Figure 33, in some cases, Zone 1, represented as Process Reservoir 1 in Figure 33, can be any desired buffer, such as Buffer A in Figure 33, and can be injected with any desired buffer. Thus, components of Zone 1, such as cell isolation devices, TFF filter assemblies, and aggregate filters, can be injected with the desired buffer. In some cases, Zones 2, 3, and 4, represented as SQZ reservoir, SQZ output reservoir, and incubation reservoir in Figure 33, can be any desired buffer, such as Buffer B in Figure 33, and can be injected with any desired buffer. Thus, components of Zones 2-4, such as cell deformation devices and aggregate filters, can be injected with the desired buffer. In some cases, Buffer B is connected only to Zone 2, but Zone 2 is in fluid communication with Zone 3, and Zone 3 is in communication with Zone 4, thereby allowing Buffer B to be injected into each of Zones 2-4. In some embodiments, zones 5 and 6, represented as process reservoir 2 and formulation reservoir in Figure 33, can be injected with a desired buffer, such as DMSO buffer. Thus, components of zones 5 and 6, such as TFF filter assemblies and aggregate filters, can be injected with the desired buffer. In some cases, the buffer, e.g., DMSO, is in fluid communication with zone 5, and zone 5 is in fluid communication with zone 6, thereby allowing the desired buffer to be injected into both zones 5 and 6.

[0124] In some embodiments, the reservoir (container) volume is approximately 0.10L to 4.00L, 0.20L to 4.00L, 0.30L to 4.00L, 0.40L to 4.00L, 0.50L to 4.00L, 0.75L to 4.00L, 1.00L to 4.00L, 1.25L to 4.00L, and 1.50L. It could be approximately 4.00L, 1.75L to 4.00L, 2.0L to 4.00L, 2.25L to 4.00L, 2.50L to 4.00L, 2.75L to 4.00L, 3.00L to 4.00L, 3.25L to 4.00L, 3.50L to 4.00L, 3.75L to 4.00L, or approximately 4.00L.

[0125] In some embodiments, the system's reservoir may be fluidly connected to the buffer supply. In some embodiments, at least one buffer line may be connected to at least one reservoir. In some embodiments, at least one first reservoir may be connected to at least one second distinct reservoir. In some embodiments, the buffer may be transported from the first reservoir to the second reservoir. In some embodiments, the system for processing cells includes at least one, at least two, or at least three injection sectors, as shown in Figure 33.

[0126] iii. Pressure test In some embodiments, the cell processing systems described herein may undergo pressure testing of one or more subsystems. For example, pressure testing may be performed as shown in the schematic diagram of Figure 32. Referring to Figure 32, the suspension preparation subsystem may be tested in a pressure range of 10–30 psi, and components such as the cell suspension device, cell isolation device, leukocyte removal filter, and aggregate filter may be subjected to pressure testing. In some embodiments, the cell deformation subsystem may be tested at approximately 80 psi, and components such as the aggregate filter and cell deformation device may be subjected to pressure testing. In some embodiments, the dilution subsystem, incubation subsystem, cell washing subsystem, and container filling subsystem may be connected to the same line for pressure testing, as shown in Figure 32, and the pressure test may be performed at approximately 10 psi. Components such as the aggregate filter and TFF filter assembly may be subjected to pressure integrity testing. In some embodiments, pressure testing may involve the use of an electronic regulator to adjust the air pressure within the system. In some embodiments, the electronic regulator adjusts the pressure for each of the pressure sectors being tested. In some embodiments, a valve regulates the airflow to each of the pressure sectors being tested, as illustrated by the bow tie shape in Figure 32. In some embodiments, pressure testing may be used to test the integrity of each disposable kit after the kit components have been assembled into the system frame. In some embodiments, each of the cell suspension subsystem (zone 1), cell deformation subsystem (zone 2), dilution subsystem (zone 3), and container filling subsystem (zone 6) is pressure tested individually, while the incubation subsystem (zone 4) and cell washing subsystem (zone 5) are pressure tested together.

[0127] c. Cells for processing In some embodiments, the system for processing cells processes red blood cells (RBCs). In some embodiments, the system for processing cells processes peripheral blood mononuclear cells (PBMCs). In some embodiments, the system for processing cells processes activated antigen carrier (AAC) cells. In some embodiments, the system for processing cells processes tolerant antigen carrier (TAC) cells. In some embodiments, the system for processing cells processes antigen-presenting cells (APCs). In some embodiments, the system for processing cells processes T cells. In some embodiments, the system for processing cells processes B cells. In some embodiments, the system for processing cells processes macrophages. In some embodiments, the system for processing cells processes natural killer (NK) cells. In some embodiments, the system for processing cells processes dendritic cells. In some embodiments, the system for processing cells processes immune cells. In some embodiments, the system for processing cells processes monocytes. In some embodiments, the system for processing cells processes monocytic leukocytes. In some embodiments, the system for processing cells processes eosinophils. In some embodiments, the system for processing cells processes basophils. In some embodiments, the system for processing cells processes natural killer T (NKT) cells. In some embodiments, the system for processing cells processes mast cells. In some embodiments, the system for processing cells processes neutrophils. In some embodiments, the cell deformation subsystem includes one or more cell deformation contractions configured to induce perturbations in the cell membrane of a cell to allow the entry of a payload into the cell. In some embodiments, the payload includes one or more reprogramming factors. In some embodiments, the payload includes one or more nucleic acids. In some embodiments, the payload includes one or more differentiation factors. In some embodiments, the payload includes one or more neuronal reprogramming factors. In some embodiments, the system processes cells for cell therapeutics, such as cell-based therapeutics.In some embodiments, the cells for processing may include cells from a concentrated leukocyte-depleting transfusion product such as LEUKOPAK, or a similar product.

[0128] d. Process flow As described above, the point-of-care systems for processing cells described herein may include one or more subsystems, i.e., zones, for processing cells. Each subsystem may be fluidly connected to a second subsystem, thereby enabling fluid exchange between zones. In some embodiments, the systems for processing cells described herein include a suspension preparation subsystem (zone 1), a cell deformation subsystem (zone 2), a dilution subsystem (zone 3), an incubation subsystem (zone 4), a cell washing subsystem (zone 5), and a container filling subsystem (zone 6). In some embodiments, process flows for the systems described herein are shown in Figures 26 to 31, as will be further described below.

[0129] The process flow for RBCs and PBMCs may be the same in some cases, but may differ in some embodiments. For example, in some embodiments, the process flow for RBCs and PBMCs differs in the suspension preparation subsystem (zone 1) and the cell deformation subsystem. In some embodiments, the process flow for the suspension preparation subsystem for RBCs and PBMCs is the same in the dilution subsystem (zone 3), incubation subsystem (zone 4), cell washing subsystem (zone 5), and container filling subsystem (zone 6).

[0130] Referring here to Figure 26, in some embodiments, the point-of-care system process flow is the flow shown in 18000. Note that the solid black line 18008 represents a flow path. The gray hexagon 18002 represents a filter, which in some embodiments is an air filter. The black circle 18010 represents a pipe joint. The gray circle 18012 represents a pump. The black bow tie 18023 represents a valve, which in some embodiments may be a proportionally controlled pinch valve. The gray bow tie 18014 represents a valve, which in some embodiments may be a pinch valve. The white diamond 18020 represents a pressure gauge. The gray crescent 18030 represents a bubble sensor. The wavy rectangle 18050 represents a flow sensor. In some embodiments, containers such as 18036, 18040, and 18044 may be fluidly connected to zone 1 via piping. In some embodiments, container 18036 may be in contact with plate 18034, which may be a temperature-controlled plate. Container 18036 may be further fluid-connected to container 18028 via piping and to filter 18032, which may be a 40 μm aggregate filter. In some embodiments, container 18040 may be in contact with plate 18038, which may be a temperature-controlled plate. Container 18040 may be fluid-connected to container 18028 via piping. In some embodiments, container 18044 may be in contact with plate 18042, which may be a temperature-controlled plate. Container 18044 may be further fluid-connected to container 18028 via piping. Container 18028 may be in contact with plate 18026, which may be a temperature-controlled plate in some embodiments. In some embodiments, container 18028 may be in contact with a weighing system such as a compression load cell or a tension load cell. In some embodiments, the container 18028 may be in fluid communication with a cell suspension device 18022, which may be a TFF filter assembly, via piping. In some embodiments, the cell suspension device 18022 may be in fluid communication with a pressure gauge, such as a pressure gauge 18020. In some embodiments, the inlet 18024 may be used as an inlet for a pressure test zone 1 of the point of care system.In some embodiments, waste liquid from Zone 1 may flow through piping into a waste liquid container 18004 which may come into contact with a metering system 18006, which may be a tensile or compression load cell in some cases. In some embodiments, the container 18028 is in fluid communication with a filter 18016, which may be a 40 μm aggregate filter, and is further in fluid communication with Zone 2 via piping and a Zone 1 outlet 18046.

[0131] Referring further to Figure 26, Zone 1 may be in fluid communication with Zone 2 via Zone 1 Outlet 18046, which serves as an inlet for Zone 2, allowing fluid, such as cells suspended in a delivery medium, to flow from Zone 1 to Zone 2. Zone 1 Outlet 18046 may be in fluid communication with container 18062 via piping. In some embodiments, container 18062 may be in contact with plate 18060, such as a temperature-controlled plate for heating, cooling, or maintaining a relatively constant temperature. In some embodiments, container 18066 may be in contact with plate 18064, which may be a temperature-controlled plate. In some embodiments, container 18066 may be in fluid communication with container 18062 via piping. In some embodiments, container 18062 may be in fluid communication with container 18058 via piping. In some embodiments, container 18058 may be in contact with plate 18056, such as a temperature-controlled plate for heating, cooling, or maintaining a relatively constant temperature. In some embodiments, the container 18058 is in fluid communication with a filter 18052, such as a 40 μm aggregate filter, and further in fluid communication with a cell deformation device 18048 via piping. In some embodiments, the inlet 18054 is used for pressure testing in zone 2. In some embodiments, the cell deformation device 18048 is in fluid communication with zone 3 via a zone 2 outlet 18059, which may also be the inlet for zone 3.

[0132] Referring further to Figure 26, in some embodiments, the zone 3 inlet 18059 may be in fluid communication with container 18072 via piping. Container 18072 may be in contact with a metering system 18070, which may be, for example, a tensile load cell or a compression load cell. Container 18078 may be in contact with a plate 18076, which may be a temperature-controlled plate. Container 18078 may be in fluid communication with container 18072 via piping. In some embodiments, container 18082 is in contact with a plate 18080, which may be a temperature-controlled plate. Container 18082 may be in fluid communication with container 18072 via piping. In some embodiments, the inlet 18074 is in fluid communication with container 18072. In some embodiments, container 18072 is in fluid communication with an aggregate filter 18068, which may be a 40 μm aggregate filter, and further in fluid communication with the zone 3 outlet 18075 via piping. In some embodiments, the Zone 3 outlet 18075 may also be the Zone 4 inlet. In some embodiments, the Zone 4 inlet 18075 is in fluid communication with a vessel 18088 via piping. In some embodiments, the vessel 18088 is in contact with a plate 18086, which may be a temperature-controlled plate. In some embodiments, the inlet 18090 may be used for pressure testing of Zones 4 and 3. In some embodiments, the vessel 18088 is in fluid communication with a filter 18084, such as a 40 μm aggregate filter, and is further in fluid communication with the Zone 4 outlet 18092 via piping. In some embodiments, the Zone 4 outlet 18092 may also be the Zone 5 inlet. In some embodiments, the Zone 4 outlet 18092 is in fluid communication with a vessel 18100 via piping. In some embodiments, the vessel 18100 may be in contact with a plate 18098, which may be a temperature-controlled plate. In some embodiments, container 18106 is in contact with plate 18104, which may be a temperature-controlled plate. In some embodiments, container 18110 may be in contact with plate 18108, which may be a temperature-controlled plate. In some embodiments, inlet 18102 may be used for a pressure test zone 5. In some embodiments, container 18100 is in fluid communication with a cell suspension device 18094, which may be a TFF filter assembly. In some embodiments, the cell suspension device 18094 is in fluid communication with container 18100 via piping.In some embodiments, the container 18100 is in fluid communication with a filter 18096, which may be a 40 μm aggregate filter, and is further fluidly connected to a zone 5 outlet 18112 via piping. In some embodiments, the zone 5 outlet 18112 is also the inlet for zone 6.

[0133] In some embodiments, the outlet 18112 of zone 5 is in fluid communication with vessel 18126 via piping. In some embodiments, vessel 18126 is in contact with a metering system 18122. In some embodiments, vessel 18126 may further be in contact with backlight illumination 18124. In some embodiments, vessel 18134 is in contact with plate 18132, which may be a temperature-controlled plate. In some embodiments, vessel 18134 is in fluid communication with vessel 18126 via piping. In some embodiments, vessel 18126 is in fluid communication with filter 18120, which may be a 40 μm aggregate filter. In some embodiments, the inlet 18118 may be in fluid communication with filter 18120. In some embodiments, the inlet 18118 may be used for pressure test zone 6. In some embodiments, vessel 18126 is in fluid communication with vessel 18130 via piping. In some cases, container 18130 comes into contact with the metering system 18128. In some embodiments, waste liquid from zone 5 or zone 6 may flow through piping into a waste liquid container 18116 that may come into contact with the metering system 18114.

[0134] Referring here to Figure 35, in some embodiments, the point-of-care system process flow is the flow shown at 19000. Note that the solid black line 19012 represents the flow path. The gray hexagon 19002 represents a filter, which in some embodiments is an air filter. The black circle 19008 represents a pipe joint. The gray circle 19010 represents a pump. The black bow tie 19014 represents a valve, which in some embodiments may be a proportionally controlled pinch valve. The gray bow tie 19016 represents a valve, which in some embodiments may be a pinch valve. The white diamond 19018 represents a pressure gauge. The gray crescent 19032 represents a bubble sensor. The wavy rectangle 19052 represents a flow sensor. In some embodiments, the container 19036 contacts a plate 19038, which may be a temperature-controlled plate. In some embodiments, container 19036 is in fluid communication with a cell isolation device 19034, such as a leukocyte removal filter, and further in fluid communication with container 19026 via piping. In some embodiments, container 19026 is in contact with a metering system 19024. In some embodiments, inlet 19030 is in fluid communication with container 19026 via piping. In some embodiments, inlet 19028 may be used for a pressure test zone 1. In some embodiments, inlet 19021 is in fluid communication with container 19026 via piping. Container 19026 may further in fluid communication with a cell suspension device 19020, such as a TFF filter assembly. In some embodiments, container 19040 is in contact with a plate 19042, which may be a temperature-controlled plate. Container 19040 may be in fluid communication with container 19026 via piping. In some embodiments, container 19046 is in contact with plate 19044, which may be a temperature-controlled plate. Container 19046 may be in further fluid communication with container 19026 via piping. In some embodiments, container 19026 is in fluid communication with aggregate filter 19022, which may be a 40 μm aggregate filter, and is in further fluid communication with zone 1 outlet 19048 via piping. In some embodiments, waste liquid from zone 1 may flow through piping into waste liquid container 19006, which may be in contact with metering system 19004.

[0135] Referring further to Figure 35, the Zone 1 outlet 19048 may be the Zone 2 inlet. In some embodiments, the Zone 1 outlet 19048 is in fluid communication with container 19064. Container 19064 may be in contact with plate 19062, which may be a temperature-controlled plate. In some embodiments, container 19070 is in contact with plate 19068, which may be a temperature-controlled plate. In some embodiments, container 19070 is in fluid communication with container 19064 via piping. In some embodiments, container 19064 is in fluid communication with container 19058 via piping. In some embodiments, container 19058 is in contact with plate 19060, which may be a temperature-controlled plate. In some embodiments, the inlet 19056 may be used for pressure testing Zone 3. In some embodiments, container 19058 is in fluid communication with filter 19054, such as a 40 μm aggregate filter, and with cell deformation device 19050 via piping. In some embodiments, the cell deformation device 19050 is in fluid communication with a zone 2 outlet 19066, which may be an inlet to zone 3. In some embodiments, the inlet 19066 is in fluid communication with a container 19076 via piping. In some embodiments, the container 19076 is in contact with a metering system 19074. In some embodiments, the inlet 19078 is in fluid communication with the container 19076 via piping. In some embodiments, the container 19076 is in fluid communication with a filter 19072, such as a 40 μm aggregate filter, and is further fluidly connected to a zone 3 outlet 19079 via piping. In some embodiments, the zone 3 outlet 19079 may be an inlet to zone 4. In some embodiments, the zone 3 outlet 19079 is in fluid communication with a container 19084 via piping. In some embodiments, the container 19084 is in contact with a plate 19082, which may be a temperature-controlled plate. In some embodiments, the inlet 19086 may be used for pressure testing of zones 4 and 3. In some embodiments, the vessel 19084 is in fluid communication with a filter 19080, such as a 40 μm aggregate filter, and further in fluid communication with the zone 4 outlet 19088 via piping. The zone 4 outlet 19088 may be an inlet to zone 5. In some embodiments, the zone 4 outlet 19088 is in fluid communication with a vessel 19098. The vessel 19098 may be in contact with a metering system 19096.In some embodiments, container 19098 is in fluid communication with container 19098 via piping. In some embodiments, container 19104 is in contact with plate 19102, which may be a temperature-controlled plate. In some embodiments, container 19108 is in contact with plate 19106, which may be a temperature-controlled plate. In some embodiments, container 19108 is in fluid communication with container 19098 via piping. In some embodiments, inlet 19100 may be used for pressure test zone 5. In some embodiments, container 19098 is in fluid communication with cell suspension device 19090, such as a TFF filter assembly, via piping. In some embodiments, inlet 19092 is in fluid communication with a container having the cell suspension device 19090 via piping. In some embodiments, container 19098 is in further fluid communication with aggregate filter 19094, which may be a 40 μm aggregate filter, and further fluid communication with zone 5 outlet 19110.

[0136] In some embodiments, the Zone 5 outlet 19110 is in fluid communication with container 19124 via piping. In some embodiments, container 19124 is in contact with a metering system 19120. In some embodiments, container 19124 may further be in contact with backlight illumination 19122. In some embodiments, container 19132 is in contact with a plate 19130, which may be a temperature-controlled plate. In some embodiments, container 19132 is in fluid communication with container 19124 via piping. In some embodiments, container 19124 is in fluid communication with a filter 19118, which may be a 40 μm aggregate filter. In some embodiments, the inlet 19116 may be in fluid communication with the filter 19118. In some embodiments, the inlet 19116 may be used for pressure test zone 6. In some embodiments, container 19124 is in fluid communication with container 19128 via piping. In some cases, container 19128 comes into contact with the metering system 19126. In some embodiments, waste liquid from zone 5 or zone 6 may flow through piping into a waste liquid container 19114 that may come into contact with the metering system 19112.

[0137] Referring here to Figure 27, in some embodiments, the PBMC process flow is the flow presented in 12000. Flow path 12006 is represented by a solid black line, an optional flow path 12021 is represented by a dashed black line, and the wastewater flow path 12004 is represented by a dashed gray line. The solid black circle 12002 represents a switch valve. The gray bow tie 12010 represents a valve. The gray circle 12012 represents a pump. In some embodiments, containers such as 12030, 12038, 12044, and 12048 may be fluidly connected to zone 1 via piping and pumps, such as pump 12028 for container 12030 or pump 12012 for container 12038. In some embodiments, container 12030 may be in contact with plate 12033, which may be a temperature-controlled plate. In some embodiments, container 12030 may be detachably coupled to a fastening element 12032, which may be a hook and may also function as part of a weighing system. In some embodiments, a sample from container 12030 may be pumped through piping to an aggregate filter 12029 by a pump 12028. The aggregate filter 12029 may be in fluid communication with container 12024 by piping and valves. In some embodiments, container 12038 is in contact with a plate 12036, which may be a temperature-controlled plate. In some embodiments, container 12038 is detachably coupled to a fastening element 12034, which may function as part of a weighing system. In some embodiments, container 12044 is in contact with a plate 12042, which may be a temperature-controlled plate. In some embodiments, container 12044 is detachably coupled to a fastening element 12046, which may function as part of a weighing system. In some embodiments, container 12044 is in contact with a plate 12040. In some embodiments, the container 12050 is in contact with a plate 12048, which may be a temperature-controlled plate. In some embodiments, the container 12050 is removably coupled to a fastening element 12052, which may function as part of a metering system. In some embodiments, the containers 12038, 12044, and 12050 are fluidly connected to zone 1 via piping and valves.In some embodiments, container 12050 is further fluidly connected to zone 2 via piping and valves. In some embodiments, each of containers 12038, 12044, and 12050 is in fluid communication with container 12024 via piping and valves. In some embodiments, container 12024 is in fluid communication with sorting chips 12020 and TFF filter assembly 12018 via piping and valves. Fluid from container 12024 can be pumped across zone 1 by pump 12012. In some embodiments, TFF filter assembly 12018 is in fluid communication with pressure gauge 12008, bubble sensor 12026, and additionally flow sensor. In some embodiments, zone 1 includes one or more sorting chips 12020, such as two sorting chips 12020. In some embodiments, the container 12022 is in fluid communication with the aggregate filter 12016 and the flow sensor 12014, and further in fluid communication with zone 2 via the outlet 12054.

[0138] Referring here to Figure 28, in some embodiments, the RBC process flow is presented as 13000. The flow path 13006 is represented by a solid black line, the optional flow path 13021 is represented by a dashed black line, and the wastewater path 13004 is represented as a dashed gray line. The solid black circle 13002 represents a switch valve. The gray bow tie 13012 represents a valve. The gray circle 13010 represents a pump. The crescent 13024 represents a bubble sensor. The black wavy rectangle 13008 represents a pressure gauge. The gray wavy rectangle 13018 represents a flow sensor. In some embodiments, the container 13030 is in contact with a plate 13031, which may be a temperature-controlled plate. In some embodiments, the container 13030 is detachably coupled to a fastening element 13032, which may also function as part of a metering system. In some embodiments, container 13030 is in fluid communication with pump 13028. In some embodiments, container 13030 is in fluid communication with aggregate filter 13026 and is further fluidly connected to container 13022. Container 13036 may be in contact with plate 13034, which may be a temperature-controlled plate. Container 13038 may be detachably coupled to fastening element 13038, which may be a hook in some cases. In some embodiments, container 13042 is in contact with plate 13040, which may be a temperature-controlled plate. In some embodiments, container 13042 is detachably coupled to fastening element 13044. In some embodiments, container 13048 is in contact with plate 13046, which may be a temperature-controlled plate. In some embodiments, container 13048 is detachably coupled to fastening element 13050. In some embodiments, each of the containers 13030, 13036, 13042, and 13048 is in fluid communication with Zone 1. In some embodiments, container 13048 is in further fluid communication with Zone 2. In some embodiments, container 13033 is in contact with plate 13020, which may be a temperature-controlled plate. In some embodiments, container 13022 is in fluid communication with TFF filter 13016. Container 13022 is in further fluid communication with leukocyte removal filter 13014 and also in further fluid communication with Zone 2 via outlet 13019.

[0139] Referring here to Figure 29, in several embodiments, PBMC process flows 14000 to zones 2 and 3 are presented. Note that the crescent 14016 represents a bubble sensor, the solid black circle 14002 represents a switch valve, the bow tie 14004 represents a valve, and the gray wavy triangle 14010 represents a flow sensor. The solid black line 14008 represents a flow path. The dashed black line 14012 represents an optional flow path. Fluid from zone 1 may enter zone 2 via an inlet 14015 that is in fluid communication with a container 14018. The container 14018 may be in contact with a plate 14020, which may be a temperature-controlled plate. The container 14018 is further in fluid communication with a cell aggregate filter 14014 and further in fluid communication with a cell deformation device 14006, such as a microfluidic chip for cell deformation. In some embodiments, the cell deformation device 14006 is in fluid communication with zone 3 via the outlet 14022. Referring to zone 3, the solid black line 14024 represents a flow path, the solid black circle 14030 represents a switch valve, the gray wavy rectangle 14026 represents a flow sensor, the gray dotted line 14032 represents an optional flow path, the bow tie 14035 represents a valve, the crescent 14036 represents a bubble sensor, and the gray circle 14028 represents a pump. In some embodiments, fluid from zone 2 enters zone 3 via the outlet 14022 from zone 2, which serves as an inlet for zone 3, and flows into a container 14038. The container 14038 is in contact with a plate 14040, which may be a temperature-controlled plate. In some embodiments, container 14038 is in fluid communication with aggregate filter 14034 and further in fluid communication with zone 4 via outlet 14037. Furthermore, container 14046 may be in fluid communication with zone 3 so that fluid from container 14046 can be pumped into zone 3 by pump 14042. In some embodiments, container 14046 is detachably coupled to fastening element 14044. In some embodiments, container 14050 is in fluid communication with zone 3 so that pump 14052 can pump fluid from container 14050 into zone 3. In some embodiments, container 14050 is detachably coupled to fastening element 14048.

[0140] Referring here to Figure 30, in several embodiments, RBC process flows 15000 for zones 2 and 3 are presented. Note that the crescent 15016 represents a bubble sensor, the solid black circle 15002 represents a switch valve, the bow tie 15004 represents a valve, and the gray wavy triangle 15010 represents a flow sensor. The solid black line 15008 represents a flow path. The dashed black line 15012 represents an optional flow path. Fluid from zone 1 may enter zone 2 via an inlet 15015 which is in fluid communication with a container 15018. The container 15018 may be in contact with a plate 15020 which may be a temperature-controlled plate. The container 15018 is further in fluid communication with a cell aggregate filter 15014 and further in fluid communication with a cell deformation device 15006 such as a microfluidic chip for cell deformation. In some embodiments, the cell deformation device 15006 is in fluid communication with zone 3 via the outlet 15022. Referring to zone 3, the solid black line 15024 represents a flow path, the solid black circle 15030 represents a switch valve, the gray wavy rectangle 15026 represents a flow sensor, the gray dotted line 15032 represents an optional flow path, the bow tie 15035 represents a valve, the crescent 15036 represents a bubble sensor, and the gray circle 15028 represents a pump. In some embodiments, fluid from zone 2 enters zone 3 via the outlet 15022 from zone 2, which serves as an inlet for zone 3, and flows into a container 15038. The container 15038 is in contact with a plate 15040, which may be a temperature-controlled plate. In some embodiments, the container 15038 is in fluid communication with the aggregate filter 15034 and further in fluid communication with zone 4 via the outlet 15037. In some embodiments, the container 15044 may be in fluid communication with zone 3 so that fluid from the container 15044 can be pumped into zone 3 by the pump 15042. In some embodiments, the container 15044 is in contact with a plate 15046, which may be a temperature-controlled plate. In some embodiments, the container 15044 may be in contact with a meter 15048.

[0141] Referring here to Figure 31, in some embodiments, the PBMC or RBC process flow may be process flow 16000. Referring to Zone 4, the solid black line 16002 represents a flow path, the solid black circle 16004 represents a switch valve, the gray circle 16008 represents a pump, the gray wavy rectangle 16006 represents a flow sensor, the bow tie 16014 represents a valve, the crescent 16018 represents a bubble sensor, and the dashed black line 16010 represents an optional flow path. In some embodiments, the fluid enters Zone 4 from Zone 3 16016 through the inlet and flows into container 16022. Container 16022 is in contact with plate 16020, which may be a temperature-controlled plate. In some embodiments, the container 16022 is in fluid communication with the aggregate filter 16012 and further in fluid communication with zone 5 via the zone 4 outlet / zone 5 inlet 16024.

[0142] Referring to Zone 5, the solid black circle 16026 represents a switch valve, the wavy black rectangle 16032 represents a pressure gauge, the crescent 16042 represents a bubble sensor, the gray circle 16034 represents a pump, the bow tie 16040 represents a valve, the wavy gray rectangle 16043 represents a flow sensor, the solid black line 16030 represents a flow path, and the dashed gray line 16028 represents a wastewater flow path. In some embodiments, fluid from Zone 4 enters Zone 5 through the inlet 16024 and flows into container 16046. Container 16046 is in contact with plate 16044, which may be a temperature-controlled plate. In some embodiments, container 16044 is in fluid communication with TFF filter assembly 16038. Furthermore, the container 16044 is in fluid communication with the aggregate filter 16036 and further in fluid communication with Zone 6 via the outlet / inlet 16076 to Zone 6.

[0143] Referring to Zone 6, the gray dashed line 16058 represents the wastewater flow path, the black solid line 16078 represents the flow path, the black dashed line 16064 represents an optional flow path, the gray circle 16068 represents the pump, the black circle 16060 represents the switch valve, the bow tie 16070 represents the valve, and the crescent moon 16084 represents the bubble sensor. In some embodiments, fluid from Zone 5 enters Zone 6 via the inlet 16076 to Zone 6 and flows into container 16080. In some embodiments, container 16080 is in contact with plate 16082, which may be a temperature-controlled plate. In some embodiments, container 16080 is in fluid communication with aggregate filter 16066. In some embodiments, container 16080 is in fluid communication with container 16074. Each container 16074 may be placed on a plate 16072, which may each be a temperature-controlled plate and may also be part of a metering system. Furthermore, zone 6 may include a wastewater tank 16062 into which wastewater flows along a wastewater channel 16058. The wastewater tank 16062 may be in contact with the metering system. In some embodiments, the metering system includes a tensile load cell or a compression load cell. Further referring to Figure 31, container 16054 may be in contact with a plate 16052, which may be a temperature-controlled plate. In some embodiments, container 16054 is fluid-connected to zones 5 and 6, so that pump 16050 can pump fluid into either zone, as directed by a switch valve 16048. In some embodiments, container 16054 is in contact with meter 16056.

[0144] 2. Methods for processing cells In some embodiments, the disclosure generally relates to a method for processing cells, wherein the method is carried out by a system comprising one or more of the following subsystems: a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, wherein the method comprises: i. in the suspension preparation subsystem: 1. receiving cells from a container; 2. optionally performing a cell isolation operation on the cells to produce isolated cells; 3. receiving a delivery medium via a delivery medium inlet; and 4. producing a cell suspension by suspending cells or isolated cells in the delivery medium to produce a cell suspension; and ii. in the cell deformation subsystem: 1. receiving a flow of cell suspension from the suspension preparation subsystem; and 2. inducing perturbations in the cell membranes of the cells to allow the entry of a payload into the cells. The system includes: iii. a dilution subsystem that: 1. receives a flow of cell suspension containing manipulated cells from the cell deformation subsystem; 2. receives a fluid or dry reagent via a buffer inlet; and iv. an incubation subsystem that: 1. receives a flow of incubated cell suspension from the incubation subsystem; 2. performs a buffer exchange operation on the cells to suspend them in a storage medium; and vi. a container filling subsystem that: receives a flow of cells suspended in a storage medium from the cell washing subsystem; and introduces the cells suspended in the storage medium into one or more containers.

[0145] In some embodiments, the system for processing cells processes red blood cells (RBCs). In some embodiments, the system for processing cells processes peripheral blood mononuclear cells (PBMCs). In some embodiments, the system for processing cells processes activated antigen carrier (AAC) cells. In some embodiments, the system for processing cells processes tolerant antigen carrier (TAC) cells. In some embodiments, the system for processing cells processes antigen-presenting cells (APCs). In some embodiments, the system for processing cells processes T cells. In some embodiments, the system for processing cells processes B cells. In some embodiments, the system for processing cells processes macrophages. In some embodiments, the system for processing cells processes natural killer (NK) cells. In some embodiments, the system for processing cells processes dendritic cells. In some embodiments, the system for processing cells processes immune cells. In some embodiments, the system for processing cells processes monocytes. In some embodiments, the system for processing cells processes monocytic leukocytes. In some embodiments, the system for processing cells processes eosinophils. In some embodiments, the system for processing cells processes basophils. In some embodiments, the system for processing cells processes natural killer T (NKT) cells. In some embodiments, the system for processing cells processes mast cells. In some embodiments, the system for processing cells processes neutrophils. In some embodiments, the method removes coagulated plasma and / or serum. In some embodiments, the method is carried out for about 5 to about 7 hours. In some embodiments, the method produces a cell therapy drug. In some embodiments, the payload contains one or more reprogramming factors. In some embodiments, the payload contains one or more nucleic acids. In some embodiments, the payload contains one or more differentiation factors. In some embodiments, the payload contains one or more neuronal reprogramming factors. In some embodiments, the cell isolation operation is not carried out as part of the method for processing cells.In some embodiments, the cell isolation procedure is not performed as part of the process for processing TACs. In some embodiments, the cell isolation procedure is not performed as part of the process for processing APCs. In some embodiments, the cell isolation procedure is not performed as part of the process for processing PBMCs.

[0146] 3. Kit In some embodiments, the present disclosure generally relates to a kit for use in a system for processing blood, the kit comprising: i. a first kit comprising detachably connectable components configured to be detachably connected to a frame of a suspension preparation subsystem of the system, wherein the first set of detachably connectable components comprises 1. a cell isolation device, or 2. a tangential flow filtration membrane assembly; ii. a second kit comprising detachably connectable components configured to be detachably connectable to a frame of a cell deformation subsystem of the system, wherein the second set of detachably connectable components comprises one or more microfluidic chips comprising one or more cell deformation contractions that can compel cells to undergo perturbations of the cell membrane; and iii. a third kit comprising detachably connectable components configured to be detachably connectable to a frame of a dilution subsystem of the system, wherein the detachably connectable components The system comprises one or more of the following: a third kit in which the third set of components includes at least one cell aggregate filter; a fourth kit in which detachably connectable components configured to be detachably connectable to the frame of the incubation subsystem of the system, wherein the fourth set of detachably connectable components includes at least one cell aggregate filter; a fifth kit in which detachably connectable components configured to be detachably connectable to the frame of the cell washing subsystem of the system, wherein the fifth set of detachably connectable components includes a second tangential flow filtration membrane assembly; and a sixth kit in which detachably connectable components configured to be detachably connectable to the frame of the container filling subsystem of the system, wherein the sixth set of detachably connectable components includes at least one cell aggregate filter.

[0147] In some embodiments, one or more detachable connectable components of a second kit of detachable connectable components are configured to be fluidically connected to one or more detachable connectable components of a first kit of detachable connectable components. In some embodiments, one or more detachable connectable components of a third kit of detachable connectable components are configured to be fluidically connected to one or more detachable connectable components of a second set of detachable connectable components. In some embodiments, one or more detachable connectable components of a fourth kit of detachable connectable components are configured to be fluidically connected to one or more detachable connectable components of a third kit of detachable connectable components. In some embodiments, one or more detachable connectable components of a fifth kit of detachable connectable components are configured to be fluidically connected to one or more detachable connectable components of a fourth kit of detachable connectable components. In some embodiments, one or more detachable connectable components of a sixth kit of detachable connectable components are configured to be fluidically connected to one or more detachable connectable components of a fifth kit of detachable connectable components.

[0148] In some embodiments, one or more components of the kit are configured to be fluidly connected to one or more components of corresponding subsystems of the system. In some embodiments, the kit includes first, second, third, fourth, fifth, and sixth kits. In some embodiments, each kit is packaged separately. In some embodiments, at least two kits are packaged together.

[0149] In some embodiments, a disposable kit for use with a point-of-care system includes the disposable kit 7000 shown in Figure 9. Referring here to Figure 9, in some cases, the kit 7000 includes input lines 7028, 7030, and 7034, which can be used, for example, to deliver a delivery medium, buffer, or sample. Inlets 7028 and 7030 may be fluidly connected to a container 7016 via piping and inlet 7015. In some embodiments, inlet 7034 is fluidly connected to an aggregate filter 7032, which is fluidly connected to the container 7016. In some embodiments, the container 7016 is fluidly connected to a TFF filter assembly 7004 via piping. In some embodiments, the kit 7000 includes a TFF filter assembly 7004, which is fluidly connected to a cell isolation device 7014 via piping 7002. In some embodiments, the cell isolation device 7014 is fluidly connected to the container 7016 via outlet 7013 and piping. Container 7016 may be fluidly connected to an aggregate filter 7006, which is fluidly connected to container 7038 via piping. In some embodiments, container 7038 includes an inlet 7037 and an outlet 7035 which are coupled to piping. In some embodiments, container 7038 is fluidly connected to an aggregate filter 7036, which is further connected to a cell deformation device 7018 via outlet 7035 and piping. In some embodiments, the cell deformation device 7018 is fluidly connected to container 7020 via piping. Inlets 7040 and 7042 are fluidly connected to container 7020 via piping and inlet 7023. Container 7020 is fluidly connected to an aggregate filter 7008, which is fluidly connected to container 7044 via piping and inlet 7043, via outlet 7021 and piping. In some embodiments, the vessel 7044 is fluidly connected to an aggregate filter 7022 via an outlet 7041 and piping, which is fluidly connected to the vessel 7024 via piping and an inlet 7025. In some embodiments, the inlet 7009 is fluidly connected to the vessel 7024 via piping and an inlet 7025. In some embodiments, the vessel 7024 is fluidly connected to a TFF assembly 7010 via an outlet 7025' and piping, which can be further fluidly connected to the vessel 7024 via piping and an inlet 7025.In some embodiments, container 7024 may be fluidly connected via outlet 7024 to an aggregate filter 7012, which is fluidly connected to container 7046 via piping and inlet 7047. In some embodiments, container 7047 may be fluidly connected via outlet 7045' to an aggregate filter 7048, which is fluidly connected to container 7046 via piping and inlet 7047. Container 7046 may be further fluidly connected to an outer container 7026 via outlet 7045 and piping. In some embodiments, kit 7000 may further include a waste liquid container 7001 which is coupled to piping.

[0150] In contrast, in some embodiments, separate kits may be used for each subsystem, as discussed below. Each kit includes components that are detachably coupled to the system frame and can be further coupled to the corresponding subsystems of the system.

[0151] In some embodiments, disposable kits for use with a point-of-care system are sterile. In some embodiments, disposable kits for use with a point-of-care system are assembled in a cleanroom environment. In some embodiments, gamma radiation is used to sterilize disposable kits for use with a point-of-care system. In some embodiments, ethanol is used to sterilize disposable kits for use with a point-of-care system. In some embodiments, disposable kits for use with a point-of-care system are packaged in a cleanroom. In some embodiments, disposable kits for use with a point-of-care system are packaged in thermoformed trays.

[0152] a. Suspension preparation subsystem (Zone 1) In some embodiments, the Zone 1 kit includes the components discussed above, as shown in Figure 12 or Figure 13.

[0153] In some embodiments, the kit includes a first kit comprising a suspension preparation subsystem kit. In some embodiments, the first kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the first kit of detachably connectable components includes a leukocyte removal filter. In some embodiments, the first kit of detachably connectable components includes a container. In some embodiments, the first kit of detachably connectable components includes a tangential flow filtration filter assembly. In some embodiments, the first kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, carboys, air filters, and tangential flow filtration filter assemblies.

[0154] b. Cell deformation subsystem (Zone 2) In some embodiments, the Zone 2 kit includes the components discussed above, as shown in Figure 14.

[0155] In some embodiments, the kit includes a second kit comprising a cell deformation subsystem. In some embodiments, the second kit of detachably connectable components includes a rigid sample container. In some embodiments, the second kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the second kit of detachably connectable components includes a preparation container. In some embodiments, the second kit of detachably connectable components includes one or more microfluidic chips. In some embodiments, the second kit of detachably connectable components includes one or more microfluidic chip cartridges. In some embodiments, the second kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, containers, bags, air filters, and barrel filters.

[0156] c. Dilution subsystem (Zone 3) In some embodiments, the Zone 3 kit includes the components discussed above, as shown in Figure 15.

[0157] In some embodiments, the kit includes a third kit comprising a dilution subsystem. In some embodiments, the third kit of detachably connectable components includes a container. In some embodiments, the third kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the third kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys.

[0158] d. Incubation subsystem (Zone 4) In some embodiments, the Zone 4 kit includes the components discussed above, as shown in Figure 16.

[0159] In some embodiments, the kit includes a fourth kit comprising a culture subsystem. In some embodiments, the fourth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the fourth kit of detachably connectable components includes one or more containers. In some embodiments, the fourth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, and clamps.

[0160] e. Cell washing subsystem (Zone 5) In some embodiments, the Zone 5 kit includes the components discussed above, as shown in Figure 17.

[0161] In some embodiments, the kit includes a fifth kit comprising a cell washing subsystem. In some embodiments, the fifth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the fifth kit of detachably connectable components includes a container. In some embodiments, the fifth kit of detachably connectable components includes a tangential flow filtration filter assembly. In some embodiments, the fifth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, sampling bulbs, air filters, and carboys.

[0162] f. Container filling subsystem (Zone 6) In some embodiments, the Zone 6 kit includes the components discussed above, as shown in Figure 18.

[0163] In some embodiments, the kit includes a sixth kit comprising a cell-filling subsystem. In some embodiments, the sixth kit of detachably connectable components includes at least one container. In some embodiments, the container is a cryopreservation bag. In some embodiments, the sixth kit of detachably connectable components includes a cell aggregate filter. In some embodiments, the sixth kit of detachably connectable components includes one or more of piping, pipe fittings, connectors, clamps, and sampling spheres.

[0164] g. Temperature control subsystem In some embodiments, the temperature-controlled subsystem kit includes components for performing functions otherwise carried out by the dilution subsystem (zone 3) and the culture subsystem (zone 4). For example, the temperature-controlled subsystem kit may include a container for receiving a cell suspension containing cells manipulated from the cell deformation subsystem (zone 2), and one or more inlets for introducing fluids and / or dry reagents into the container containing the cell suspension. The container may come into contact with a temperature-controlled plate when mounted on the system frame.

[0165] In some embodiments, the kit includes a temperature control subsystem kit. In some embodiments, the temperature control kit includes a container. In some embodiments, the temperature control kit components include a cell aggregate filter. In some embodiments, the temperature control kit includes one or more of the following: piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys.

[0166] h. Waste liquid container kit In some embodiments, the kit includes a waste liquid container kit, such as a waste liquid container kit 9000. Referring here to Figure 11, the waste liquid container kit 900 includes a waste liquid container 9002 which is coupled to a cap 9004. The cap 9004 may be coupled to a pipe 9006, such as through an outlet 9005.

[0167] i. Kit installation In some embodiments, a kit containing one or more kits for zones 1 through 6 may be packaged in an accordion tray package. In some embodiments, such a kit may be installed by coupling it to a system frame, as shown in Figure 22. Referring to Figure 22, kit 10002 may be detachably coupled to frame 10004, for example, by attaching a panel to a device containing one or more subsystems. The remaining subsystems may be installed by unfolding the accordion tray kit 10002. For example, a subsystem configured on panel 10006 of kit 10002 may be detachably connected to frame 10004 by unfolding the accordion tray. In some embodiments, panel 10010 may contain one or more subsystems that can be connected to frame 10004 by unfolding the panel.

[0168] In some embodiments, a kit containing one or more kits for zones 1 through 6 may be packaged as a rewound sheet. In some embodiments, such a kit may be installed by coupling it to a frame of the system, as shown in Figure 23. Referring to Figure 23, the rewound sheet 11002 may be removably coupled to a frame 110004 by unwinding the sheet and connecting it to the frame 11004. For example, a partially unwinded sheet 11006 may be further unwinded to become a fully unwinded sheet 11008 and then connected to the frame.

[0169] 4. Further systems for processing cells A system for processing cells is provided. The system may be configured to receive a cell input, such as in the form of blood or cells suspended in any suitable medium, and to automatically process the cells to perform one or more operations on the cells, such as isolation, suspension in one or more mediums, buffer exchange, payload delivery, dilution, incubation, container filling, temperature control, filtration, agitation, pressurization, and / or cryopreservation.

[0170] In some embodiments, the system may include multiple subsystems, each subsystem configured to perform one or more cell processing operations. In some embodiments, one or more subsystems may be in fluid communication with one or more other subsystems so that a fluid (e.g., cell suspension) can flow automatically from one subsystem to the next (e.g., by gravity and / or under pressure). In some embodiments, the system may be configured to pass the cells being processed from one subsystem to the next (and / or from system input to system output) without manual (e.g., physical) human intervention. In some embodiments, one or more subsystems may be configured to be controlled independently of one or more other subsystems (e.g., temperature, pressure, etc.).

[0171] In some embodiments, the system may include a suspension preparation subsystem configured to receive cells, perform cell isolation operations on the cells, receive a delivery medium, and / or create a cell suspension by suspending the cells in the delivery medium. The cells suspended in the delivery medium may then flow from the suspension preparation subsystem to a cell deformation subsystem.

[0172] In some embodiments, the cell deformation subsystem may be configured to flow a cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membrane of the cells, thereby allowing the payload to enter the cells. After the perturbation and / or delivery of the payload, the cell suspension may then flow from the cell deformation subsystem to the dilution subsystem.

[0173] In some embodiments, the dilution subsystem may be configured to create a diluted cell suspension by mixing the received cell suspension with a buffer. After dilution of the cell suspension, the diluted cell suspension may flow from the dilution subsystem to the incubation subsystem.

[0174] In some embodiments, the incubation subsystem may be configured to control the temperature of the diluted cell suspension, for example, by raising, lowering, or maintaining the temperature. After incubation, the diluted cell suspension may then flow from the incubation subsystem to the cell washing subsystem.

[0175] In some embodiments, the cell washing subsystem may be configured to perform a buffer exchange operation on the accepted diluted cell suspension and to suspend the cells in a medium such as a cryoprotective medium. The cells suspended in the medium can then flow from the cell washing subsystem to the container filling subsystem.

[0176] In some embodiments, the container filling subsystem may be configured to flow cells suspended in a medium into one or more bags for storage and / or preservation, such as cryopreservation.

[0177] In some embodiments, one or more operations of the system may be configured to be automatically controlled and may be controlled by one or more processors of the system. A user may perform input to one or more user interfaces to control the functions of the system.

[0178] In some embodiments, the system may be configured to accept one or more disposable components, which may be inserted into the system, attached to the system, or otherwise configured for use with the system. One or more of the disposable components may form all or part of a fluid channel for cells flowing through the system. The disposable components may be configured for use with specific subsystems of the system. The disposable components may be supplied as one or more kits. In some embodiments, the disposable components may include one or more helical inertia separation microfluidic consumables, a tangential flow filtration membrane assembly, a cell aggregate filter, a leukocyte removal filter, a sampling bulb, piping, pipe fittings, connectors, clamps, bags, carboys, rigid sample containers, components including one or more cell deformation contractions that can compel cells to cause perturbations of the cell membrane, and cartridges for containing fluid and delivering fluid to and from the components, which include one or more cell deformation constructs.

[0179] The following exemplary embodiments are provided.

[0180] 1. A system for processing cells, the system is A suspension preparation subsystem configured to receive cells, perform cell isolation operations on the cells, receive a delivery medium, and create a cell suspension by suspending the cells in the delivery medium, A cell deformation subsystem that is in fluid communication with a suspension preparation subsystem, wherein the cell deformation subsystem is configured to receive a cell suspension from the suspension preparation subsystem and to flow the cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membrane of a cell, thereby enabling the entry of a payload into the cell, A dilution subsystem that is in fluid communication with a cell deformation subsystem, wherein the dilution subsystem is configured to receive a cell suspension from the cell deformation subsystem, receive a fluid or dry reagent, and create a diluted cell suspension by mixing the cell suspension with the fluid or dry reagent, A culture subsystem that is in fluid communication with a dilution subsystem, wherein the culture subsystem is configured to receive a diluted cell suspension from the dilution subsystem and to regulate the temperature of the diluted cell suspension, A cell washing subsystem that is in fluid communication with a culture subsystem, wherein the cell washing subsystem is configured to receive a diluted cell suspension from the culture subsystem and to perform a first buffer exchange operation on the cells to suspend the cells in a storage medium, A system comprising a container filling subsystem that is in fluid communication with a culture subsystem, wherein the container filling subsystem is configured to receive cells suspended in a storage medium from a cell washing subsystem and to allow the cells suspended in the storage medium to flow into one or more containers.

[0181] 2. The system according to Embodiment 1, wherein the suspension preparation subsystem is configured to receive a washing medium.

[0182] 3. The system according to Embodiment 1 or 2, wherein the suspension preparation subsystem is configured to accept a diluent.

[0183] 4. The system according to any one of Embodiments 1 to 3, wherein the suspension preparation subsystem includes a helical inertial isolation system configured to perform a cell isolation operation on cells.

[0184] 5. The system according to any one of Embodiments 1 to 4, wherein the suspension preparation subsystem includes a leukocyte removal filter system configured to perform a cell isolation operation on cells.

[0185] 6. The system according to any one of embodiments 1 to 5, wherein the suspension preparation subsystem includes a first tangential flow filtration system configured to perform a second buffer exchange operation.

[0186] 7. The system according to any one of embodiments 1 to 6, wherein the cell deformation subsystem includes a pressurizing system configured to generate pressure to pass a cell suspension through one or more cell deformation contractions.

[0187] 8. The system according to any one of embodiments 1 to 7, wherein the cell deformation subsystem includes a first temperature control system configured to control the temperature of the cell suspension.

[0188] 9. The system according to any one of embodiments 1 to 8, wherein the cell deformation subsystem includes a first stirring system configured to agitate the cell suspension to promote the homogeneity of the cell suspension.

[0189] 10. The system according to any one of Embodiments 1 to 9, wherein the cell reception in the suspension preparation subsystem includes one or more of the following: reception of blood and reception of cells suspended in a fluid other than blood.

[0190] 11. The system according to any one of embodiments 1 to 10, wherein the dilution subsystem includes a first metering system configured to measure the amount of fluid or dry reagent added to cells in a cell suspension.

[0191] 12. The system according to any one of embodiments 1 to 11, wherein the dilution subsystem includes a second stirring system configured to agitate the cell suspension to promote homogeneity of the cell suspension.

[0192] 13. The system according to any one of embodiments 1 to 12, wherein the dilution subsystem includes a first illumination system configured to illuminate one or more of the cell suspension and the diluted cell suspension.

[0193] 14. The system according to any one of Embodiments 1 to 13, wherein the culture subsystem includes a temperature control device configured to regulate the temperature of a diluted cell suspension.

[0194] 15. The system according to any one of embodiments 1 to 14, wherein the dilution subsystem includes a third stirring system configured to stir the diluted cell suspension to promote homogeneity of the diluted cell suspension.

[0195] 16. The system according to any one of embodiments 1 to 15, wherein the cell washing subsystem includes a second tangential flow filtration system configured to perform a first buffer exchange operation.

[0196] 17. The system according to any one of embodiments 1 to 16, wherein the cell washing subsystem includes a second metering system configured to measure the amount of a second buffer added to the cells during a first buffer exchange operation.

[0197] 18. The system according to any one of embodiments 1 to 17, wherein the cell washing subsystem includes a fourth stirring system configured to agitate and promote the homogeneity of one or more of the diluted cell suspension and the cells suspended in the storage medium.

[0198] 19. The system according to any one of embodiments 1 to 18, wherein the cell washing subsystem includes a second illumination system configured to illuminate one or more of the diluted cell suspension and cells suspended in a storage medium.

[0199] 20. The system according to any one of Embodiments 1 to 19, wherein the storage medium is a freeze-protected medium.

[0200] 21. The system according to any one of Embodiments 1 to 20, wherein the container filling subsystem includes a third weighing system configured to measure the amount of cells suspended in a preservation medium added to one or more containers.

[0201] 22. The system according to any one of embodiments 1 to 21, wherein the container filling subsystem includes a fifth stirring system configured to agitate and promote the homogeneity of cells suspended in a storage medium in one or more containers.

[0202] 23. The system according to any one of embodiments 1 to 22, wherein the container filling subsystem includes a third illumination system configured to illuminate cells suspended in a storage medium in one or more containers.

[0203] 24. The system according to any one of embodiments 1 to 23, wherein the system is configured such that a fluid flows gravitationally between two or more subsystems.

[0204] 25. The system according to any one of embodiments 1 to 34, wherein the system includes one or more pumps configured to pass fluid between two or more subsystems.

[0205] 26. The system according to any one of embodiments 1 to 25, wherein one or more subsystems can be made separate to test the integrity of one or more components of the subsystem.

[0206] 27. A method for processing cells, wherein the method is carried out by a system comprising a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, a culture subsystem, a cell washing subsystem, and a container filling subsystem, In the suspension preparation subsystem, Receiving cells and Performing cell isolation procedures on cells, Receiving the delivery medium, Creating a cell suspension by suspending cells in a delivery medium, In the cell deformation subsystem, Receiving the flow of cell suspension from the suspension preparation subsystem, The process involves flowing a cell suspension through one or more cell deformation contractions configured to induce perturbations in the cell membrane, thereby allowing the payload to enter the cell. In the dilution subsystem, Receiving the flow of cell suspension from the cell deformation subsystem, The ability to accept fluid or dry reagents, Diluted cell suspensions are prepared by mixing the cell suspension with a fluid or dry reagent. In the culture subsystem, Receiving the flow of diluted cell suspension from the dilution subsystem, Adjusting the temperature of the diluted cell suspension, In the cell washing subsystem, Receiving a flow of diluted cell suspension from the culture subsystem, The first buffer exchange procedure is performed on the cells to suspend them in the storage medium, In the container filling subsystem, The cell washing subsystem receives a stream of cells suspended in the storage medium, A method comprising flowing cells suspended in a storage medium into one or more containers.

[0207] 28. A disposable kit for use in a system for processing blood, the kit is A first set of disposable components configured for use in the suspension preparation subsystem of the system, wherein the first set of disposable components is Helical inertia isolation microfluidic consumables, and A first set of disposable components, including one or both of the first tangential flow filtration membrane assemblies, A second set of disposable components configured for use in the system's cell deformation subsystem, wherein the second set of disposable components is A component comprising one or more cell...

Claims

1. An automated point-of-care system for processing cells for cell-based therapeutics, wherein the automated point-of-care system is: A frame comprising multiple zones and multiple subsystems that can be detachably coupled to each zone, A first zone of the frame for receiving a suspension preparation subsystem, wherein the suspension preparation subsystem is A first cell suspension inlet and a delivery medium inlet, each of which is fluidly connected to a cell suspension device, the cell suspension device being configured to suspend cells in a delivery medium and thereby create a cell suspension. The first zone includes, A second zone of the frame for receiving a cell deformation subsystem, wherein the cell deformation subsystem is A second cell suspension inlet, which can be fluidly connected to the suspension preparation subsystem, One or more cell deformation contractions are configured to induce a perturbation in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and creating a cell suspension containing the manipulated cells. The second zone, which includes, A third zone of the frame for receiving a dilution subsystem, wherein the dilution subsystem is A third cell suspension inlet, which can be fluidly connected to the cell deformation subsystem, Buffer inlet and A container configured to receive the cell suspension containing the manipulated cells through the third cell suspension inlet, and to receive a fluid or dry reagent, which is mixed with the cell suspension containing the manipulated cells to create a diluted cell suspension, through the buffer inlet. The third zone, which includes, A fourth zone of the frame for receiving an incubation subsystem, wherein the incubation subsystem is The dilution subsystem includes a dilution cell suspension inlet that can be fluidly connected, A container configured to receive the diluted cell suspension through the diluted cell suspension inlet, A plate configured to regulate the temperature of the diluted cell suspension in the container to create an incubated cell suspension, The fourth zone, which includes, A fifth zone of the frame for receiving a cell washing subsystem, wherein the cell washing subsystem is The incubation subsystem includes an inlet for an incubated cell suspension that can be fluidly connected to the container by piping, Storage medium inlet, A container configured to receive the incubated cell suspension through the incubated cell suspension inlet, and to receive a storage medium to be mixed with the incubated cell suspension within the container through the storage medium inlet, thereby suspending the cells in the storage medium. The fifth zone, which includes, A sixth zone of the frame for receiving a container filling subsystem, wherein the container filling subsystem is The cell washing subsystem has an inlet that is fluid-connectable and configured to receive cells suspended in a storage medium, One or more containers configured to receive cells suspended in a preservation medium, One or more pumps configured to pump the cells suspended in a storage medium into one or more containers, The sixth zone, which includes The plurality of subsystems comprises at least the cell suspension preparation subsystem, the cell deformation subsystem, the incubation subsystem, and the container filling subsystem. The dilution subsystem can be omitted from the automated point-of-care system, and if omitted, the cell deformation subsystem can be fluidly connected to the incubation subsystem. The cell washing subsystem can be omitted from the automated point-of-care system. If omitted, the suspension preparation subsystem performs cell washing. An automated point-of-care system comprising a pump for pumping fluid through the automated point-of-care system to produce cells processed for cell-based therapeutics in 5 to 7 hours.

2. (i) The cell suspension inlet and buffer inlet of the dilution subsystem are the same inlet, and / or (ii) The automated point-of-care system according to claim 1, wherein the buffer inlet is used to deliver fluid to the dilution subsystem.

3. The suspension preparation subsystem (i) Cleaning medium inlet, and / or (ii) Dilution medium inlet, and / or (iii) One or more elutriation systems, leukocyte removal filter systems, or tangential flow filtration systems configured to perform a cell isolation procedure on the cells, and / or (iv) at least one outlet configured to be attached to at least one container for receiving cells suspended in a delivery medium. An automated point-of-care system according to claim 1 or 2, further comprising:

4. The cell deformation subsystem, (i) A pressurizing system configured to generate pressure to pass the cell suspension through one or more cell deformation contraction sections, and / or (ii) A preparation container configured to flow the cell suspension through one or more cell deformation contraction sections, and / or (iii) A temperature control system including a heated plate configured to control the temperature of the cell suspension, and / or (iv) At least one outlet configured to be attached to at least one container for receiving a cell suspension or a cell suspension containing manipulated cells. An automated point-of-care system according to any one of claims 1 to 3, further comprising:

5. The automated point-of-care system according to any one of claims 1 to 4, wherein the dilution subsystem includes at least one outlet configured to be coupled to the container configured to receive the cell suspension containing the manipulated cells.

6. The automated point-of-care system according to any one of claims 1 to 5, wherein the incubation subsystem includes a temperature control device configured to regulate the temperature of the diluted cell suspension.

7. The cell washing subsystem, (i) A tangential flow filtration system configured to perform a buffer exchange operation, and / or (ii) at least one outlet configured to be connected to the container An automated point-of-care system according to any one of claims 1 to 6, including the above.

8. An automated point-of-care system according to any one of claims 1 to 7, wherein one or more of the suspension preparation subsystem, the dilution subsystem, the incubation subsystem, the cell washing subsystem, and the container filling subsystem include a weighing device.

9. The automatic point-of-care system according to claim 8, wherein the weighing instrument includes a tension load cell, a compression load cell, or a straight bar load cell.

10. An automated point-of-care system according to any one of claims 1 to 9, wherein one or more of the cell deformation subsystem, the dilution subsystem, the incubation subsystem, the cell washing subsystem, and the container filling subsystem include a stirring system that includes a platform in contact with the container, configured to oscillate the container up and down.

11. The automated point-of-care system according to any one of claims 1 to 10, wherein the container filling subsystem includes one or more outlets configured to be coupled to one or more containers.

12. (i) The suspension preparation subsystem further includes one or more pumps configured to move fluid within the subsystem or between fluid-connected subsystems, and / or (ii) The dilution subsystem includes at least one pump configured to move fluid between fluid-connected subsystems, and / or (iii) The incubation subsystem includes at least one pump configured to move fluid between fluid-connected subsystems, and / or (iv) The automated point-of-care system according to any one of claims 1 to 11, wherein the container filling subsystem includes at least one pump configured to move fluid within the subsystem or between fluid-connected subsystems.

13. The automated point-of-care system according to any one of claims 1 to 12, wherein the automated point-of-care system includes one or more pumps configured to pump fluid between two or more subsystems.

14. The automated point-of-care system according to any one of claims 1 to 13, wherein the automated point-of-care system is used in a non-sterile environment.

15. (i) The suspension preparation subsystem further includes one or more of the following: a cell aggregate filter, a leukocyte removal filter, piping, pipe fittings, connectors, clamps, sampling bulbs, carboys, and air filters, and / or (ii) The cell deformation subsystem further includes one or more of the following: a rigid sample container, a cell aggregate filter, a rigid preparation container, one or more microfluidic chip cartridges, one or more microfluidic chips, piping, pipe fittings, connectors, clamps, air filters, and barrel filters, and / or (iii) The dilution subsystem further comprises one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, sampling bulbs, and carboys, and / or (iv) The incubation subsystem further includes one or more of the following: cell aggregate filters, piping, pipe fittings, connectors, and clamps, and / or (v) The cell washing subsystem further includes one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, sampling bulbs, air filters, and carboys, and / or (vi) The automated point-of-care system according to any one of claims 1 to 14, wherein the container filling subsystem further comprises one or more of the following: a cell aggregate filter, piping, pipe fittings, connectors, clamps, and sampling spheres.

16. An automated method for processing cells, wherein the automated method is carried out by a point-of-care system comprising one or more of a suspension preparation subsystem, a cell deformation subsystem, a dilution subsystem, an incubation subsystem, a cell washing subsystem, and a container filling subsystem, and the automated method is i. In the suspension preparation subsystem: (1) Receiving cells from a container, (2) Performing a cell isolation procedure on the aforementioned cells to produce isolated cells, (3) Receiving the delivery medium through the delivery medium inlet, (4) To produce a cell suspension by suspending the isolated cells in the delivery medium, and thereby produce a cell suspension, ii. In the aforementioned cell deformation subsystem: (1) Receiving the flow of the cell suspension from the suspension preparation subsystem, (2) Flowing the cell suspension through one or more cell deformation contraction units configured to induce perturbations in the cell membrane of the cells, thereby enabling the entry of a payload into the cells and thereby producing a cell suspension containing the manipulated cells, iii. In the aforementioned dilution subsystem: (1) Receiving the flow of the cell suspension containing cells manipulated from the cell deformation subsystem, (2) Receiving a fluid or dry reagent through the buffer inlet, (3) Producing a diluted cell suspension by mixing the cell suspension containing the manipulated cells with the fluid or dry reagent, iv. In the incubation subsystem: (1) Receiving the flow of the diluted cell suspension from the dilution subsystem, (2) Adjusting the temperature of the diluted cell suspension to produce an incubated cell suspension, v. In the cell washing subsystem: (1) Receiving the flow of the incubated cell suspension from the incubation subsystem, (2) Performing a buffer exchange operation on the cells to suspend the cells in a storage medium, vi. In the container filling subsystem: (1) Receiving a stream of cells suspended in a storage medium from the cell washing subsystem, (2) Introducing the cells suspended in the storage medium into one or more containers The automated method, including the above, is carried out in 5 to 7 hours. The aforementioned cells, (i) Red blood cells (RBCs) and / or vesicles derived from red blood cells, (ii) Peripheral blood mononuclear cells (PBMCs), or (iii) T cells, B cells, dendritic cells, monocytes, macrophages, eosinophils, basophils, natural killer (NK) cells, natural killer T (NKT) cells, mast cells, or neutrophils Automated methods, including [specific methods].

17. The automated method according to claim 16, wherein the payload comprises one or more initialization factors and / or one or more differentiation factors.

18. The method according to claim 16 or 17, wherein the automated method is carried out by a sterilization system in a non-sterile environment.

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