Systems and methods for micromanufacturing and bioprinting in a micro gravity environment thereof

Abstract

The system, methods, and uses of micromanufacturing materials, scaffolds, and living tissues in microgravity environment that is found in space such as Low-Earth-Orbit, on a spacecraft in-transit between celestial bodies, or on other celestial bodies such as the Moon or Mars.

Description

SYSTEMS AND METHODS FOR MICROMANUFACTURING AND BIOPRINTINGIN A MICRO GRAVITY ENVIRONMENT THEREOFINCORPORATION BY REFERENCE

[0001] This application claims the benefit of U.S. Provisional Application No. 63 / 589,314, filed October 10, 2023, which application is incorporated herein by reference in its entirety .BACKGROUND OF THE INVENTION

[0002] Traditional additive manufacturing processes in a microgravity environment rely on the controlled extrusion of material through nozzles or syringes to dispense material layer-by- layer until a complete three-dimensional object is formed. However, these processes provide for limited precision and accuracy of three-dimensional objects which are produced using such methods.SUMMARY

[0003] It is appreciated by the Applicant that there are inherent limitations to controlled extrusion and layer-by-layer approaches to three-dimensional printing systems which are currently used in microgravity environments. For example, by manufacturing through the extrusion of material, the overall spatial resolution of the features of a printed construct is limited to the size of the extruding component, while drastically longer fabrication sequences are required when using extrusion technologies to manufacture high resolution features. Further, such manufacturing processes do not permit the fabrication of hard tissues, can only dispense liquids with low viscosities which prevents them from forming rigid three dimensional tissues or scaffolds. There remains a need in the art for methods and systems which provide for manufacturing of three dimensional tissues or scaffolds in a microgravity environment.

[0004] An additive manufacturing system comprising: a housing defining a first sealable compartment, the housing being configurable from a first position in which the housing is sealed, to a second position in which the housing is unsealed, the housing comprising: a door providing access to the compartment when the housing is in the second position; one or more light sources; a removable cartridge defining a second sealable compartment comprising: a) a plurality of fluid reservoirs; b) a print chamber defining a third sealable compartment; c) a movable stage positioned within the print chamber; d) one or more transparent walls permitting for transmission of light into the removable cartridge.

[0005] In some embodiments, the removable cartridge is enclosed within the first compartment, wherein the print chamber is enclosed within the second compartment. In some embodiments, the print chamber is hermetically sealed from the second compartment. In some embodiments, the removable cartridge is hermetically sealed from the first compartment. In some embodiments, the print chamber comprises one or more transparent walls permitting for transmission of light into the print chamber. In some embodiments, the print chamber is fully filled with a liquid and comprises essentially no gas. In some embodiments, the one or more transparent walls are gas permeable. In some embodiments, the gas permeability permits an active or passive flow of oxygen. In some embodiments, the one or more transparent walls comprise a gas inlet. In some embodiments, the system further comprises a pressurized oxygen source coupled to an oxygen line, wherein the oxygen line is directly coupled to the one or more transparent walls. In some embodiments, the system further comprises one or more fans configured to direct an active flow of oxygen to the one or more transparent walls. In some embodiments, the system comprises an influx vent, an efflux vent, or a combination thereof. In some embodiments, the system comprises an influx vent, an efflux vent, or a combination thereof, wherein the influx vent or efflux vent further comprise HEPA filters. In some embodiments, the system comprises HEPA filters in the 1st compartment. In some embodiments, the system comprises HEPA filters in the 1 st compartment, wherein the system is not air tight.

[0006] In some embodiments, the removable cartridge comprises a gas inlet directing gas proximally to the one or more transparent walls. In some embodiments, the removable cartridge is interchangeable within the system. In some embodiments, the plurality of fluid reservoirs are removable and interchangeable within the removable cartridge. In some embodiments, the removable cartridge is thereby modular. In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the plurality of fluid reservoirs, and provides an outlet of fluid to at least one of the plurality of fluid reservoirs.

[0007] In some embodiments, the plurality of fluid reservoirs comprise a wash fluid and a photopolymerizable fluid, wherein the outlet of fluid to at least one of the plurality of fluid reservoirs is to the fluid reservoir comprising the photopolymerizable fluid. In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage. In some embodiments, the perfusionpath comprises one or more pumps coupled to the fluid reservoirs and the print chamber. In some embodiments, the removable cartridge is moveable within the system along at least 2 of an x-axis, a y-axis, and a z-axis. In some embodiments, the removable cartridge is movable relative to at least one light source, thereby allowing a larger exposure area than a single field of view of light form the at least one light source onto the movable stage. In some embodiments, the moveable stage comprises a patterned surface configured to enhance adhesion between the patterned surface and a fluid or a solid in contact with the patterned surface. In some embodiments, the patterned surface comprises a plurality of raised regions and recessed regions arranged in a predetermined pattern. In some embodiments, the raised regions have a height from 0.1 mm to 1 mm. In some embodiments, the one or more transparent walls comprise an anti -adhesion surface configured to reduce adhesion between a fluid or a solid in contact with the anti-adhesion surface. In some embodiments, the antiadhesion surface comprises a coating of a hydrophobic or a superhydrophobic material. In some embodiments, the anti-adhesion surface comprises a micropattem or nanopattern to induce hydrophobic or superhydrophobic properties. In some embodiments, the micropattern or nanopattern comprises dimensions less than 500 nm. In some embodiments, the micropattern or nanopattem comprises dimensions less than 400 nm. In some embodiments, the removable cartridge further comprises a heating element. In some embodiments, the system further comprises one or more imaging devices configured to image the first compartment, or the second compartment.

[0008] In some embodiments, the system further comprises one or more imaging devices configured to image one or more components of the system. In some embodiments, the system further comprises one or more imaging devices configured to image an object manufactured within the system. In some embodiments, the system further comprises one or more imaging devices configured to image the internals of the system before, during, and after a fabrication sequence. In some embodiments, the system further comprises one or more imaging devices configured to image the removable cartridge during a fabrication sequence. In some embodiments, one or more imaging devices are oriented to image a completed 3D object after the fabrication steps. In some embodiments, the one or more imaging devices comprise one or more optical cameras. In some embodiments, the system can be placed in any orientation and maintains functionality without regard to a gravitational field. In some embodiments, the system is configured to operate in microgravity, e.g., low earth orbit, or any orbit around a celestial object. In some embodiments, the system can be placed in any orientation and maintains functionality without regard to a gravitational field, wherein thesystem is configured to operate in microgravity. In some embodiments, the removable cartridge further comprises a thermometer. In some embodiments, the system is configured to maintain the temperature within the removable cartridge below or above a critical level. In some embodiments, the system further comprises an avionics spine comprising a plurality of electrical and data connections electrically coupling the system to a spacecraft. In some embodiments, the one or more transparent walls comprises glass, quartz, transparent plastic, silicon, silica, poly dimethylsiloxane (PDMS), transparent ceramic, fluorinated ethylene propylene (FEP), or combinations thereof. In some embodiments, at least one of the plurality of fluid reservoirs comprise a photopolymerizable material, a photo initiator, a chemical quencher, (2,2,6,6-Tetramethylpiperidin-l-yl)oxyl, (2,2,6,6-tetramethylpiperidin-l- yl)oxidanyl (TEMPO), a physical quencher (color dye), a wash solution, a biologic solution, cell media, suspension of cells, microparticles, nanoparticles, therapeutic loaded nanoparticles, therapeutic loaded microparticles, electrolytic or electrically conductive materials, or combinations thereof. In some embodiments, the wash solution comprises water, a buffer, an organic material, a solution containing antibiotics, an antimicrobial agent, or combinations thereof. In some embodiments, the biologic solution comprises nutrients and / or growth factors suitable for cell culture and proliferation.

[0009] In some embodiments, the biologic solution comprises a suspension of cells. In some embodiments, the one or more light sources are movable within the first compartment. In some embodiments, the removable cartridge is moveable with the first compartment. In some embodiments, the system further comprises a plurality of spatial light modulators configured to direct light from the one or more light sources. In some embodiments, the plurality of spatial light modulators are configured to direct light along an x-axis, and a y-axis. In some embodiments, the plurality of spatial light modulators are configured to direct light along an x-axis, a y-axis, and a z-axis. In some embodiments, the plurality of spatial light modulators comprise one or more digital micro-mirror devices. In some embodiments, the plurality of spatial light modulators comprise one or more liquid crystal devices. In some embodiments, at least one of the one or more light sources comprises a light emitting diode.

[0010] In some embodiments, at least one of the one or more light sources comprises a laser. In some embodiments, at least one of the one or more light sources is movable along an x- axis, and a y-axis.

[0011] In some embodiments, at least one of the one or more light sources is movable along an x-axis, a y-axis, and a z-axis. In some embodiments, the housing is configured to interfacewith a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform.

[0012] In some embodiments, the removable cartridge comprises a plurality of movable stages, each stage in line with at least one of the one or more transparent walls. In some embodiments, at least one of the one or more light sources is movable along two or more of an x-axis, a y-axis, and a z-axis relative to the removable cartridge such that multiple optical field-of-views can be exposed onto at least one of the plurality of movable stages, thereby allowing a larger print area than a single field of view.

[0013] In some embodiments, the system further comprises one or more computer processors comprising a computer readable medium configured to implement a method of additively manufacturing a 3D object comprising: a) sealing one or more of the first compartment, the second compartment, or the third compartment from one another; b) filling the print chamber with a photopolymerizable fluid comprised by the plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from the one or more light sources through the one or more transparent walls onto the movable stage and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid, wherein the method takes place in a reduced gravity environment.

[0014] In some embodiments, the computer readable medium further comprises instructions for producing a scaffold, a biological tissue, or a microfluidic device.

[0015] In some embodiments, the instructions comprise a 3D model of the 3D object in discrete layers which correspond to a printing resolution of the system about a z axis defined by the movement of the movable stage. In some embodiments, the method further comprises monitoring the 3D object with one or more imaging devices. In some embodiments, the method further comprises monitoring the 3D object during any of c. -e. with one or more imaging devices. In some embodiments, the monitoring occurs before, during, and after a fabrication sequence. In some embodiments, the method further comprises monitoring a component of the system, the first compartment, or the second compartment with one or more imaging devices. In some embodiments, the method further comprises monitoring a component of the system, the first compartment, or the second compartment during any of c.- e. with one or more imaging devices. In some embodiments, the one or more imaging devices comprise one or more optical cameras.

[0016] In some embodiments, the monitoring occurs before, during, and after a fabrication sequence. In some embodiments, the monitoring 3D object comprises monitoring a dimensionality of the solid for compliance with the 3D model. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model is within 0.6 microns.

[0017] In some embodiments, the computer readable medium further comprises instructions for producing the 3D object, the 3D object comprising a plurality of channels. In some embodiments, the plurality of fluid reservoirs comprise a wash fluid and a photopolymerizable fluid, where the removable cartridge comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage, wherein the channels align with the inlet, wherein the method further comprise flushing the plurality of channels with the wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels. In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising flushing the print chamber with the wash fluid to remove the photopolymerizable fluid, and directing the wash fluid and the removed photopolymerizable fluid into one of the fluid reservoirs initially holding the photopolymerizable fluid.

[0018] In some embodiments, the directing the light comprises directing two-dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage. In some embodiments, the method further comprises moving the one or more light sources relative to the movable stage and exposing multiple optical field-of- views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the method further comprises moving the removeable cartridge relative to the movable stage and exposing multiple optical field-of-views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the moving the movable stage comprises moving the movable stage away from the optical plane such that the projected two-dimensional cross-sections of the three-dimensional object are incident on photopolymerizable fluid in contact with and adjacent to previously formed layers of the object.

[0019] In some embodiments, the moving the movable stage comprises moving the movable stage continuously. In some embodiments, the moving the movable stage comprises movingthe movable stage incrementally. In some embodiments, the directing light comprises intermittently activating the one or more light sources during the formation of the three- dimensional object. In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with a thermometer and maintaining the temperature above or below a critical level.

[0020] In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with a thermometer, and ceasing operation of the system if a critical temperature is reached. In some embodiments, the one or more transparent walls are gas permeable, the method further comprising providing an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring against the one or more transparent walls. In some embodiments, the method further comprising contacting the top layer of the photopolymerizable fluid with the one or more transparent walls. In some embodiments, the top layer comprises a thickness of between about 1 to about 200 micrometers. In some embodiments, the plurality of fluid reservoirs comprise a chemical quencher, physical quencher, or a combination thereof that quenches free-radicals, light intensity, irradiance, or light exposure and prevents polymerization, or over polymerization. In some embodiments, the method further comprises filling the print chamber with the physical quencher and preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber.

[0021] In some embodiments, the target polymerization depth is between about 1 to about 200 micrometers. In some embodiments, the target polymerization depth is between about 10 to about 50 micrometers. In some embodiments, a concentration of the physical quencher in the print chamber is up to 1 -5% v / v. In some embodiments, the concentration of the physical quencher in the print chamber is at least about 4%. In some embodiments, the method further comprises mounting the system in an orientation without regard to a gravitational field. In some embodiments, the method further comprises the operating the system in an orientation without regard to a gravitational field. In some embodiments, any of the preceding methods, a printed construct is created. In some embodiments, the plurality of fluid reservoirs comprises a cell media fluid reservoir.

[0022] In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising perfusing the cell media throughthe newly printed construct. In some embodiments, the moving the stage occurs at a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

[0023] In some embodiments, the method further comprises additively manufacturing the 3D object at a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

[0024] In some embodiments, the sealing one or more of the first compartment, the second compartment, or the third compartment from one another comprising hermetically sealing one or more of the first compartment, the second compartment, or the third compartment from one another.

[0025] A method of additively manufacturing a 3D object comprising: a) sealing one or more of a first compartment, a second compartment, or a third compartment from one another, wherein the second compartment and third compartment comprise one or more transparent walls; b) filling a print chamber with a photopolymerizable fluid comprised by a plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from a light source through one or more transparent walls of the second compartment or the third compartment onto a movable stage within the print chamber and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid, wherein the method takes place in a reduced gravity environment.

[0026] In some embodiments, the method further comprises interfacing the first compartment with a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform. In some embodiments, the 3D object is a scaffold, a biological tissue, or a microfluidic device. In some embodiments, the photopolymerizing comprises printingthe 3D object in discrete layers which correspond to a printing resolution of the method about a z axis defined by the movement of the movable stage. In some embodiments, the method further comprises monitoring the 3D object with one or more imaging devices. In some embodiments, the method further comprises monitoring the 3D object during any of c. -e. with one or more imaging devices. In some embodiments, the one or more imaging devices comprise one or more optical cameras. In some embodiments, the monitoring occurs before, during, and after a fabrication sequence comprising performance of a. -e. In some embodiments, the monitoring occurs before, during, and after a fabrication sequence. In some embodiments, the monitoring the 3D object comprises monitoring a dimensionality of the solid for compliance with a 3D model. In some embodiments, the monitoring thedimensionality of the solid for compliance with the 3D model is within 0.6 microns. In some embodiments, the method further comprises flushing a plurality of channels of the 3D object with a wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels. In some embodiments, the method further comprises directing the wash fluid and the removed photopolymerizable fluid into a fluid reservoir initially holding the photopolymerizable fluid.

[0027] In some embodiments, the directing light from a light source comprises directing two- dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage. In some embodiments, the method further comprises moving the light source relative to the movable stage and exposing multiple optical field -of- views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the method further comprises moving a removeable cartridge defining the second compartment relative to the movable stage and exposing multiple optical field-of-views onto the moveable stage, thereby allowing a larger print area than a single field of view.

[0028] In some embodiments, the moving the movable stage comprises moving the movable stage away from the optical plane such that the projected two-dimensional cross-sections of the three-dimensional object are incident on photopolymerizable fluid in contact with and adjacent to previously formed layers of the object. In some embodiments, the moving the movable stage comprises movingthe movable stage continuously. In some embodiments, the moving the movable stage comprises moving the movable stage incrementally. In some embodiments, the directing light from a light source comprises intermittently activating the light source during the formation of the three-dimensional object. In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with a thermometer, and maintaining the temperature below a critical level. In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with a thermometer, and ceasing operation of the method if a critical temperature is reached. In some embodiments, the method further comprises providing an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring in the top layer of the photopolymerizable fluid. In some embodiments, the method further comprises contacting a top layer of the photopolymerizable fluid with one or more transparent walls comprising an anti -adhesive surface, a hydrophobic surface, or combinations thereof. In some embodiments, a top layer of the photopolymerizable fluid comprises a thickness of about 1 to about 200 microns. In someembodiments, the method further comprises introducing a chemical quencher into the photopolymerizable fluid, and preventing polymerization, or over polymerization, by quenching a free radical.

[0029] In some embodiments, the method further comprises at least partially filling the print chamber with the chemical quencher and preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber. In some embodiments, the top layer is between about 1 to about 200 microns. In some embodiments, the top layer is between about 10 to about 50 microns. In some embodiments, the movingthe movable stage occurs at a speed of atleast 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min. In some embodiments, the method further comprises additively manufacturing the 3D objectat a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min. In some embodiments, the sealing one or more of a first compartment, a second compartment, or a third compartment from one another comprises hermetically sealing one or more of a first compartment, a second compartment, or a third compartment from one another.BRIEF DESCRIPTION OF THE DRAWINGS

[0030] The novel features of the invention are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present invention will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the invention are utilized, and the accompanying drawings of which:

[0031] FIG. 1A depicts an additive manufacturing system, according to some embodiments.

[0032] FIG. IB depicts an additive manufacturing system internal subsystem, according to some embodiments.

[0033] FIG. 1C depicts an additive manufacturing system, according to some embodiments.

[0034] FIG. ID depicts an additive manufacturing system print cartridge, according to some embodiments.

[0035] FIG. 2A depicts an additive manufacturing system subsystem internals and print cartridge, according to some embodiments.

[0036] FIG. 2B depicts an additive manufacturing system subsystem internals, according to some embodiments.

[0037] FIG. 3A depicts an additive manufacturing enclosure system from a closed position, according to some embodiments.

[0038] FIG. 3B depicts an additive manufacturing enclosure system from an open position, according to some embodiments.

[0039] FIG. 4A depicts a first side perspective view of an additive manufacturing system internals, according to some embodiments.

[0040] FIG. 4B depicts a second side perspective view of an additive manufacturing system internals, according to some embodiments.

[0041] FIG. 4C depicts a third side perspective view of an additive manufacturing system internals, according to some embodiments.

[0042] FIG. 5A depicts a partial side perspective view of an additive manufacturing system internals, according to some embodiments.

[0043] FIG. 5B depicts a fourth side perspective view of an additive manufacturing system internals, according to some embodiments.

[0044] FIG. 6A depicts a first perspective view of an additive manufacturing system print cartridge, according to some embodiments.

[0045] FIG. 6B depicts a second perspective view of an additive manufacturing system print cartridge, according to some embodiments.

[0046] FIG. 6C depicts a third perspective view of an additive manufacturing system print cartridge, according to some embodiments.

[0047] FIG. 7A depicts an additive manufacturing system print chamber, according to some embodiments.

[0048] FIG. 7B depicts an additive manufacturing system print chamber, according to some embodiments.

[0049] FIG. 8A depicts a disassembled additive manufacturing system print cartridge, according to some embodiments.

[0050] FIG. 8B depicts a disassembled additive manufacturing system print cartridge, according to some embodiments.

[0051] FIG. 8C depicts a disassembled additive manufacturing system print cartridge, according to some embodiments.DETAILED DESCRIPTION

[0052] It is appreciated by the Applicant that there are inherent limitations to controlled extrusion and layer-by-layer approaches to three-dimensional printing systems which are currently used in microgravity environments. For example, by manufacturing through the extrusion of material, the overall spatial resolution of the features of a printed construct islimited to the size of the extruding component, as well as drastically longer fabrication sequences when using extrusion technologies to manufacture high resolution features. Further, such manufacturing processes do not permit the fabrication of hard tissues, can only dispense liquids with low viscosities which prevents them from forming rigid three dimensional tissues or scaffolds. There remains a need in the art for methods and systems which provide for manufacturing.

[0053] For example, by manufacturing through the extrusion of material, the overall spatial resolution of the features of a printed construct is limited to the size of the extruding component. Although the extruding component can be made to dispense small volumes of material at any given time, there is then a tradeoff with the overall time it takes to make the entire object. To allow for manufacture of high-resolution features on an object using an extrusion based manufacturing, one must accept drastically longer fabrication sequences. In addition to prolonged fabrication sequences, on the order of hours to manufacture objects, these manufacturing processes limit the material selection available to the user. In order to accommodate another material, additional print heads would need to be added which can take up more space than is available and further complicates the fabrication sequence. Further, when applied to bioprinting, these manufacturing processes do not permit the fabrication of hard tissues. Due to their innate characteristics, these processes can only dispense liquids with low viscosities which prevents them from forming rigid three dimensional tissues or scaffolds.

[0054] The disclosure provided herein overcomes the limitations of these manufacturing processes to establish robust manufacturing capabilities in a microgravity environment, such as an off-Earth setting as might be observed in satellite in orbit above earth.

[0055] Provided herein are systems, methods, and uses of micromanufacturing three- dimensional objects using digital photolithography in a microgravity environment. In some embodiments, the systems, methods, and uses of micromanufacturing are provided to fabricate materials, scaffolds, and living tissues in microgravity environment. Such systems methods provide for fast fabrication sequences, on the order of minutes to produce fully fabricated objects, the ability to manufacture very high resolution features using the photolithography based processes, while overcoming challenges presented by the operation of such systems and methods in a microgravity environment. For example, layers of 3D printed objects can be manufactured within seconds, and entire objects within minutes.

[0056] When using an additive manufacturing system in a microgravity environment, the placement of liquid matter cannot precisely be controlled due to the lack of an appreciablegravitational field, which is traditionally relied upon for the separation of gas and liquids on Earth. Therefore, in order to provide for the correct placement of liquids in an additive manufacturing system, for astronaut safety, e.g., preventing outgassing, and as well as for high tolerance and high precision manufacturing, it becomes necessary to account for placement of liquid matter in a microgravity environment, as well as direction of light precisely within the fluid if using a photopolymerizable printing approach. One solution to such manufacturing and safety problem presented with additive manufacturing systems using liquids in a microgravity environment are systems, methods, and uses of additive manufacturing disclosed herein which utilize an additive manufacturing system comprising multiple sealed enclosures, sealed print chambers with limited, essentially none, or no gaseous content, printing stages which provide for the movement of polymerized objects within a sealed print chamber, light sources to direct light for polymerization to a polymerizable fluid, and transparent features of a print chamber which provide for transmission of light to a photopolymerizable fluid, among other relevant aspects of the system.

[0057] In some embodiments, the system comprises multiple sealed enclosures which are fully sealed so that fluids (e.g., liquids, or gas) cannot pass between these enclosures. In some embodiments, at least one enclosure comprises a print chamber, wherein the print chamber is separate from other system compartments. In some embodiments, the print chamber is completely filled with a liquid, and comprises substantially no gas, or no gas. In some embodiments, the print chamber comprises one or more transparent walls or windows to allow a light to be transmitted to the photopolymerizable fluid within a print chamber, and form a solid on a portion of the fluid in contact with a movable stage, or adjacent portion of already polymerized fluid (e.g., 3D printed object being manufactured).

[0058] The systems, methods and uses of additive manufacturing systems provided which allow for consistent and reliable placement of fluid, polymerized fluid (e.g., a solid), and light within a print chamber in a microgravity environment disclosed herein provide a beneficial technical effect of allowing high precision and high tolerance manufacturing in a microgravity environment. By using photolithography, for example, light based polymerization, the additive manufacturing system provides for very high manufacturing tolerances, potentially as precise as the wavelength of the light used for photo polymerization, e.g., on the order of a couple hundred of nanometers, as opposed to other methods, e.g., an extrusion, which are generally only capable of manufacturing tolerances on the order of millimeters at any commercially practicable lengths of manufacturing sequences.

[0059] In some embodiments, the additive manufacturing system can be placed in any orientation without regard to a gravitational field, and can be operated and utilized to produce a 3D printed object in any orientation. In some embodiments, the additive manufacturing system is configured to operate in a microgravity environment, such as in low earth orbit or in any orbit around a celestial object. In some embodiments, the additive manufacturing system can be placed in any orientation without regard to a gravitational field, wherein the system is configured to operate in microgravity, low earth orbit or in any orbit around a celestial object.

[0060] In some embodiments, a system is provided for additive manufacturing of a three- dimensional object. In some embodiments, the additive manufacturing system comprises a housing defining a first sealable compartment. In some embodiments, the housing is configurable from a first position in which the housing is sealed, to a second position in which the housing is unsealed. In some embodiments, the housing comprises: a door providing access to the compartment when the housing is in the second position; one or more light sources; a removable cartridge defining a second sealable compartment comprising: a plurality of fluid reservoirs; a print chamber defining a third sealable compartment; a movable stage positioned within the print chamber; one or more transparent walls permitting for transmission of light into the removable cartridge.

[0061] In some embodiments, the system (130) is comprised of a housing (110) that serves as an interface between the internal subsystem (120) and the host spacecraft or module. In some embodiments, the housing (110) contains an avionics spine (150) that passes all electrical power and data between the host spacecraft or module and the internal subsystem. In some embodiments, the internal subsystem (120) comprises a projector (950) containing a light source and spatial light modulator, projection optics (940), linear axis (910), and removable cartridge (140). In some embodiments, the removable cartridge (140) contains a print chamber (1250), heaters, sensors, one or more fluid reservoirs (1220) to contain the photopolymerizable material, one or more fluid reservoirs (1220) to contain a wash or perfusion solution, a pump (1270), a microcontroller (1210), and the necessary tubing.

[0062] FIG. 1A depicts an embodiment of an additive manufacturing system (130) with the housing (110) door closed, with the housing in a first position in which the first compartment is sealed (e.g., the system is sealed). FIG. IB depicts an embodiment of an additive manufacturing system (130) internal subsystem (120). FIG. 1C depicts an embodiment of an additive manufacturing system (130) with the housing (110) door open and the internalsubsystem (120) tray extended, for example, with the housing in the first position in which the compartment is unsealed (e.g., the door is open).

[0063] FIG. 3A depicts an embodiment of the housing of the additive manufacturing system (130) in a first position (e.g., closed position). FIG. 3B depicts an embodiment of the housing of the additive manufacturing system (130) in a second position (e.g., open position).

[0064] In some embodiments, the additive manufacturing system includes a housing (110) which serves as a system enclosure. In some embodiments, the housing (110) is suitable for interface with a host module or spacecraft. In some embodiments, the additive manufacturing system (130) comprises a housing defining a first compartment (111). In some embodiments, the housing is configurable from a first position in which the housing is sealed, to a second position in which the housing is unsealed. In some embodiments, the housing can be fabricated from metals, steel, stainless steel, aluminum, copper, alloys, polymers, plastics, PEEK, or combinations thereof. In some embodiments, the housing comprises a plurality of layers, wherein at least one layer comprises a conductive material and at least one layer comprises a dielectric material. In some embodiments, the housing is constructed from a material that shields the internal subsystem from electromagnetic interference (EMI). In some embodiments, the housing forms a barrier to EMI. In some embodiments, the housing is configured to prevent the transmission of electromagnetic waves to the internal subsystem.

[0065] In some embodiments, the housing comprises a door providing access to the first compartment (111) when the housing is the second position; one or more light sources; and a removable cartridge (140) defining a second compartment. In some embodiments, the housing is configured to interface with a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform. In some embodiments, the housing comprises an influx vent and an efflux vent, each of which is connected to a gaseous filtration system. In some embodiments, the filtration system may comprise a HEPA filter, an ultraviolet light source, or other components to remove contaminants from the air entering or exiting the compartment.

[0066] In some embodiment, the system may comprise a plurality of sealed compartments comprise various components of the additive manufacturing system, and which prevent the movement of fluids between the various compartments. In some cases, the plurality of sealed compartments contain the fluids within the various compartments to prevent the inadvertent movement of fluid within the system or within a microgravity environment where there is no gravitational field, or a very weak gravitational field, such that liquids are not pulled to the bottom of the system by gravity. In some embodiments, the housing may define a firstsealable compartment, the housing being configurable from a first position in which the housing is sealed, to a second position in which the housing is unsealed. In some embodiments, the housing may comprise and may enclose a removable cartridge defining a second sealable compartment. In some embodiments, the removable cartridge may comprise and may enclose a print chamber defining a third sealable compartment. The plurality of sealed compartments within the additive manufacturing system can provide for user (e.g., astronaut) safety when operating the system in a microgravity environment. For instance, the removable cartridge defining the second sealable compartment may comprise a plurality of fluid reservoirs, and, in some cases, all of the fluid and all fluid reservoirs, utilized in the additive manufacturing process may be entirely contained within the removable cartridge and / or the second sealable compartment. By containing all fluid within a second sealable compartment within a first sealable compartment, all fluid can be contained within the second compartment and separated from other components of the system in the first compartment which could be damaged by exposure to fluids (e.g., electronics). Further, it is possible to ensure that there is no inadvertent exposure of fluid to an environment external to the additive manufacturing system, in the event of sealant failure in the second compartment, such that user (e.g., astronaut) safety is not compromised even in the event of sealant failure in one or more compartments.

[0067] In some cases, the plurality of sealed compartments within the additive manufacturing system can provide for highly controlled manufacturing when operating the system in a microgravity environment. For instance, a print chamber defining a third sealable compartment which is contained within the second sealable compartment may comprise and contain all fluid which is utilized in a photolithography based (e.g., photopolymerization based) manufacturing process, such that there is no free movement of fluid outside of a perfusion system (e.g., a fluid line or tubing leading to a fluid reservoir), for example, between the second sealable compartment and the third sealable compartment. The print chamber and / or third sealable compartment may be degassed and completely filled with a liquid such as a completely filled waterbottle, such that there is a substantially homogenous distribution of fluid within the print chamber. In some embodiments, the print chamber is completely filled with a liquid to establish a homogeneous distribution of liquid within the print chamber without any air or air bubbles in the chamber which could create gaps in the 3D printed object if the photopolymerizable fluid were polymerized around it. By providing a print chamber which defines a third sealable compartment which comprises essentially no gas and a substantial homogenous liquid therein, a beneficial technical effect of improvingmanufacturing a three dimensional object using a photolithography based additive manufacturing method in a microgravity environment without regard to polymerization of the fluid around air or air bubbles in the 3D printed object, and without regards to an orientation of the system relative to a gravitational field.

[0068] FIG. 4A-4C depict example embodiments of the additive manufacturing internal subsystem. FIG. 4A shows a first side perspective view of an embodiment of an additive manufacturing internal subsystem. FIG. 4B depicts a second side perspective view of an embodiment of an additive manufacturing internal subsystem. FIG. 4C depicts a third perspective view of an embodiment of an additive manufacturing internal subsystem.

[0069] In some embodiments, the system (130) comprises linear axis (910) to control build plane motion, a mirror (920), a shutter (930), projection optics (940), and a projector (950). In some embodiments, a linear axis (910) to control build plane motion, a mirror (920), a shutter (930), projection optics (940), and a projector (950) are comprised within the first compartment defined by the housing. In some embodiments, the projector (950) comprises at least one light source and at least one spatial light modulator. In some embodiments, the system (130) comprises a digital microcontroller (960) to control the internal subsystem, linear axis (970) for post-print imaging camera, and post-print imaging camera (980).

[0070] FIG. ID and FIG. 6A-6C depict example embodiments of additive manufacturing system removable cartridge (140). FIG. ID depicts an embodiment of an additive manufacturing system removable cartridge (140). FIG. 6A depicts a first perspective view of an embodiment of an additive manufacturing system removable cartridge. FIG. 6B depicts a second perspective view of an embodiment of an additive manufacturing system removable cartridge (140). FIG. 6C depicts a third perspective view of an embodiment of an additive manufacturing system removable cartridge (140). In some embodiments, the additive manufacturing system comprises a cartridge. In some embodiments, the additive manufacturing system comprises a removable cartridge (140). In some embodiments the removable cartridge (140) comprises a digital microcontroller (1210) to the removable cartridge, reservoir reservoirs (1220), and a movable stage (1230). In some embodiments, the removable cartridge (140) comprises a print chamber (1250) and fluid flow connection points (1260). In some embodiments, the removable cartridge (140) contains a print chamber (1250), heaters, sensors, one or more fluid reservoirs (1220) to contain the photopolymerizable material, one or more fluid reservoirs (1220) to contain a wash or perfusion solution, a pump (1270), a microcontroller (1210), and the necessary tubing.

[0071] In some embodiments, the removable cartridge (140) is prefilled with the desired photopolymerizable material in one fluid reservoir (1220), and the desired wash solution in another fluid reservoir (1220). In some embodiments, once inserted, the microcontroller (1210) is connected to the avionics spine (150) via the digital microcontroller connection interface (1240).

[0072] In some embodiments, the housing comprises the removable cartridge (140) defining the second compartment. In some embodiments, the removable cartridge (140) is enclosed within the first compartment (111), wherein the print chamber (1250) is enclosed within the second compartment. The print chamber can define the third sealable compartment. In some embodiments, the removable cartridge (140) is hermetically sealed from the first compartment (111). In some embodiments, the removable cartridge (140) is interchangeable within the system with another like removable cartridge. In some embodiments, the removable cartridge (140) comprises a plurality of fluid reservoirs which contain different liquids therein, and which are interchangeable within the cartridge, and is thereby modular. In some embodiments, the removable cartridge may be about 1-15 cm, by about 5-10 cm. In some embodiments, the size of the removable cartridge may vary based upon the size of the 3D object to be manufactured.

[0073] In some embodiments, the removable cartridge (140) is moveable with the first compartment (111). In some embodiments, the additive manufacturing system is configured to move the removeable cartridge (140) relative to the movable stage (1230) and exposing multiple optical field-of-views onto the moveable stage (1230), thereby allowing a larger print area than a single field of view.

[0074] FIG. 8A-8C depict example embodiments of additive manufacturing system print cartridge (140). FIG. 8A depicts an embodiment of a first outlined additive manufacturing system disassembled removable cartridge (140). FIG. 8B depicts an embodiment of a second outlined additive manufacturing system disassembled removable cartridge. In some embodiments, the movable stage plunger (1310) further comprises one or more gaskets (1320). FIG. 8C depicts an embodiment of a third outlined additive manufacturing system disassembled removable cartridge. In some embodiments, the print chamber (1250) comprises a build chamber housing (1410), O-rings (1420) and an optically transparent cartridge window (1430). In some embodiments, the top layer of the photopolymerizable fluid is in contact with the one or more transparent walls.

[0075] In some embodiments, the top layer of the photopolymerizable fluid comprises a thickness of at least about 1 micrometer. In some embodiments, the top layer of thephotopolymerizable fluid comprises a thickness of between about 1 to about 200 micrometers. In some embodiments, the top layer of the photopolymerizable fluid comprises a thickness of up to about 200 micrometers. In some embodiments, the print chamber (1250) is filled with the chemical quencher, thereby preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber. In some embodiments, the target polymerization depth is between about 1 to about 200 micrometers. In some embodiments, the target polymerization depth is at least 10 micrometers. In some embodiments, the target polymerization depth is between 10-50 micrometers. In some embodiments, the target polymerization depth is up to 50 micrometers.

[0076] In some embodiments, the removable cartridge (140) comprises one or more fluid reservoir (e.g., fluid bags) (1220). In some embodiments, the fluid reservoirs (1220) comprise bio ink (e.g., a photopolymerizable fluid), a wash solution, or a combination thereof. In some embodiments, the fluid reservoirs (1220) are coupled to the movable stage (1230). In some embodiments, the pump (1270) is configured to pump fresh wash solution into the bio print chamber (1250). In some embodiments, the pump (1270) is configured to remove the unpolymerized ink.

[0077] In some embodiments, the plurality of fluid reservoirs (1220) comprise a chemical quencher that quenches free-radicals and prevents polymerization, or over polymerization. In some embodiments, the plurality of fluid reservoirs (1220) comprise a physical quencher that is partially or fully opaque which reduces the transmission of light to the fluid, and prevents polymerization, or over polymerization. In some embodiments, the plurality of fluid reservoirs (1220) comprise a cell media fluid reservoir. In some embodiments, a concentration of the chemical quencher in the print chamber is at least 1% w / v. In some embodiments, the concentration of the chemical quencher in the print chamber is between 1 - 5% w / v. In some embodiments, the concentration of the chemical quencher in the print chamber is up to 5% w / v. In some embodiments, a concentration of the phy sical quencher in the print chamber is about 4% w / v. In some embodiments, a concentration of the physical quencher in the print chamber is about 0.05 % w / v to about 5 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber is about 0.05 % w / v to about 0. 1 % w / v, about 0.05 % w / v to about 0.25 % w / v, about 0.05 % w / v to about 0.5 % w / v, about 0.05 % w / v to about 0.75 % w / v, about 0.05 % w / v to about 1 % w / v, about 0.05 % w / v to about 1.5 % w / v, about 0.05 % w / v to about 2 % w / v, about 0.05 % w / v to about 2.5 % w / v, about 0.05 % w / v to about 3 % w / v, about 0.05 % w / v to about 4 % w / v, about 0.05 % w / v to about 5 % w / v, about 0.1 % w / v to about 0.25 % w / v, about 0.1 % w / v to about 0.5 % w / v,about 0.1 % w / v to about 0.75 % w / v, about 0.1 % w / v to about 1 % w / v, about 0.1 % w / v to about 1.5 % w / v, about 0.1 % w / v to about 2 % w / v, about 0.1 % w / v to about 2.5 % w / v, about 0.1 % w / v to about 3 % w / v, about 0.1 % w / v to about 4 % w / v, about 0.1 % w / v to about 5 % w / v, about 0.25 % w / v to about 0.5 % w / v, about 0.25 % w / v to about 0.75 % w / v, about 0.25 % w / v to about 1 % w / v, about 0.25 % w / v to about 1.5 % w / v, about 0.25 % w / v to about 2 % w / v, about 0.25 % w / v to about 2.5 % w / v, about 0.25 % w / v to about 3 % w / v, about 0.25 % w / v to about 4 % w / v, about 0.25 % w / v to about 5 % w / v, about 0.5 % w / v to about 0.75 % w / v, about 0.5 % w / v to about 1 % w / v, about 0.5 % w / v to about 1.5 % w / v, about 0.5 % w / v to about 2 % w / v, about 0.5 % w / v to about 2.5 % w / v, about 0.5 % w / v to about 3 % w / v, about 0.5 % w / v to about 4 % w / v, about 0.5 % w / v to about 5 % w / v, about 0.75 % w / v to about 1 % w / v, about 0.75 % w / v to about 1.5 % w / v, about 0.75 % w / v to about2 % w / v, about 0.75 % w / v to about 2.5 % w / v, about 0.75 % w / v to about 3 % w / v, about 0.75 % w / v to about 4 % w / v, about 0.75 % w / v to about 5 % w / v, about 1 % w / v to about 1.5 % w / v, about 1 % w / v to about 2 % w / v, about 1 % w / v to about 2.5 % w / v, about 1 % w / v to about 3 % w / v, about 1 % w / v to about 4 % w / v, about 1 % w / v to about 5 % w / v, about 1.5 % w / v to about 2 % w / v, about 1.5 % w / v to about 2.5 % w / v, about 1.5 % w / v to about 3 % w / v, about 1.5 % w / v to about 4 % w / v, about 1.5 % w / v to about 5 % w / v, about 2 % w / v to about 2.5 % w / v, about 2 % w / v to about 3 % w / v, about 2 % w / v to about 4 % w / v, about 2 % w / v to about 5 % w / v, about 2.5 % w / v to about 3 % w / v, about 2.5 % w / v to about 4 % w / v, about 2.5 % w / v to about 5 % w / v, about 3 % w / v to about 4 % w / v, about 3 % w / v to about 5 % w / v, or about 4 % w / v to about 5 % w / v, including increments therein. In some embodiments, a concentration of the physical quencher in the print chamber is about 0.05 % w / v, about 0.1 % w / v, about 0.25 % w / v, about 0.5 % w / v, about 0.75 % w / v, about 1 % w / v, about 1.5 % w / v, about2 % w / v, about2.5 % w / v, about3 % w / v, about4 % w / v, or about 5 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber is at least about 0.05 % w / v, about 0.1 % w / v, about 0.25 % w / v, about 0.5 % w / v, about 0.75 % w / v, about 1 % w / v, about 1.5 % w / v, about 2 % w / v, about 2.5 % w / v, about 3 % w / v, or about 4 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber is at most about 0.1 % w / v, about 0.25 % w / v, about 0.5 % w / v, about 0.75 % w / v, about 1 % w / v, about 1.5 % w / v, about2 % w / v, about2.5 % w / v, about3 % w / v, about 4 % w / v, or about 5 % w / v.

[0078] In some embodiments, as the movable stage (1230) is pulled away from the window once a layer of the fluid has been polymerized to a solid, and additional fluid is pulled to the top of the movable stage (1230) from within the print chamber, and contacts contact with thetran sparent walls (e.g., window). In some embodiments, the additional fluid moves around the exterior circumference of the movable stage in a gap between a wall of the print chamber as the polymerized fluid (e.g., printed object) is pulled down into the print chamber. In some embodiments, additional fluid is actively pumped into the print chamber through the fluid flow connection points (1260) as the polymerized fluid (e.g., printed object) is pulled down into the print chamber. In some embodiments, the moveable stage comprises one or more through holes, and the additional fluid moves through the through the through holes in the movable stage as the polymerized fluid (e.g., printed object) is pulled down into the print chamber. In some embodiments, the moveable stage comprises a textured surface to increase adhesion to the polymerized fluid and its movement with the movable stage. In some embodiments, the transparent wall (e.g., window) comprise a surface modification (e.g., smooth texturing, hydrophobic coating) which prevents adhesion between the polymerized fluid and the transparent wall (e.g., window), as to allow for movement of the polymerized fluid (e.g., printed object) with the movable stage, and for new unpolymerized fluid within the chamberto take the place of the object and contact the transparent wall (e.g., window). In some embodiments, as the movable stage (1230) is pulled away from the window, addition fluid is pulled into sealed print chamber (1250). In some embodiments, as the movable stage (1230) is pulled away from the window, additional fluid is moved to the top of print chamber (1250) in contact with the window, and the polymerized fluid (e.g., printed object).

[0079] In some embodiments, the print chamber is degassed. In some embodiments, the print chamber is completely filled to prevent movement of the fluid, for example, such as a completely filled water bottle, such that there is a substantially homogenous distribution of fluid within the print chamber. In some embodiments, the projection optics align at the movable stage (1230), and polymerize fluid which are illuminated by the projection optics (e.g., one or more light sources). In some embodiments, the movable stage comprises a textured movable stage (1230). In some embodiments, the textured movable stage (1230) provides more contact (e.g., surface) area to increase adhesion between the polymerized fluid on the movable stage (1230) vs adhesion between the polymerized fluid and one or more transparent walls of the print chamber (e.g., window).

[0080] In some embodiments, the additive manufacturing system comprises one or more fluid reservoirs (1220). In some embodiments, the removable cartridge (140) defines a second compartment comprises a plurality of fluid reservoirs (1220).

[0081] In some embodiments, the removable cartridge (1 0) comprises one or more fluid reservoirs (1220) within the second sealable compartment. In some embodiments, theremovable cartridge (140) comprises 1 fluid bag. In some embodiments, the removable cartridge (140) comprises 2 fluidbags. In some embodiments, the removable cartridge (140) comprises 3 fluid bags. In some embodiments, the removable cartridge (140) comprises 4 fluid bags. In some embodiments, the removable cartridge (140) comprises 5 fluid bags. In some embodiments, the removable cartridge (140) comprises 6 fluid bags.

[0082] In some embodiments, the removable cartridge (140) comprises one or more fluid reservoirs (1220), wherein the one or more fluid reservoir comprise a wash or perfusion solution.

[0083] In some embodiments, the removable cartridge (140) is prefilled with the desired photopolymerizable material in one fluid reservoir (1220), and the desired wash solution in another fluid reservoir (1220). In some embodiments, the plurality of fluid reservoirs (1220) comprise a wash fluid and a photopolymerizable fluid, wherein the outlet of fluid to at least one of the fluid reservoirs is to the fluid reservoir comprising the photopolymerizable fluid.

[0084] In some embodiments, the plurality of fluid reservoirs are removable and interchangeable within the removable cartridge. In some embodiments, after a print is complete, at least one fluid reservoir in removed and replaced with a replacement fluid reservoir. In some embodiments, the replacement fluid reservoir comprises a fluid, e.g., cell media, used to perfuse through the printed object, e.g., for culturing a tissue that was printed.

[0085] In some embodiments, the removable cartridge (140) is assembled and filled on earth. In some embodiments, the removable cartridge (140) is minimally modified once in space.

[0086] In some embodiments, after a print is complete, a first removable cartridge (140) is removed and replaced with a second removable cartridge. In some embodiments, the second removable cartridge comprises a different solution, e.g., cell media, than the first removable cartridge (140). In other embodiments, the second removable cartridge comprises the solution as the first removable cartridge (140).

[0087] In some embodiments, the removable cartridge (140) further comprises a perfusion path between the fluid reservoirs (1220) and the print chamber (1250) which provides an inlet of fluid to the print chamber (1250) from at least one of the fluid reservoirs (1220), and provides an outlet of fluid to at least one of the fluid reservoirs (1220). In some embodiments, the removable cartridge (140) further comprises a perfusion path between the fluid reservoirs (1220) and the print chamber (1250) which provides an inlet of fluid to the print chamber (1250) from atleast one of the fluid reservoirs (1220), and provides an outlet of fluid to at least one of the fluid reservoirs (1220). In some embodiments, the removable cartridge (140) is configured to perfuse the cell media through the newly printed construct.

[0088] In some embodiments, the plurality of fluid reservoirs (1220) are movable within the removable cartridge. In some embodiments, the plurality of fluid reservoirs (1220) are movable relative to the at least one light source, thereby allowing a larger exposure area than a single field of view of light form the light source onto the movable stage. In some embodiments, the removable cartridge (140) is moveable within the system along at least 2 of x-axis, a y-axis, and a z-axis.

[0089] In some embodiments, the plurality of fluid reservoirs comprise a photopolymerizable material, a photo initiator, a chemical quencher, (2,2,6,6-Tetramethylpiperidin-l-yl)oxyl, (2,2,6,6-tetramethylpiperidin-l-yl)oxidanyl (TEMPO), a physical quencher, a wash solution, a biologic solution, a suspension of cells, microparticles, nanoparticles, therapeutic loaded nanoparticles, therapeutic loaded microparticles, electrolytic or electrically conductive materials, or combinations thereof. In some embodiments, at least one of the plurality of fluid reservoirs comprise a photopolymerizable material, a photo initiator, a chemical quencher, (2,2,6,6-Tetramethylpiperidin-l-yl)oxyl, (2,2,6,6-tetramethylpiperidin-l-yl)oxidanyl (TEMPO), a physical quencher, a wash solution, a biologic solution, a suspension of cells, microparticles, nanoparticles, therapeutic loaded nanoparticles, therapeutic loaded microparticles, electrolytic or electrically conductive materials, or combinations thereof. In some embodiments, the wash solution comprises water, a buffer, an organic material a solution containing antibiotics, an antimicrobial agent, or combinations thereof. In some embodiments, the biologic solution comprises nutrients and / or growth factors suitable for cell culture and proliferation. In some embodiments, the biologic solution comprises a suspension of cells. In some embodiments, the plurality of fluid reservoirs (1220) comprise a chemical quencher that quenches free-radicals and prevents polymerization, or over polymerization. In some embodiments, the plurality of fluid reservoirs (1220) comprise a physical quencher that quenches the light intensity of the light source and prevents polymerization, or over polymerization. In some embodiments, the plurality of fluid reservoirs (1220) comprises a cell media fluid reservoir.

[0090] In some embodiments, one fluid reservoir may be utilized. In some embodiments, more than one fluid reservoir may be utilized. Each reservoir may contain a different photopolymerizable material or other solution to be utilized during fabrication. In some multiple fluid reservoir embodiments, some of the fluid reservoirs may comprise the same solution. In some embodiments, the fluid reservoir may be considered as a combination of multiple reservoirs, and each individual reservoir may be considered as a subreservoir or subcomponent of the fluid reservoir.

[0091] FIG. 7A-7B depict example embodiments of additive manufacturing system print chamber (1250). In some embodiments, the removable cartridge (140) defines a second compartment. FIG. 7A depicts an embodiment of an additive manufacturing system print chamber tall version. FIG. 7B depicts an embodiment of an additive manufacturing system print chamber (1250) short version. In some embodiments, the removable cartridge (140) defines a second sealable compartment comprising a print chamber (1250). In some embodiments, the removable cartridge (140) defines a second compartment comprises a print chamber (1250) tall version, wherein the movable stage plunger (1310) is approximately 10- 20 cm inches in height. In some embodiments, the removable cartridge (140) defines a second compartment comprises a print chamber (1250) short version, wherein the movable stage plunger (1310) is approximately less than 10 inches in height.

[0092] In some embodiments, the print chamber (1250) comprises fluid flow connection points (1260), and a movable stage plunger (1310). In some embodiments, the print chamber (1250) defines a third sealable compartment, and is hermetically sealed from the second compartment. In some embodiments, the print chamber (1250) sealed such that the seal is liquid tight from the second compartment. In some embodiments, the print chamber (1250) sealed such that the seal is gas tight from the second compartment. In some embodiments, the print chamber (1250) is sealed such that the seal is impermeable to liquid from the second compartment. In some embodiments, the print chamber (1250) is sealed, wherein a completely filled print chamber will not leak into the second compartment. In some embodiments, the print chamber (1250) is fully filled with a liquid and comprises essentially no gas. In some embodiments, the print chamber is configured to maintain a liquid seal within the print chamber, and prevent entry of gas into the print chamber from the second compartment.

[0093] In some embodiments, the print chamber (1250) is empty. In some embodiments, the print chamber (1250) is empty at the beginning of a print sequence. In some embodiments, the print chamber (1250) is empty and without gas at the beginning of a print sequence. In some embodiments, the print chamber (1250) fills with a fluid before or during the printing sequence. In some embodiments, the print chamber (1250) completely fills with a liquid and is degassed prior to initiation of a print sequence, such that it comprises essentially no gas, or no gas.

[0094] In some embodiments, the removable cartridge (140) further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage (1230).

[0095] In some embodiments, the removable cartridge (140) defining a second compartment comprises one or more transparent walls (1430). In some embodiments, the one or more transparent walls (1430) permitfor transmission of light into the removable cartridge (140).

[0096] In some embodiments, the print chamber comprises one or more transparent walls (1235) permitting for the transmission of light to a liquid within the print chamber. In some embodiments, the one or more transparent walls (1235) of the print chamber comprise an anti-adhesion surface configured to reduce adhesion between a fluid or solid in contact with the anti-adhesion surface. In some embodiments, the anti-adhesion surface comprises a coating of a hydrophobic or a superhydrophobic material. In some embodiments, the antiadhesion surface comprises a micropattern or nanopattern to induce hydrophobic or superhydrophobic properties. In some embodiments, the micropattem or nanopattem comprises dimensions of, of about, of at most, or at most about 500 nm, about 490 nm, about 480 nm, about 470 nm, about460 nm, about450 nm, about 440 nm, about430 nm, about 420 nm, about 410 nm, about 400 nm, about 390 nm, about 380 nm, about 370 nm, about 360 nm, about 350 nm, about 340 nm, about 330 nm, about 320 nm, about 310 nm, about 300 nm, about 290 nm, about 280 nm, about 270 nm, about 260 nm, about 250 nm, about 240 nm, about 230 nm, about 220 nm, about 210 nm, about 200 nm, about 190 nm, about 180 nm, about 170 nm, about 160 nm, about 150 nm, about 140 nm, about 130 nm, about 120 nm, about 110 nm, about 100 nm, about 90 nm, about 80 nm, about 70 nm, about 60 nm, about 50 nm, about 40 nm, about 30 nm, about 20 nm, or about 10 nm, or any range of values therebetween. For example, in some embodiments, the micropattern or nanopattern comprises dimensions in any one of the following ranges: 10 nm - 500 nm, 10 nm - 400 nm, 10 nm - 300 nm, 10 nm - 200 nm, 10 nm - 100 nm, or 100 nm - 300 nm.

[0097] In some embodiments, the one or more transparent walls (1235) comprise a gas inlet. In some embodiments, the one or more transparent walls (1235) are gas permeable. In some embodiments, the gas permeability permits an active or passive flow of oxygen. In some cases, the flow of oxygen through a gas permeable transparent wall (1235) of the print chamber can establish a dead zone adjacent to the transparent wall (1235) where no fluid can be photopolymerized. In some embodiments, the dead zone adjacent to the transparent wall (1235) prevents adhesion of a 3D printed object to the transparent wall (1235), and permits for the movement of the 3D printed object with the movable stage and further into the print chamber upon polymerization of fluid which forms the 3D printed objects. In some embodiments, the one or more transparent walls (1235) comprises glass, quartz, transparentplastic, silicon, silica, poly dimethylsiloxane (PDMS), transparent ceramic, fluorinated ethylene propylene (FEP), or combinations thereof.

[0098] In some embodiments, the removable cartridge (140) comprises a pressurized oxygen source. In some embodiments, the pressurized oxygen source is coupled to an oxygen line. In some embodiments, the oxygen line is directly coupled to the one or more transparent walls. In some embodiments, the removable cartridge (1 0) comprises a gas inlet directing gas proximally to the one or more transparent walls (1430). In some embodiments, the one or more transparent walls (1430) are gas permeable. In some embodiments, the one or more transparent walls (1430) are configured to provide an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring against the one or more transparent walls (1430). In some embodiments, the additive manufacturing system comprises one or more fans configured to direct an active flow of oxygen to the one or more transparent walls. In some embodiments, the active flow is achieved via pressurized air, fans or a combination thereof.

[0099] FIG. 7A-7D depicts print chamber tall defining a third sealable compartment, of the additive manufacturing system of present embodiments. Shown are tall and short version with the movable stage (1230) at the top and bottom positions, according to some embodiments. In some embodiments, the print chamber (1250) comprises a moveable stage (1230) upon which a 3D object is printed and adheres to upon polymerization of fluid. In some embodiments, the removable cartridge (140) defining a second compartment comprises a movable stage (1230). In some embodiments, the removable cartridge (140) defining a second compartment comprises a movable stage (1230) positioned within the print chamber (1250).

[0100] In some embodiments, the moveable stage (1230) comprises a patterned surface configured to enhance adhesion between the patterned surface and a fluid or a solid in contact with the patterned surface. In some embodiments, the removable cartridge (140) comprises a plurality of movable stages. In some embodiments, wherein the removable cartridge (140) comprises a plurality of movable stages (1230), each movable stage (1230) is in line with at least one of the one or more transparent walls.

[0101] In some embodiments, the additive manufacturing system is configured to move the movable stage (1230). In some embodiments, the removable cartridge (140) is configured to move the movable stage (1230). In some embodiments, moving the movable stage (1230) comprises moving the movable stage (1230) away from the optical plane such that the projected two-dimensional cross-sections of the three-dimensional object are incidenton photopolymerizable fluid in contact with and adjacent to previously formed layers of the object. In some embodiments, moving the movable stage (1230) comprises moving the movable stage (1230) continuously. In some embodiments, the rate at which the movable stage is moved is determined by a combination of factors e.g., formulation of photopolymer, photopolymer ingredients, the brightness of the light source, or a combination thereof.

[0102] In some embodiments, the movable stage (1230) is moved at rates of at least about 0.1 mm / min. In some embodiments, the movable stage is moved at rates up to about 0.5 mm / min. In some embodiments, the movable stage is moved at rates between about 0.1 mm / min to about 0.5 mm / min. In some embodiments, the movable stage is moved at rates of 0.1, 0.2, 0.3, 0.4 or 0.5 mm / min. In some embodiments, the movable stage is moved at rates of at least about 0.1 mm / min to about 2 mm / min. In some embodiments, the movable stage is moved at rates of at least about 0.1 mm / min to about 0.2 mm / min, about 0.1 mm / min to about 0.3 mm / min, about 0.1 mm / min to about 0.4 mm / min, about 0.1 mm / min to about 0.5 mm / min, about 0.1 mm / min to about 0.6 mm / min, about 0.1 mm / min to about 0.7 mm / min, about 0.1 mm / min to about 0.8 mm / min, about 0.1 mm / min to about 0.9 mm / min, about 0.1 mm / min to about 1 mm / min, about 0.1 mm / min to about 1.5 mm / min, about 0.1 mm / min to about 2 mm / min, about 0.2 mm / min to about 0.3 mm / min, about 0.2 mm / min to about 0.4 mm / min, about 0.2 mm / min to about 0.5 mm / min, about 0.2 mm / min to about 0.6 mm / min, about 0.2 mm / min to about 0.7 mm / min, about 0.2 mm / min to about 0.8 mm / min, about 0.2 mm / min to about 0.9 mm / min, about 0.2 mm / min to about 1 mm / min, about 0.2 mm / min to about 1.5 mm / min, about 0.2 mm / min to about 2 mm / min, about 0.3 mm / min to about 0.4 mm / min, about 0.3 mm / min to about 0.5 mm / min, about 0.3 mm / min to about 0.6 mm / min, about 0.3 mm / min to about 0.7 mm / min, about 0.3 mm / min to about 0.8 mm / min, about 0.3 mm / min to about 0.9 mm / min, about 0.3 mm / min to about 1 mm / min, about 0.3 mm / min to about 1.5 mm / min, about 0.3 mm / min to about 2 mm / min, about 0.4 mm / min to about 0.5 mm / min, about 0.4 mm / min to about 0.6 mm / min, about 0.4 mm / min to about 0.7 mm / min, about 0.4 mm / min to about 0.8 mm / min, about 0.4 mm / min to about 0.9 mm / min, about 0.4 mm / min to about 1 mm / min, about 0.4 mm / min to about 1.5 mm / min, about 0.4 mm / min to about 2 mm / min, about 0.5 mm / min to about 0.6 mm / min, about 0.5 mm / min to about 0.7 mm / min, about 0.5 mm / min to about 0.8 mm / min, about 0.5 mm / min to about 0.9 mm / min, about 0.5 mm / min to about 1 mm / min, about 0.5 mm / min to about 1.5 mm / min, about 0.5 mm / min to about 2 mm / min, about 0.6 mm / min to about 0.7 mm / min, about 0.6 mm / min to about 0.8 mm / min, about 0.6 mm / min to about 0.9 mm / min, about 0.6 mm / min to about 1 mm / min, about 0.6 mm / min to about 1.5 mm / min, about 0.6 mm / min to about 2 mm / min,about 0.7 mm / min to about 0.8 mm / min, about 0.7 mm / min to about 0.9 mm / min, about 0.7 mm / min to about 1 mm / min, about 0.7 mm / min to about 1.5 mm / min, about 0.7 mm / min to about 2 mm / min, about 0.8 mm / min to about 0.9 mm / min, about 0.8 mm / min to about 1 mm / min, about 0.8 mm / min to about 1.5 mm / min, about 0.8 mm / min to about 2 mm / min, about 0.9 mm / min to about 1 mm / min, about 0.9 mm / min to about 1.5 mm / min, about 0.9 mm / min to about 2 mm / min, about 1 mm / min to about 1.5 mm / min, about 1 mm / min to about 2 mm / min, or about 1.5 mm / min to about 2 mm / min, including increments therein. In some embodiments, the movable stage is moved at rates of at least about 0.1 mm / min, about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, about 1.5 mm / min, or about 2 mm / min. In some embodiments, the movable stage is moved at rates of at least at least about 0.1 mm / min, about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, or about 1.5 mm / min. In some embodiments, the movable stage is moved at rates of at least at most about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, about 1.5 mm / min, or about 2 mm / min.

[0103] In some embodiments, the 3D object is manufactured at rates of at least about 0.1 mm / min to about 2 mm / min. In some embodiments, the 3D object is manufactured at rates of at least about 0.1 mm / min to about 0.2 mm / min, about 0.1 mm / min to about 0.3 mm / min, about 0.1 mm / min to about 0.4 mm / min, about 0.1 mm / min to about 0.5 mm / min, about 0.1 mm / min to about 0.6 mm / min, about 0.1 mm / min to about 0.7 mm / min, about 0.1 mm / min to about 0.8 mm / min, about 0.1 mm / min to about 0.9 mm / min, about 0.1 mm / min to about 1 mm / min, about 0.1 mm / min to about 1.5 mm / min, about 0.1 mm / min to about 2 mm / min, about 0.2 mm / min to about 0.3 mm / min, about 0.2 mm / min to about 0.4 mm / min, about 0.2 mm / min to about 0.5 mm / min, about 0.2 mm / min to about 0.6 mm / min, about 0.2 mm / min to about 0.7 mm / min, about 0.2 mm / min to about 0.8 mm / min, about 0.2 mm / min to about 0.9 mm / min, about 0.2 mm / min to about 1 mm / min, about 0.2 mm / min to about 1.5 mm / min, about 0.2 mm / min to about 2 mm / min, about 0.3 mm / min to about 0.4 mm / min, about 0.3 mm / min to about 0.5 mm / min, about 0.3 mm / min to about 0.6 mm / min, about 0.3 mm / min to about 0.7 mm / min, about 0.3 mm / min to about 0.8 mm / min, about 0.3 mm / min to about 0.9 mm / min, about 0.3 mm / min to about 1 mm / min, about 0.3 mm / min to about 1.5 mm / min, about 0.3 mm / min to about 2 mm / min, about 0.4 mm / min to about 0.5 mm / min, about 0.4 mm / min to about 0.6 mm / min, about 0.4 mm / min to about 0.7 mm / min, about 0.4mm / min to about 0.8 mm / min, about 0.4 mm / minto about 0.9 mm / min, about 0.4 mm / min to about 1 mm / min, about 0.4 mm / min to about 1.5 mm / min, about 0.4 mm / min to about 2 mm / min, about 0.5 mm / min to about 0.6 mm / min, about 0.5 mm / min to about 0.7 mm / min, about 0.5 mm / min to about 0.8 mm / min, about 0.5 mm / min to about 0.9 mm / min, about 0.5 mm / min to about 1 mm / min, about 0.5 mm / min to about 1.5 mm / min, about 0.5 mm / min to about 2 mm / min, about 0.6 mm / min to about 0.7 mm / min, about 0.6 mm / min to about 0.8 mm / min, about 0.6 mm / min to about 0.9 mm / min, about 0.6 mm / min to about 1 mm / min, about 0.6 mm / min to about 1.5 mm / min, about 0.6 mm / min to about 2 mm / min, about 0.7 mm / min to about 0.8 mm / min, about 0.7 mm / minto about 0.9 mm / min, about 0.7 mm / min to about 1 mm / min, about 0.7 mm / min to about 1.5 mm / min, about 0.7 mm / min to about 2 mm / min, about 0.8 mm / min to about 0.9 mm / min, about 0.8 mm / min to about 1 mm / min, about 0.8 mm / min to about 1.5 mm / min, about 0.8 mm / min to about 2 mm / min, about 0.9 mm / min to about 1 mm / min, about 0.9 mm / min to about 1.5 mm / min, about 0.9 mm / min to about 2 mm / min, about 1 mm / min to about 1 .5 mm / min, about 1 mm / min to about 2 mm / min, or about 1.5 mm / min to about 2 mm / min, including increments therein. In some embodiments, the 3D object is manufactured at rates of at least about 0.1 mm / min, about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, about 1.5 mm / min, or about 2 mm / min. In some embodiments, the 3D object is manufactured at rates of at least at least about 0.1 mm / min, about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, or about 1.5 mm / min. In some embodiments, the 3D object is manufactured at rates of at least at most about 0.2 mm / min, about 0.3 mm / min, about 0.4 mm / min, about 0.5 mm / min, about 0.6 mm / min, about 0.7 mm / min, about 0.8 mm / min, about 0.9 mm / min, about 1 mm / min, about 1.5 mm / min, or about 2 mm / min.

[0104] In some embodiments, moving the movable stage comprises moving the movable stage incrementally. In some embodiments, the movable stage is capable of moving in increments of about 1 micron. In some embodiments, the movable stage is capable of moving in increments of about 0.001 microns to about 10 microns. In some embodiments, the movable stage is capable of moving in increments of about 0.001 microns to about 0.01 microns, about 0.001 microns to about 0.1 microns, about 0.001 microns to about 0.5 microns, about 0.001 microns to about 1 micron, about 0.001 microns to about 2 microns, about 0.001 microns to about 5 microns, about 0.001 microns to about 10 microns, about 0.01 microns to about 0.1 microns, about 0.01 microns to about 0.5 microns, about 0.01 microns toabout 1 micron, about 0.01 microns to about 2 microns, about 0.01 microns to about 5 microns, about O.Ol microns to about 10 microns, about 0.1 microns to about 0.5 microns, about 0.1 microns to about 1 micron, about 0. 1 microns to about 2 microns, about 0.1 microns to about 5 microns, about 0.1 microns to about 10 microns, about 0.5 microns to about 1 micron, about 0.5 microns to about 2 microns, about 0.5 micronsto about 5 microns, about 0.5 microns to about 10 microns, about 1 micron to about 2 microns, about 1 micron to about 5 microns, about 1 micron to about 10 microns, about 2 microns to about 5 microns, about 2 microns to about 10 microns, or about 5 microns to about 10 microns. In some embodiments, the movable stage is capable of moving in increments of about 0.001 microns, about O.Ol microns, about 0.1 microns, about 0.5 microns, about 1 micron, about 2 microns, about 5 microns, or about 10 microns. In some embodiments, the movable stage is capable of moving in increments of at least about 0.001 microns, about O.Ol microns, about 0.1 microns, about 0.5 microns, about 1 micron, about 2 microns, or about 5 microns. In some embodiments, the movable stage is capable of moving in increments of at most about 0.01 microns, about 0.1 microns, about 0.5 microns, about 1 micron, about 2 microns, about 5 microns, or about 10 microns.

[0105] In some embodiments, the movable stage (1230) comprises one or more perfusable through holes. In some embodiments, the one or more perfusable through holes provided in the movable stage have a diameter of about 1 millimeter (mm). In some embodiments, the one or more perfusable through holes provided in the movable stage have a diameter of about 0.05 mm to about 10 mm. In some embodiments, the one or more perfusable through holes provided in the movable stage have a diameter of about 0.05 mm to about 0.1 mm, about 0.05 mm to about 0.5 mm, about 0.05 mm to about 1 mm, about 0.05 mm to about 2 mm, about 0.05 mm to about 3 mm, about 0.05 mm to about 5 mm, about 0.05 mm to about 10 mm, about 0.1 mm to about 0.5 mm, about 0. 1 mm to about 1 mm, about 0.1 mm to about 2 mm, about 0.1 mm to about 3 mm, about 0.1 mm to about 5 mm, about 0.1 mm to about 10 mm, about 0.5 mm to about 1 mm, about 0.5 mm to about 2 mm, about 0.5 mm to about 3 mm, about 0.5 mm to about 5 mm, about 0.5 mm to about 10 mm, about 1 mm to about 2 mm, about 1 mm to about 3 mm, about 1 mm to about 5 mm, about 1 mm to about 10 mm, about 2 mm to about 3 mm, about 2 mm to about 5 mm, about 2 mm to about 10 mm, about 3 mm to about 5 mm, about 3 mm to about 10 mm, or about 5 mm to about 10 mm. In some embodiments, the one or more perfusable through holes provided in the movable stage have a diameter of about 0.05 mm, about 0.1 mm, about 0.5 mm, about 1 mm, about 2 mm, about 3 mm, about 5 mm, or about 10 mm. In some embodiments, the one or more perfusablethrough holes provided in the movable stage have a diameter of at least about 0.05 mm, about 0.1 mm, about 0.5 mm, about 1 mm, about 2 mm, about 3 mm, or about 5 mm. In some embodiments, the one or more perfusable through holes provided in the movable stage have a diameter of at most about 0.1 mm, about 0.5 mm, about 1 mm, about 2 mm, about 3 mm, about 5 mm, or about 10 mm.

[0106] In some embodiments, the movable stage (1230) comprises about 1 through hole to about 20 through holes. In some embodiments, the movable stage comprises about 1 through hole to about 2 through holes, about 1 through hole to about 3 through holes, about 1 through hole to about 5 through holes, about 1 through hole to about 7 through holes, about 1 through hole to about 10 through holes, about 1 through hole to about 15 through holes, about 1 through hole to about 20 through holes, about 2 through holes to about 3 through holes, about 2 through holes to about 5 through holes, about 2 through holes to about 7 through holes, about 2 through holes to about 10 through holes, about 2 through holes to about 15 through holes, about 2 through holes to about 20 through holes, about 3 through holes to about 5 through holes, about 3 through holes to about 7 through holes, about 3 through holes to about 10 through holes, about 3 through holes to about 15 through holes, about 3 through holes to about 20 through holes, about 5 through holes to about 7 through holes, about 5 through holes to about 10 through holes, about 5 through holes to about 15 through holes, about 5 through holes to about 20 through holes, about 7 through holes to about 10 through holes, about 7 through holes to about 15 through holes, about 7 through holes to about 20 through holes, about 10 through holes to about 15 through holes, about 10 through holes to about 20 through holes, or about 15 through holes to about 20 through holes. In some embodiments, the movable stage comprises about 1 through hole, about 2 through holes, about 3 through holes, about 5 through holes, about 7 through holes, about 10 through holes, about 15 through holes, or about 20 through holes. In some embodiments, the movable stage comprises at least about 1 through hole, about 2 through holes, about 3 through holes, about 5 through holes, about 7 through holes, about 10 through holes, or about 15 through holes. In some embodiments, the movable stage comprises at most about 2 through holes, about 3 through holes, about 5 through holes, about 7 through holes, about 10 through holes, about 15 through holes, or about 20 through holes. The through holes may all be of a uniform size or may differ in size.

[0107] In some embodiments, the through holes provide passages for unpolymerized material to pass through the movable stage. In such embodiments, the apertures allow for the photopolymerizable material to fill in above the previously formed layer of the 3D objectbeing fabricated, quicker than if no holes or perfusions are provided in the movable stage. In some embodiments, the apertures allow for the photopolymerizable material of high viscosities to fill in above the previously formed layer of the 3D object being fabricated, more quickly than if no holes or perfusions are provided in the movable stage.

[0108] In some embodiments, the movable stage is raised or lowered up to 10 centimeters (cm). In some embodiments, the range of movement of the movable stage is about 5 cm to about 20 cm. In some embodiments, the range of movement of the movable stage is about 5 cm to about 7 cm, about 5 cm to about 10 cm, about 5 cm to about 15 cm, about 5 cm to about 20 cm, about 7 cm to about 10 cm, about 7 cm to about 15 cm, about 7 cm to about 20 cm, about 10 cm to about 15 cm, about 10 cm to about 20 cm, or about 15 cm to about 20 cm. In some embodiments, the range of movement of the movable stage is about 5 cm, about 7 cm, about 10 cm, about 15 cm, or about 20 cm. In some embodiments, the range of movement of the movable stage is at least about 5 cm, about 7 cm, about 10 cm, or about 15 cm. In some embodiments, the range of movement of the movable stage is at most about 7 cm, about 10 cm, about 15 cm, or about 20 cm.

[0109] In some embodiments, the movable stage (1230) descends. In some embodiments, the movable stage (1230) descends within the print chamber (1250). In some embodiments, the movable stage (1230) is at the top of the print chamber (1250) at the beginning of a print sequence. In some embodiments, the movable stage (1230) is at the bottom of the print chamber (1250) at the end of the print sequence. In some embodiments, the fluid polymerized is on top of the movable stage (1230). In some embodiments, as the movable stage (1230) moves down, fluid flows over the top of the movable stage. In some embodiments, as fluid flows over the top of the movable stage (1230) the fluid is polymerized by the at least one light source.

[0110] In some embodiments, wherein the volume has been removed from the system, the movable stage plunger (1310) draws fluid from the fluid reservoir (1220), e.g., bio ink fluid reservoir. In some embodiments, the print chamber comprises a movable stage plunger (1310). In some embodiments, extending the movable stage plunger (1310) requires replacement of fluid, e.g., from the fluid reservoir (1220).

[0111] In some embodiments, the plurality of fluid reservoirs (1220) comprises a wash fluid, a photopolymerizable fluid, a biologies fluid, or a combination thereof. In some embodiments, the removable cartridge (140) comprises a perfusion path between the fluid reservoirs (1220) and the print chamber. In some embodiments, the perfusion path provides an inlet of fluid to the print chamber (1250) with the inlet aligning with an opening in themoveable stage (1230). In some embodiments, the channels align with the inlet. In some embodiments, the plurality of channels are flushed with the wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels.

[0112] In some embodiments, the removable cartridge (140) further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber (1250) from at least one of the fluid reservoirs (1220), and provides an outlet of fluid to at least one of the fluid reservoirs ( 1220). In some embodiments, flushing the print chamber (1250) with the wash fluid removes the photopolymerizable fluid, and directs the wash fluid and the removed photopolymerizable fluid into one of the fluid reservoirs (1220) initially holding the photopolymerizable fluid.

[0113] In some embodiments, the system comprises a perfusion apparatus. In some embodiments, the perfusion apparatus comprises a pump. In some embodiments, the pump comprises one or more reservoirs for retaining a solution. In some embodiments, the perfusion apparatus is configured to remove unpolymerized material from a lumen or passage of the 3D fabricated object. In some embodiments, the perfusion apparatus is configured to introduce a solution into the lumen or passage of the 3D fabricated object.

[0114] In some embodiments, the perfusion apparatus is provided with an inlet conduit for introducing a solution to the fabricated object. In some embodiments, the terminal end of the inlet conduit is provided with an inlet syringe to facilitate insertion into a lumen or passage of the fabricated object. In some embodiments, the inlet syringe passes through and aperture provided in the movable stage (1230) prior to entering an aperture of a lumen or passage of the fabricated object.

[0115] In some embodiments, the perfusion apparatus is provided with an outlet conduit for removing a solution or unpolymerized material from the fabricated object. In some embodiments, the terminal end of the outlet conduit is provided with an outlet syringe to facilitate insertion into a lumen or passage of the fabricated object. In some embodiments, the outlet syringe passes through and aperture provided in the movable stage (1230) prior to entering an aperture of a lumen or passage of the fabricated object. In some embodiments, the outlet conduit for removing a solution or fluid from the system if fluidically coupled to the one or more fluid reservoir. In some embodiments, the one or more fluid reservoirs are reused as waste reservoirs following completion of a print sequence. In some embodiments, the fluid reservoirs reused as waste retain solutions (e.g., wash fluids) being pumped into print chamber following fabrication of an to prevent contamination of a workspace. In some embodiments, where the fabricated object is a biomimetic implant, the reservoir maycomprise a solution to coat the implant to sterilize or to otherwise prepare the implant for insertion into a living being in need thereof.

[0116] In some embodiments, after fabrication, 3D fabricated objects having a lumen or passage can be uniformly filled with stem cells, blood, synthetic blood, cell media, cells, particles, or other desirable solutions using the perfusion apparatus. In some embodiments, the lumen or passage can be filled with neural stem cells, hematopoietic stem cells, mesenchymal stem cells, epithelial stem cells, endothelial stem cells, intestinal stem cells, muscle stem cells, satellite cells, dermal stem cells, or a combination thereof. In some embodiments, the lumen or passage are uniformly filled with stem cells. In some embodiments, the lumen or passage are filled after the 3d object is fabricated. In some embodiments, after fabrication, channels can be uniformly filled with neural stem cells. In some embodiments, stem cell-derived cells in the channels can express a neuronal marker, such as, but not limited to, Hu or NeuN. In other embodiments, the stem cell -derived cells in the channels can express an oligodendrocyte marker, such as but not limited to, Olig2 or an astrocyte marker such as, but not limited to, GFAP. In some embodiments, the cells can express two or more of the above.

[0117] In some embodiments, the interior walls of a 3D fabricated object are coated with stem cells, blood, synthetic blood, cell media, cells, particles, or other desirable solutions or suspensions using the perfusion apparatus. In some embodiments, the exterior walls of a 3D fabricated object are coated with stem cells, blood, synthetic blood, cell media, cells, particles, or other desirable solutions or suspensions using the perfusion apparatus.

[0118] In some embodiments, the perfusion path comprises one or more pumps coupled to the fluid reservoirs and the print chamber. In some embodiments, the additive manufacturing system further comprises a pump system. In some embodiments, the pump system comprises one or more pumps having fluid reservoirs (1 0) contained within. In some embodiments, each pump may be connected to a single fluid reservoir (140). In some embodiments, a first pump is connected to the first fluid reservoir via a first fluid line and first reservoir interface, such that the first pump is in fluid communication with the first fluid reservoir. In some embodiments, a second pump may be connected to the second fluid reservoir via a second fluid line and second reservoir interface, such that the second pump is in fluid communication with the second fluid reservoir.

[0119] In some embodiments, the first pump and first interface are configured to fill to the first fluid reservoir with a solution contained within a reservoir of the first pump. Insome embodiments, the first pump and first interface are configured to remove a solution from the first fluid reservoir and transport the solution to a reservoir within the first pump.

[0120] In some embodiments, the second pump and second interface are configured to fill to the second fluid reservoir with a solution contained within a reservoir of the second pump. In some embodiments, the second pump and second interface are configured to remove a solution from the second fluid reservoir and transport the solution to a reservoir within the second pump.

[0121] In some embodiments, the fluid pumps comprise more than one reservoir to store more than one solution. In some embodiments, while the 3D object is being fabricated in one reservoir, the pump system removes and fills solution in the reservoir not being utilized for fabrication. In some embodiments, wherein fluid reservoirs are utilized, each reservoir being connected to a pump system, a first print sequence may be complete within a first reservoir wherein the first print sequence utilizes a first photopolymerizable material. In some embodiments, after completion of the first sequence, the partially fabricated 3D object may be transferred to the second reservoir to undergo a second print sequence. In some embodiments, as the second print sequence is underway, a pump system may remove the first photopolymerizable material from the first reservoir, then fill the first reservoir with a different solution or photopolymerizable material. In some embodiments, once the second print sequence is complete, the first reservoir will then be ready, such that the object being fabricated can be transferred back to the first reservoir to begin a new processing stage within the first reservoir. In some embodiments, although the example is provided with two reservoirs, it is possible to conduct similar processing with any embodiment wherein more than one fluid reservoiris provided. In some embodiments, the processing method allows for little delay between processing steps taking place with different solution to reduce overall production time of a 3D object being fabricated. In some embodiments, an object can be fabricated with multiple materials in the same build chamber that is connected to multiple fluid reservoirs containing multiple materials. In some embodiments, the same cartridge system can contain multiple build chambers, each with their own plurality of fluid reservoirs, and allow the one or more light sources to fabricate a three-dimensional structure within more than one build chamber in a single print sequence.

[0122] FIG. 5A-5B depict example embodiments of the additive manufacturing internal subsystem. FIG. 5A depicts a partial side perspective view of an embodiment of an additive manufacturing internal subsystem. FIG. 5B depicts a fourth side perspective view of an embodiment of an additive manufacturing internal subsystem. In some embodiments, theadditive manufacturing system (130) comprises one or more light sources. In some embodiments, the system comprises linear axis (910) to control build plane motion, a mirror (920), a shutter (930), projection optics (940), and a projector (950). In some embodiments, the projector (950) contains at least one light source and at least one spatial light modulator.

[0123] In some embodiments, the one or more light sources are movable within the first compartment (111). In some embodiments, the at least one light source comprises a light emitting diode (LED). In some embodiments, the at least one light source comprises a laser.

[0124] In some embodiments, the at least one light source is movable along an x-axis, and a y-axis. In some embodiments, the at least one light source is movable along an x-axis, a y-axis, and a z-axis. In some embodiments, wherein the at least one light source is movable along two or more of an x-axis, a y-axis, and a z-axis relative to the removable cartridge such that multiple optical field-of-views can be exposed onto the at least one movable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the multiple optical field-of-views are configured to print multiple 3D objects simultaneously. In some embodiments, the multiple optical field-of-views allow for multiple prints to occur in one or more build chambers during the same print sequence. In some embodiments, the at least one light source comprises a plurality of spatial light modulators configured to direct light from the one or more light sources. In some embodiments, the plurality of spatial light modulators are configured to direct light along an x-axis, and a y-axis. In some embodiments, the plurality of spatial light modulators are configured to direct light along an x-axis, a y-axis, and a z-axis. In some embodiments, the plurality of spatial light modulators comprise one or more digital micro-mirror devices. In some embodiments, the plurality of spatial light modulators comprise one or more liquid crystal devices. In some embodiments, the plurality of spatial light modulators display the cross section of the first layer to be printed.

[0125] In some embodiments, the additive manufacturing system is configured to direct the light. In some embodiments, the additive manufacturing system is configured to direct two-dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage. In some embodiments, the additive manufacturing system is configured to move the one or more light sources relative to the movable stage and expose multiple optical field-of-views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the additive manufacturing system is configured to intermittently activate the one or more light sources during the formation of the three-dimensional object.

[0126] In some embodiments, the light source is a divergent light source. The divergent light source may be comprised of a light emitting diode (LED) or a plurality of LEDs. In some embodiments, other divergent light sources are utilized. In some embodiments, the light projector is a digital light projector which utilizes a liquid crystal panel or digital micromirror array to create a digital photomask.

[0127] Digital light processing (DLP) methods may allow the user to digitally control an array of micromirrors to expose an entire plane of photo-active material at once. By programming each micromirror individually, a user can selectively expose the layer of photo - active material to a predefined pattern with a spatial resolution corresponding to the individually programmed pixels (micromirrors). An entire three-dimensional object may be fabricated through the incremental motion of a stage coordinated with the successive exposure of multiple two-dimensional polymerized layers.

[0128] In some embodiments, the MCU connects the computing device to the controller of the digital light projector (DLP). In some embodiments, the DLP controller sends pattern data and a clocking pulse to the digital micromirror device (DMD) to activate specific micro mirrors, manipulating light from the light source to project the pattern of the two-dimensional cross-section for the 3D object being fabricated. In some embodiments, an analog DMD control is provided, wherein the analog control converts the digital signal from the digital controller prior to sending the signal to the DMD.

[0129] In some embodiments, the DMD is replaced by a liquid crystal display (LCD) screen. Similar to embodiments using a DMD, the LCD is manipulated by controller to project two-dimensional cross-sections of the 3D object being fabricated.

[0130] In some embodiments, the light projector further comprises a collimating optical system comprised of one or more optical lenses to collimate light from the light source onto the digital micromirror device or liquid crystal display. In some embodiments, the light projector further comprises a focal optical system comprised of one or more optical lenses to focus a light pattern emitted by the digital micromirror device or LCD onto an incident plane. In some embodiments, the optical lenses are disposed between the spatial light modulator and the incident plane.

[0131] In some embodiments, the incident plane is provided within a fluid reservoir such that the light pattern representing a two-dimensional cross-section of the object being fabricated is incident on unpolymerized material provided within the fluid reservoir. In some embodiments, the incident plane is provided within a build chamber containing photopolymerizable material and coupled to one of more fluid reservoirs such that the lightpattern representing a two-dimensional cross-section of the object being fabricated is incident on unpolymerized material provided within the build chamber. Upon exposure to the patterned light, the photopolymerizable material begins cross-linking and solidifying to form a layer of the 3D object being fabricated. In some embodiments, the 3D object is fabricated onto the movable stage. In some embodiments, after exposure of the projected light pattern, the MCU outputs a signal to the stepper motor, such that the printer frame and / or stage are lowered. The previously polymerized layer of the 3D object being fabricated is then further submerged within the photopolymerizable material and the next cross-sectional pattern is emitted onto the incident plane. This process is repeated until at least a first portion of the 3D object being fabricated is formed. In some embodiments, the stepper motor continuously lowers the movable stage into the build chamber as the digital micromirror device or LCD is continuously varied to emit the light pattern of the appropriate two-dimensional cross-section. In some embodiments, the stepper motor continuously lowers the movable stage into the fluid reservoir as the digital micromirror device or LCD is continuously varied to emit the light pattern of the appropriate two-dimensional cross-section. In such embodiments, continuous polymerization allows for the 3D object to be produced with a high tolerance and little to no separation between the two-dimensional cross-sections.

[0132] In some embodiments, the system combines multiple light projectors to create an incident plane having a larger surface area than possible with a single light projector, n some embodiments, four light projectors are used to produce the two-dimensional cross section patterns of the 3D object being fabricated, wherein each projector comprises a quadrant of the pattern. In some embodiments, a single light projector is used having multiple digital micromirror devices or liquid crystal displays to create an incident plane having a larger surface area than possible with a single DMD or LCD. In some embodiments, four DMDs or LCDs are used to produce the two-dimensional cross section patterns of the 3D object being fabricated, wherein each DMD or LCD comprises a quadrant of the pattern.

[0133] In stereolithography (SLA), the light source is in the form of a focused laser thatis rastered across a plane of the photoactive material coincident to the laser’s focal point. Using a computer to control the motion and activation of the laser, a predetermined two- dimensional pattern can be selectively polymerized from the photo-active solution onto the movable stage. By coordinating the incremental motion of the movable stage along an axis orthogonal to the plane of polymerization, an entire three-dimensional object can be fabricated through the sequential polymerization of multiple two dimensional layers.

[0134] In some embodiments, the light source is a laser light source and the DMD or LCD is replaced by one or more scanning mirrors or galvanometers used to reflect the laser across the incident plane to polymerize the photopolymerizable material provided in the reservoir 531.

[0135] In some cases, stereolithography apparatus (SLA) may be utilized as the light projector in the additive manufacturing system. In some embodiments, the system comprises a computing device. The computer assisted design model (CAD) may be loaded onto the computing device. The CAD model may be stored in a memory of the computing device which may include an internal data storage system or an external data storage system. In some embodiments, the computing device may include an installed 3D printing software. The 3D printing software may comprise a graphical user interface (GUI). In some embodiments, the 3D printing software analyzes the loaded 3D object to be fabricated and initializes cross-sections of the 3D object for a printing sequence. In some embodiments, the thickness and / or point at which each cross-section is formed is dependent upon a desired tolerance set by the user.

[0136] In some embodiments, a micro controller unit (MCU) is provided as an interface to external components of the computing device. In some embodiments, a user interacts with the computing device and manipulate the model via a user interface which connects to the computing device via the MCU. In some embodiments, the 3D printing software allows the user to choose the orientation of the object to be fabricated relative to the movable stage. In some embodiments, the MCU is connected to the stepper motor which actuates the printer to raise or lower the movable stage relative to the fluid reservoir, build chamber, or optically transparent window. In some embodiments, the stepper motor actuates the printer frame to raise or lower the movable stage relative to the fluid reservoir, build chamber, or optically transparent window.

[0137] In some embodiments, the MCU connects the computing device to the controller of the stereolithography apparatus (SLA). In some embodiments, the SLA controller sends positioning data and a clocking pulse to the one or more scanning mirrors (SM) or galvanometers to position the SM, manipulating light from the light source to raster the pattern of the two-dimensional cross-section for the 3D object being fabricated. In some embodiments, an analog SM control is provided, wherein the analog control converts the digital signal from the digital controller prior to sending the signal.

[0138] In some embodiments, the light projector further comprises a collimating optical system comprised of one or more optical lenses to further collimate light from thelight source onto the SM. In some embodiments, the light projector further comprises a collimating optical system comprised of one or more optical lenses to further collimate light from the light source from the SM. In some embodiments, the light projector further comprises a focal optical system comprised of one or more optical lenses to focus the laser beam emitted by the onto an incident plane.

[0139] In some embodiments, the incident plane is provided within in fluid reservoir such that the light pattern representing a two-dimensional cross-section of the object being fabricated is incident on unpolymerized material provided within the fluid reservoir. In some embodiments, the incident plane is provided within in fluid reservoir such that the light pattern representing a two-dimensional cross-section of the object being fabricated is incident on unpolymerized material provided within the build chamber. Upon exposure to the laser beam, the photopolymerizable material begins cross-linking and solidifying to form a layer of the 3D object being fabricated as the beam is rastered across the incident plane. In some embodiments, after completion of a cross-sectional layer, the MCU 686 outputs a signal to the stepper motor, such that the printer frame and / or the stage are lowered. The previously polymerized layer of the 3D object being fabricated is then further submerged within the photopolymerizable material and the next cross-sectional rastered onto the incident plane. This process is repeated until at least a first portion of the 3D object being fabricated is formed. In some embodiments, the movable stage is lowered incrementally after each cross- sectional layer is formed.

[0140] In some embodiments, the additive manufacturing system utilizes multiple spatial light modulators in conjunction with one or more light sources. This allows for the selective exposure of a much larger area on the focal plane and therefore allows much larger layer areas to be fabricated throughout a given build sequence. In some embodiments, the multiple spatial light modulators each form a section of the projected two -dimensional crosssections of the 3D object to be fabricated.

[0141] The light projector includes at least one light source configured to project patterned light towards the fluid reservoir. In some embodiments, the light projector includes at least one light source configured to project patterned light towards the build chamber. In some embodiments, the fluid reservoir or build chamber is configured to contain a volume of photopolymerizable material. Light may be emitted from the projector with sufficient energy and intensity to polymerize a liquid solution of photo -active monomers.

[0142] In some embodiments, the additive manufacturing system utilizes multiple optically transparent windows between the projection optics and the movable stage in thebuild chamber. In some embodiments, the multiple optically transparent windows allow for additional levels of containment of the contained fluid inside the build chamber. In some embodiments, a fluid is placed between each of the optically transparent windows. In some embodiments, the refractive index of the fluid is equal to the refractive index of the optically transparent windows. In some embodiments, the fluid comprises a first refractive index. In some embodiments, the optically transparent windows comprise a second refractive index. In some embodiments, the first refractive index is equal to the second refractive index. In some embodiments, the first refractive index is substantially similar to the second refractive index. In some embodiments, the first refractive index is greater than the second refractive index. In some embodiments, the second refractive index is greater than the first refractive index. In some embodiments, the fluid is an immersion oil. In some embodiments, the fluid allows for the patterned image projected onto the optical plane to retain its intended geometry and not be skewed by passing through the optically transparent windows. In some embodiments, the fluid is configured to ensure that the patterned image projected onto the optical plane retains its intended geometric configuration without distortion as it traverses the optically transparent windows.

[0143] In some embodiments, the light emitted from the projector is comprised of UV light, blue light, or light within the 450 to 495 nm range. In some embodiments, the light emitted from the projector comprises a wavelength of about lOOnm to about 500 nm, or any range of values therebetween. In some embodiments, the light emitted from the projector comprises a wavelength of about 100 nm to about 280 nm. In some embodiments, the light emitted from the projector comprises a wavelength of about 280 nm to about 315 nm. In some embodiments, the light emitted from the projector comprises a wavelength of about 315 nm to about 400 nm. In some embodiments, the light emitted from the projector comprises a wavelength of about 380 nm to about 500 nm. In some embodiments, the light emitted from the projector comprises a wavelength of about 400 nm to about 450 nm. In some embodiments, light emitted from the projector comprises a wavelength of about 450 nm to about 495 nm. In some embodiments, light emitted from the projector comprises a wavelength of about 300 nm to about 310 nm, about 3 lOnm to about 320 nm, about 320 nm to about 330 nm, about 330 nm to about 340 nm, about 340 nm to about 350 nm, about 350 nm to about 360 nm, about 360 nm to about 370 nm, about 370 nm to about 380 nm, about 380 nm to about 390 nm, about 390 nm to about 400 nm, about 400 nm to about 410 nm, about 410 nm to about 420 nm, about 420 nm to about 430 nm, about 430 nm to about 440 nm, about 440 nm to about 450 nm, about 450 nm to about 455 nm, about 450 nm to about460 nm, about 450 nm to about 465 nm, about 450 nm to about 470 nm, about 450 nm to about 475 nm, about 450 nmto about 480 nm, about 450 nm to about 485 nm, about 450 nm to about 490 nm, about 450 nm to about 495 nm, about 455 nm to about 460 nm, about 455 nm to about465 nm, about455 nm to about 470 nm, about 455 nm to about 475 nm, about 455 nm to about 480 nm, about 455 nm to about 485 nm, about 455 nm to about 490 nm, about455 nm to about 495 nm, about460 nm to about 465 nm, about 460 nm to about 470 nm, about 460 nm to about 475 nm, about 460 nm to about 480 nm, about 460 nm to about 485 nm, about 460 nm to about 490 nm, about 460 nm to about 495 nm, about 465 nm to about 470 nm, about 465 nmto about475 nm, about465 nm to about480 nm, about 465 nm to about 485 nm, about 465 nm to about 490 nm, about 465 nm to about 495 nm, about 470 nm to about475 nm, about470 nm to about 480 nm, about 470 nm to about 485 nm, about 470 nm to about 490 nm, about 470 nm to about 495 nm, about 475 nm to about 480 nm, about 475 nm to about 485 nm, about475 nm to about 490 nm, about 475 nm to about 495 nm, about 480 nm to about 485 nm, about 480 nm to about 490 nm, about 480 nm to about 495 nm, about 485 nm to about 490 nm, about 485 nm to about 495 nm, or about 490 nm to about 495 nm, including increments therein and any range of values therebetween. In some embodiments, light emitted from the projector comprises a wavelength of about 450 nm, about455 nm, about 460 nm, about 465 nm, about 470 nm, about 475 nm, about 480 nm, about 485 nm, about 490 nm, or about 495 nm. In some embodiments, light emitted from the projector comprises a wavelength of at least about 450 nm, about 455 nm, about 460 nm, about 465 nm, about 470 nm, about 475 nm, about 480 nm, about 485 nm, or about 490 nm. In some embodiments, light emitted from the projector comprises a wavelength of at most about455 nm, about 460 nm, about 465 nm, about 470 nm, about 475 nm, about 480 nm, about 485 nm, about 490 nm, or about 495 nm. In some embodiments, light emitted from the projector comprises a frequency of about 750 terahertz. In some embodiments, blue light is preferentially transmitted from the projection optics of the light source, through the one or more transparent walls, and into the photopolymerizable fluid, as compared to other wavelengths of light, or UV light. In some embodiments, the method comprises transmitting light in the within the 450 to 495 nm range to the fluid. In some embodiments, the method comprises increasing a transmission efficiency of the light by transmitting light in the within the 450 to 495 nm range through the system projection optics, the one or more transparent walls, and into the fluid.

[0144] In some embodiments, the photopolymerizable materials comprise biologically acceptable polymers. In some embodiments, the polymer can includepoly ethylene glycol based polymers such as, but not limited to polyethylene glycol diacrylate (PEGDA) and poly(ethyelene glycol) diacrylate. In some embodiments, the polymer can include gelatin methacryloyl hydrogels, gelatin methacrylate (GelMA) hydrogels, or a combination thereof. In some embodiments, the polymers can include combinations of polyethylene glycol diacrylate, poly(ethyelene glycol) diacrylate, gelatin methacrylate, and gelatin methacryloyl. In some embodiments, the polymers can include combinations of polyethylene glycol diacrylate and gelatin methacryloyl. In some embodiments, the polymers can include combinations of poly(ethyelene glycol) diacrylate and gelatin methacryloyl.

[0145] In some embodiments, the additive manufacturing system utilizes a top-down approach, wherein the light source is positioned above the fluid reservoir filled with polymerizable material. A plurality of projection optics is coupled to the light projector and configured to focus light from the projector onto a plane. The incident plane may be perpendicular to the direction at which light is emitted from the projector. In some embodiments, the movable stage is submersible within the fluid reservoir. In some embodiments, the movable stage is submersible within the build chamber. In some embodiments, the movable stage is positioned just below the incident plane which light from the projector is focused on. In some embodiments, the projector illuminates a thin layer of photo-active material sitting on top of the movable stage at the incident plane. In some embodiments, after polymerizing the layer, the movable stage descends into the build chamber and allows another layer of fluid to sit atop the first layer. In some embodiments, after polymerizing the layer, the movable stage descends into the build chamber containing photopolymer material and allows another layer of fluid to sit atop the first layer. The next layer of fluid is then exposed to light from the projector to form a successive layer of the 3D object being manufactured. This process repeats for all layers of the object, where each layer corresponds to the object’s desired cross section.

[0146] In some embodiments, the removable cartridge (140) comprises a heating element. In some embodiments, the removable cartridge (140) comprises a heating element and one or more circulating fans positioned in the first compartment. The one or more circulating fans can be useful in reaching and maintain a temperature within the removable cartridge and within the system in a microgravity environment, where heat does not dissipate or transfer as it ordinarily would in a gravitational field. For example, in a microgravity environment in a sealed compartment, the one or more circulating fans can provide for a uniform temperature within the system and among system components. In some embodiments, the heating element is engaged and the one or more circulating turned on toinduced convection heat transfer with the system. In some embodiments, the heating element warms the print chamber (1250) and fluid reservoirs (1220) to the desired temperature. In some embodiments, the desired temperature is between about 37°C to about 42°C. In some embodiments, the removable cartridge (140) comprises a thermometer, wherein the thermometer measures the temperature of the print chamber and fluid reservoirs. In some embodiments, the methods comprise heating the system, the first compartment, the second compartment, the print chamber, the fluid reservoirs, or any combination thereof, to at least 37°C and maintaining the temperature. In some embodiments, the methods comprise heating the system, the first compartment, the second compartment, the print chamber, the fluid reservoirs, or any combination thereof, to a temperature not exceeding 45°C. In some embodiments, the additive manufacturing system maintains the print chamber and fluid reservoirs at the desired temperature via the heating element contained in the removable cartridge (140).

[0147] In some embodiments, the removable cartridge (140) further comprises a thermometer. In some embodiments, the removable cartridge (140) further comprises a thermometer in communication with the heating element. In some embodiments, the removable cartridge (140) further comprises a thermometer in communication with the heating element and one or more circulating fans positioned in the second compartment. In some embodiments, the thermometer is configured to monitor a temperature within the removable cartridge (140). In some embodiments, the additive manufacturing system is configured to maintain the temperature within the removable cartridge (140) below a critical level. In some embodiments, the critical level is about 45°C. In some embodiments, the critical level is above 45°C. In some embodiments, the thermometer is configured to monitor a temperature within the print chamber (1250). In some embodiments, the additive manufacturing system is configured to maintain the temperature within the print chamber (1250) below a critical level. In some embodiments, the additive manufacturing system is configured to cease operation of the system if a critical temperature is reached. In some embodiments, the additive manufacturing system is configured to cease operation of the printing process if a critical temperature is reached.

[0148] In some embodiments, the additive manufacturing system comprises one or more imaging devices (980). In some embodiments, the housing (110) comprises one or more imaging devices (980). In some embodiments, the one or more imaging devices (980) are configured to image an object manufactured within the system. In some embodiments, the one or more imaging devices (980) are configured to image the first compartment (111), orthe second compartment. In some embodiments, the one or more imaging devices (980) are configured to image one or more components of the system. In some embodiments, the one or more imaging devices (980) are configured to image the internals of the system before, during, and after a fabrication sequence. In some embodiments, the one or more imaging devices (980) are configured to image the removable cartridge (140) during a fabrication sequence. In some embodiments, the one or more imaging devices (980) are oriented to be used to image a completed 3D object after the fabrication steps. In some embodiments, the one or more imaging devices (980) are configured to monitor the 3D object before, during, and after a fabrication sequence. In some embodiments, the one or more imaging devices may an optical camera, radar device, sonar device, or the like. In some embodiments, the one or more imaging devices comprises an optical camera.

[0149] In some embodiments, the monitoring the 3D object comprises monitoring a dimensionality of the solid for compliance with the 3D model. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model is defined by a maximum optical resolution of the camera. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model is within 0.6 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.2 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.3 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.4 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.5 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.6 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 1 micron.

[0150] In some embodiments, the additive manufacturing system comprises an avionics spine (150). In some embodiments, the avionics spine (150) comprises a plurality of electrical and data connections electrically coupling the additive manufacturing system to a spacecraft. In some embodiments, the additive manufacturing system comprises an avionics spine (150) that passes all electrical power and data between the host spacecraft or module and the additive manufacturing internal subsystem (120).

[0151] In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allow the system (130) to be powered by the host spacecraft and controlled remotely.In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allow the system (130) to be powered by the host spacecraft and controlled manually.

[0152] In some embodiments, the system and methods are provided to manufacture biomimetic structures or medical implants to be grafted, transplanted, or otherwise inserted into a living structure or a patient in need thereof. In some embodiments, the biomimetic structure may be a biomimetic scaffold for promoting neural tissue growth and proliferation in a subject. In some embodiments, the biomimetic structure may be a biomimetic blood vessel forreplacement or repair of a damaged blood vessel or blood lumen. The subject may be an animal with a complex nerve system, such as a mammal, like a human, primate, or companion animal. The tissue scaffolds according to the present disclosure may thus be devices implanted in such a subject.

[0153] The printed medical implants may assist in restoring bodily function. The function can be restored in a mammal that can include a human, a horse, a pig, a cow, a bull, a goat, a sheep, a dolphin, a dog, a cat, a camel, or the like. In one embodiment, the mammal is a human. These medical implants can be used in some embodiments to promote axonal regeneration after spinal cord or peripheral nerve injury. In some embodiments, the three - dimension printed medical implants comprise stem cells. These implants can be custom designed to fit a particular patient's anatomy.

[0154] In some embodiments, a scaffold of the desired length is 3D-printed and then placed in the injury site. In some embodiments, a neural repair scaffold comprising an inner microchannel architecture of the desired length is 3D-printed and then placed in the injury site. The proximal and distal nerve stumps are then inserted into the overhangs of the scaffold where they are aligned to the microchannel scaffold. The nerve epineurium is then sutured to the overhang or sheath, thus fixing the scaffold in place. Regenerating axons from the proximal side enter the scaffold and are guided through the injury site to the distal nerve stump. In some embodiments, the channels of the scaffold are filled with Schwann cells or stem cells via a perfusion apparatus to further support axonal regeneration. In some embodiments, the 3D printed scaffold comprises microparticles, nanoparticles, drug loaded microparticles, drug loaded nanoparticles, conductive materials, or any combination thereof.

[0155] In some embodiments, the additive manufacturing system is configured to fabricate biomimetic scaffolds comprising a plurality of microchannels respectively defining a longitudinal major axis. In accordance with certain variations of the present disclosure, a "microchannel" preferably has at least one spatial dimension (e.g., a diameter) that is lessthan about 1,000 pm. In certain aspects, each microchannel has an inner diameter of greater than or equal to about 10 pm to less than or equal to about 1 ,000 pm, optionally greater than or equal to about 10 pm to less than or equal to about 500 pm, optionally greater than or equal to about 50 pm to less than or equal to about 450 pm, optionally greater than or equal to about 50 pm to less than or equal to about 300 pm.

[0156] In certain aspects, the microchannels may be treated with a biofunctional agent or active ingredient; have different surface properties or surface roughness; or have surfaces with different moieties exposed, which can be useful in designing spatially guided cellular growth and in certain aspects to facilitate adhesion of cells or tissue or to promote release of biofunctional agents, which include biofunctional materials and active ingredients (e.g., pharmaceutical active ingredients), and the like, into the surrounding environment.

[0157] The biodegradable material forming the microchannel may dissolve, referring to physical disintegration, erosion, disruption and / or dissolution of a material and may include the resorption of such material by a living organism. In certain variations, biodegradable polymeric material may dissolve or erode upon exposure to a solvent comprising a high concentration of water, such as blood, serum, growth or culture media, bodily fluids, saliva, and the like. Thus, upon implantation, the material may dissolve or disintegrate into small pieces. For structural scaffold members, the dissolution rate (e.g., a rate at which the structural member is resorbed by surrounding cells) can be designed so that sufficient cellular growth occurs prior to the structure dissolving or disintegrating via the resorption process. In various embodiments, the tissue scaffold device is designed to have a degradation time or dissolution rate that coincides with an amount of time that permits adequate neural tissue regrowth through the scaffold to a target tissue in the subject. Depending upon the subject and the time needed for recuperation and regeneration of the tissue, by way of non-limiting example, the degradation time may be greater than or equal to about 1 month to less than or equal to about 3 years, greater than or equal to about 1 month to less than or equal to 1 year, and in certain variations, greater than or equal to about 1 month to less than or equal to 6 months. In this manner, the cellular scaffold structure supports and promotes cell growth, cell proliferation, cell differentiation, cell repair, and / or cell regeneration in three-dimensions, especially for neural tissue growth.

[0158] Using focused light for polymerization generates printing resolution of 1 pm, a 50- fold improvement over nozzle-based inkjet printers. In inkjet or extrusion -based approaches, mechanical integrity may be compromised by artificial interfaces between the drops or lines and can cause mechanical failure during or after in vivo application. Byproviding layer less resolution in the Z direction, the structures may not exhibit these planar artifacts (interfaces) induced by a movement of a linear stage to a new position. Thus, systems and methods described herein can improve mechanical integrity of 3D printed implants and offer rapid fabrication of complex 3D biomimetic structures at microscale resolution.

[0159] In some embodiments, a printed biomimetic implant is printed in a single portion for implantation at a site of spinal cord transection or a peripheral nerve injury. In some embodiments, the implant is printed in two or more portions whereby a damaged, but not transected, spinal cord, or peripheral nerve, can be treated with an implant disclosed herein. If the spinal cord injury (SCI), or peripheral nerve injury, is not a transection, the one or more portions of the implant can be implanted surrounding the surviving tissue and the portions adhered to each other using a biologically acceptable adhesive. Thus, any surviving tissue can be maintained and regeneration of the host spinal cord, or peripheral nerve, encouraged at the injury site.

[0160] An implant may be customized for each patient. Upon imaging of a patient spinal cord, or peripheral nerve, injury a 3D model is created using CAD software. This model is then used to print a patient specific implant that fit and fill the lesion. In some embodiments, post-processing systems and methods to treat a fabricated biomimetic implant can increase the likelihood that the implant will be accepted by the body of which it is implanted into.

[0161] There is more to be gained by incorporating multiple materials in a given build sequence. Different materials exhibit different physical, chemical, or biological properties that may be suitable for certain parts of an object (but not others), and these additional materials may contain components (such as cells or particles) that are desired in preselected areas of that object. The combination of multiple materials in a single build sequence may allow for heterogeneous constructs to be fabricated that may serve more utility than its homogenous counterpart. In some embodiment, different reservoirs can be part of the same fabrication apparatus, where the movable stage or build stage can alternate from one reservoir to another as is appropriate to create that section of the object. In some embodiments, the different materials may be contained in separate containers connected to the same reservoir. In such examples, the material options can be pumped in or pumped out as directed by the controlling software.

[0162] In some embodiments, the system for the microfabrication of objects includes the continuous motion of the movable stage through the build chamber. Such a configurationallows objects to be created without layers, decreasing fabrication times, and provides isotropic mechanical properties and smooth features along the structure of the newly fabricated object.

[0163] In some embodiments, the system for the microfabrication of objects allows for the light source to be turned on and off throughout a given print sequence (“blinking”). When coordinated with the incremental motion of the movable stage through the build chamber, this allows layers of precise thicknesses to be created without the threat of over- curing the photopolymer solution. The advantages become especially apparent when the light source used is a laser with very fast blinking or pulsing capabilities. When combined with very precise incremental motion of the movable stage, such embodiments allow very fine structures to be created that most closely approximate the intended structures designed and displayed by the spatial light modulators.

[0164] In some embodiments, the system for the microfabrication of objects is configured to create a prevascularized three-dimensional tissue or organ structure. In such an embodiment, the outer contour of the structure can be biomimetic to the target structure’s natural appearance while comprising an internal architecture comprising perfusable lumens that mimic a native vascular network. In some embodiments, the perfusable vascular network can have its fluid inlet and outlet aligned with the through holes on the system’s movable stage that allow for the active perfusion of fluid through the printed construct. In some embodiments, the fluid perfused through the vascular network comprises a wash solution configured to remove unpolymerized photopolymer. In some embodiments, the fluid perfused through the vascular network comprises synthetic blood, cell culture media, or similar fluids intended for cell proliferation and maturation. In some embodiments, a suspension of cells and fluid can be perfused through the vascular network to line the internal lumens with a cell type of choice. In some embodiments, the cell type comprises endothelial cells. In some embodiments, the printed structure comprises a biocompatible photopolymer or mixture of biocompatible photopolymers comprising a suspension of parenchymal cells. The combination of printing parenchymal cells in the main printed structure with active perfusion of cell culture media allows for the fabrication of functional thick tissues.

[0165] In some embodiments, the system for the microfabrication of objects is configured to create a vascular network. In some embodiments the vascular network comprises a perfusable vascular branches, wherein the perfusable vascular branches can sustain active perfusion of fluid passed through them. In some embodiments the vascular network comprises vascular branches, wherein the vascular branches are in proximity of one-another such that the orientation of the vascular branches allows for nutrients and waste to be diffused between the walls of the vascular branches and the surroundings of the vascular branches. In some embodiments, the vascular network can be cast with a suspension of cells and biomaterial of choice, allowing for culture of three-dimensional thick tissues. In some embodiments, the suspension of cells and biomaterial(s) of choice comprises a third component forming a resultant solution. In some embodiments, the resultant solution comprises a third refractive index. In some embodiments, the suspension of cells and biomaterial(s) of choice comprises a fourth refractive index. In some embodiments, the third refractive index is equal to the fourth refractive index. In some embodiments, the third component causes the third refractive index. In some embodiments, the third component reduces light scattering and enables high-resolution fabrication sequences.

[0166] In some embodiments, the system for the microfabrication of objects is configured to create a Printed Circuit Board (PCB). In some embodiments, some fluids contained in the plurality of fluid reservoirs comprise of electrically conductive photopolymers, electrically insulative photopolymers, photopolymers containing electrically conductive materials, photopolymers containing electrically insulative materials, wash solutions, or any combination thereof. In some embodiments, the system creates electrically conductive wires on portions of any given layer and creates electrically insulative regions on other portions of any given layer. In some embodiments, the combination of patterning electrically conductive regions and electrically insulative regions on any given layer of a multi-layered construct allows for an on-demand circuitry. In some embodiments, the on- demand circuitry is designed and configured to act as a sensor. In some embodiments, the on- demand circuitry is designed and configured to act as an electrical -stimulation device. In some embodiments, the on-demand circuitry is designed and configured to perform logic operations. In some embodiments, the on-demand circuitry is designed and configured to interface or integrate with other digital or analog devices.

[0167] In some embodiments, the system comprises a software interface. In some embodiments, a user establishes connection with the machine via the software interface. In some embodiments, the user prompts the software to initiate a heating sequence of the removable cartridge. In some embodiments, the system comprises software to receive the design to be printed. In some embodiments, at any time, the user can load a design file into the software for an object that is intended to be fabricated. In some embodiments, the software receives the design file, slices the file into cross sections, and generates instructions that control the motor of the linear axis. In some embodiments, the design file is a CAD file.In some embodiments, the user may elect to upload their own instructions to control the motors.

[0168] In some embodiments, the system comprises a computing device. The computer assisted design model (CAD) may be loaded onto the computing device. The CAD model may be stored in a memory of the computing device which may include an internal data storage system or an external data storage system. In some embodiments, the computing device may include an installed 3D printing software. The 3D printing software may comprise a graphical user interface (GUI). In some embodiments, the 3D printing software analyzes the loaded 3D object to be fabricated and initializes cross-sections of the 3D object for a printing sequence. In some embodiments, the thickness and / or point at which each cross-section is formed is dependent upon a desired tolerance set by the user.

[0169] In some embodiments, a programable micro controller unit (MCU) is provided as an interface to external components of the computing device. In some embodiments, a user interacts with the computing device and manipulate the model via a user interface which connects to the computing device via the MCU. In some embodiments, the 3D printing software allows the user to choose the orientation of the object to be fabricated relative to the movable stage. The MCU, may also connect to the stepper motor which actuates the printer frame to raise or lower the movable stage relative to the fluid reservoir.

[0170] In some embodiments, the additive manufacturing system comprises one or more computer processors comprising a computer readable medium configured to implement a method of additively manufacturing a 3D object as disclosed herein. In some embodiments, the method of additive manufacturing further comprises one or more computer processors comprising a computer readable medium configured to implement a method comprising: a) sealing one or more of the first compartment, the second compartment, or the third compartment from one another; b) filling the print chamber with a photopolymerizable fluid comprised by the plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from the light source through the one or more transparent walls onto the movable stage and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid.

[0171] In some embodiments, the additive manufacturing system further comprises one or more computer processors comprising a computer readable medium configured to implement a method of additively manufacturing a 3D object comprising: a) sealing one ormore of the first compartment, the second compartment, or the third compartment from one another; b) filling the print chamber with a photopolymerizable fluid comprised by the plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from the light source through the one or more transparent walls onto the movable stage and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid, wherein the method takes place in a reduced gravity environment. In some embodiments, the computer readable medium further comprises instructions for producing a scaffold, a biological tissue, or a microfluidic device. In some embodiments, the computer readable medium further comprises instructions for producing electronic components. In some embodiments, the computer readable medium further comprises instructions for producing mechanical components. In some embodiments, the computer readable medium further comprises instructions for producing aerospace, medical device, dentistry, mechanical engineering, automotive, tools, manufacturing, robotic components, or a combination thereof.

[0172] In some embodiments, the computer readable medium further comprises instructions for producing any 3D object. In some embodiments, the instructions comprise a 3D model of the 3D object in discrete layers which correspond to a printing resolution of the system about a z axis defined by the movement of the movable stage.

[0173] In some embodiments, the housing is configured to interface with a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform. In some embodiments, the method further comprises monitoring the 3D object with one or more imaging devices. In some embodiments, the method further comprises monitoring the 3D object during any of c.-e. with one or more imaging devices. In some embodiments, the monitoring occurs before, during, and after a fabrication sequence. In some embodiments, the method further comprises monitoring a component of the system, the first compartment, or the second compartment with one or more imaging devices. In some embodiments, the method further comprises monitoring a component of the system, the first compartment, or the second compartment during any of c.-e. with one or more imaging devices. In some embodiments, the monitoring occurs before, during, and after a fabrication sequence. In some embodiments, the monitoring 3D object comprises monitoring a dimensionality of the solid for compliance with the 3D model. In some embodiments, the monitoring 3D object comprises monitoring a dimensionality of the solid for compliance with the 3D model withina maximum resolution of the imaging devices. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model is within 0.6 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.2 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.3 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.4 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.5 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 0.6 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model within 1 microns. In some embodiments, the monitoring the dimensionality of the solid for compliance with the 3D model is with the highest degree of precision enabled by the camera.

[0174] In some embodiments, the method further comprises moving the removeable cartridge relative to the movable stage and exposing multiple optical field -of -views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the method further comprises moving the light source relative to the movable stage and exposing multiple optical field -of-views onto the moveable stage, thereby allowing a larger print area than a single field of view.

[0175] In some embodiments, the plurality of fluid reservoirs comprise a chemical quencher that quenches free-radicals and prevents polymerization, or over polymerization. In some embodiments, the plurality of fluid reservoirs comprises a cell media fluid reservoir. In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising perfusing the cell media through the newly printed construct.

[0176] In some embodiments, the computer readable medium further comprises instructions for producing the 3D object, the 3D object comprising a plurality of channels. In some embodiments, the plurality of fluid reservoirs comprise a wash fluid and a photopolymerizable fluid, where the removable cartridge comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage, wherein the channels align with the inlet, wherein the method further comprise flushing the plurality of channelswith the wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels.

[0177] In some embodiments, the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising flushing the print chamber with the wash fluid to remove the photopolymerizable fluid, and directing the wash fluid and the removed photopolymerizable fluid into one of the fluid reservoirs initially holding the photopolymerizable fluid.

[0178] In some embodiments, method further comprises contacting the top layer of the photopolymerizable fluid with the one or more transparent walls. In some embodiments, the top layer comprises a thickness of up to 1 -200 micrometers. In some embodiments, the method further comprises filling the print chamber with the chemical quencher and preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber. In some embodiments, the target polymerization depth is 1-200 micrometers. In some embodiments, the target polymerization depth is between 10-50 micrometers. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is up to 1-5% v / v. In some embodiments, a concentration of the physical quencher in the print chamber is about 4% v / v. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is about 0.1 % w / v to about 5 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is about 0.1 % w / v to about 0.5 % w / v, about 0.1 % w / v to about 2 % w / v, about 0.1 % w / v to about 2.5 % w / v, about 0.1 % w / v to about 3 % w / v, about 0.1 % w / v to about 3.5 % w / v, about 0.1 % w / v to about 4 % w / v, about 0.1 % w / v to about 4.5 % w / v, about 0.1 % w / v to about 5 % w / v, about 0.5 % w / v to about 2 % w / v, about 0.5 % w / v to about 2.5 % w / v, about 0.5 % w / v to about 3 % w / v, about 0.5 % w / v to about3.5 % w / v, about 0.5 % w / v to about 4 % w / v, about 0.5 % w / v to about 4.5 % w / v, about 0.5 % w / v to about 5 % w / v, about 2 % w / v to about 2.5 % w / v, about 2 % w / v to about 3 % w / v, about 2 % w / v to about 3.5 % w / v, about 2 % w / v to about 4 % w / v, about 2 % w / v to about4.5 % w / v, about 2 % w / v to about 5 % w / v, about 2.5 % w / v to about 3 % w / v, about 2.5 % w / v to about 3.5 % w / v, about 2.5 % w / v to about 4 % w / v, about 2.5 % w / v to about 4.5 % w / v, about 2.5 % w / v to about 5 % w / v, about 3 % w / v to about 3.5 % w / v, about 3 % w / v toabout 4 % w / v, about 3 % w / v to about 4.5 % w / v, about 3 % w / v to about 5 % w / v, about 3.5 % w / v to about 4 % w / v, about 3.5 % w / v to about 4.5 % w / v, about 3.5 % w / v to about 5 % w / v, about 4 % w / v to about 4.5 % w / v, about 4 % w / v to about 5 % w / v, or about 4.5 % w / v to about 5 % w / v, including increments therein. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is about 0.1 % w / v, about 0.5 % w / v, about 2 % w / v, about 2.5 % w / v, about 3 % w / v, about 3.5 % w / v, about 4 % w / v, about 4.5 % w / v, or about 5 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is at least about 0.1 % w / v, about 0.5 % w / v, about 2 % w / v, about 2.5 % w / v, about 3 % w / v, about 3.5 % w / v, about 4 % w / v, or about4.5 % w / v. In some embodiments, a concentration of the physical quencher in the print chamber utilized to modify the depth of top layer which is being polymerized is at most about 0.5 % w / v, about2 % w / v, about 2.5 % w / v, about 3 % w / v, about 3.5 % w / v, about 4 % w / v, about 4.5 % w / v, or about 5 % w / v. In some embodiments, the target depth of the layer of fluid comprising the chemical quencher being polymerized is about 1 micrometers to about 500 micrometers. In some embodiments, the target depth of the layer of fluid comprising the chemical quencher being polymerized is about 1 micrometers to about 2 micrometers, about 1 micrometers to about 3 micrometers, about 1 micrometers to about 4 micrometers, about 1 micrometers to about 5 micrometers, about 1 micrometers to about 10 micrometers, about 1 micrometers to about 25 micrometers, about 1 micrometers to about 50 micrometers, about 1 micrometers to about 100 micrometers, about 1 micrometers to about 150 micrometers, about 1 micrometers to about 300 micrometers, about 1 micrometers to about 500 micrometers, about 2 micrometers to about 3 micrometers, about 2 micrometers to about 4 micrometers, about 2 micrometers to about 5 micrometers, about 2 micrometers to about 10 micrometers, about 2 micrometers to about 25 micrometers, about 2 micrometers to about 50 micrometers, about 2 micrometers to about 100 micrometers, about 2 micrometers to about 150 micrometers, about 2 micrometers to about 300 micrometers, about 2 micrometers to about 500 micrometers, about 3 micrometers to about 4 micrometers, about 3 micrometers to about 5 micrometers, about 3 micrometers to about 10 micrometers, about 3 micrometers to about 25 micrometers, about 3 micrometers to about 50 micrometers, about 3 micrometers to about 100 micrometers, about 3 micrometers to about 150 micrometers, about 3 micrometers to about 300 micrometers, about 3 micrometers to about 500 micrometers, about 4 micrometers to about 5 micrometers, about 4 micrometers to about 10 micrometers, about 4 micrometers to about 25 micrometers, about 4 micrometers to about 50 micrometers,about 4 micrometers to about 100 micrometers, about 4 micrometers to about 150 micrometers, about 4 micrometers to about 300 micrometers, about 4 micrometers to about 500 micrometers, about 5 micrometers to about 10 micrometers, about 5 micrometers to about 25 micrometers, about 5 micrometers to about 50 micrometers, about 5 micrometers to about 100 micrometers, about 5 micrometers to about 150 micrometers, about 5 micrometers to about 300 micrometers, about 5 micrometers to about 500 micrometers, about 10 micrometers to about 25 micrometers, about 10 micrometers to about 50 micrometers, about 10 micrometers to about 100 micrometers, about 10 micrometers to about 150 micrometers, about 10 micrometers to about 300 micrometers, about 10 micrometers to about 500 micrometers, about 25 micrometers to about 50 micrometers, about 25 micrometers to about 100 micrometers, about 25 micrometers to about 150 micrometers, about 25 micrometers to about 300 micrometers, about 25 micrometers to about 500 micrometers, about 50 micrometers to about 100 micrometers, about 50 micrometers to about 150 micrometers, about 50 micrometers to about 300 micrometers, about 50 micrometers to about 500 micrometers, about 100 micrometers to about 150 micrometers, about 100 micrometers to about 300 micrometers, about 100 micrometers to about 500 micrometers, about 150 micrometers to about 300 micrometers, about 150 micrometers to about 500 micrometers, or about 300 micrometers to about 500 micrometers, including increments therein. In some embodiments, the target depth of the layer of fluid comprising the chemical quencher being polymerized is about 1 micrometer, about 2 micrometers, about 3 micrometers, about 4 micrometers, about 5 micrometers, about 10 micrometers, about 25 micrometers, about 50 micrometers, about 100 micrometers, about 150 micrometers, about 300 micrometers, or about 500 micrometers. In some embodiments, the target depth of the layer of fluid comprising the chemical quencher being polymerized is at least about 1 micrometer, about 2 micrometers, about 3 micrometers, about 4 micrometers, about 5 micrometers, about 10 micrometers, about 25 micrometers, about 50 micrometers, about 100 micrometers, about 150 micrometers, or about 300 micrometers. In some embodiments, the target depth of the layer of fluid comprising the chemical quencher being polymerized is at most about 2 micrometers, about 3 micrometers, about 4 micrometers, about 5 micrometers, about 10 micrometers, about 25 micrometers, about 50 micrometers, about 100 micrometers, about 150 micrometers, about 300 micrometers, or about 500 micrometers.

[0179] In some embodiments, the method comprises moving the movable stage comprises moving the movable stage away from the optical plane such that the projected two- dimensional cross-sections of the three-dimensional object are incident onphotopolymerizable fluid in contact with and adjacent to previously formed layers of the object. In some embodiments, the movingthe movable stage comprises moving the movable stage continuously. In some embodiments, movingthe movable stage comprises moving the movable stage incrementally. In some embodiments, the moving the movable stage comprises moving the movable stage between about 0.1 mm / min to about 0.5 mm / min.

[0180] In some embodiments, the method further comprises directing the light. In some embodiments, the method further comprises directing two-dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage. In some embodiments, the method further comprises movingthe one or more light sources relative to the movable stage and exposing multiple optical field -of-views onto the moveable stage, thereby allowing a larger print area than a single field of view. In some embodiments, the directing light comprises intermittently activating the one or more light sources during the formation of the three-dimensional object. In some embodiments, the method further comprising mounting the system in an orientation without regard to a gravitational field. In some embodiments, the method, further comprises operating the system in an orientation without regard to a gravitational field.

[0181] In some embodiments, the one or more transparent walls are gas permeable, the method further comprising providing an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring against the one or more transparent walls.

[0182] In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with the thermometer and maintaining the temperature below a critical level. In some embodiments, the method further comprises monitoring a temperature within the removable cartridge with the thermometer, and ceasing operation of the system if a critical temperature is reached. In some embodiments, the critical temperature is above 45°C.

[0183] In some embodiments, any of the above methods create a 3D printed construct.

[0184] In some embodiments, the therapeutic agent is released into the lumens of the newly printed construct. The present disclosure provides computer systems that are programmed to implement methods of the disclosure. The computer system can be an electronic device of a user or a computer system that is remotely located with respect to the electronic device. The electronic device can be a mobile electronic device. The computer system includes a central processing unit (CPU, also “processor” and “computer processor”herein), which can be a single core or multi core processor, or a plurality of processors for parallel processing. The computer system also includes memory or memory location (e.g., random -access memory, read-only memory, flash memory), electronic storage unit (e.g., hard disk), communication interface (e.g., network adapter) for communicating with one or more other systems, and peripheral devices, such as cache, other memory, data storage and / or electronic display adapters. The memory, storage unit, interface and peripheral devices are in communication with the CPU through a communication bus (solid lines), such as a motherboard. The storage unit can be a data storage unit (or data repository) for storing data. The computer system can be operatively coupled to a computer network (“network”) with the aid of the communication interface. The network can be the Internet, an internet and / or extranet, or an intranet and / or extranet that is in communication with the Internet. The network in some cases is a telecommunication and / or data network. The network can include one or more computer servers, which can enable distributed computing, such as cloud computing. The network, in some cases with the aid of the computer system, can implement a peer-to-peer network, which may enable devices coupled to the computer system to behave as a client or a server.

[0185] The CPU can execute a sequence of machine-readable instructions, which can be embodied in a program or software. The instructions may be stored in a memory location, such as the memory. The instructions can be directed to the CPU, which can subsequently program or otherwise configure the CPU to implement methods of the present disclosure. Examples of operations performed by the CPU can include fetch, decode, execute, and writeback.

[0186] The CPU can be part of a circuit, such as an integrated circuit. One or more other components of the system can be included in the circuit. In some cases, the circuit is an application specific integrated circuit (ASIC).

[0187] The storage unit can store files, such as drivers, libraries and saved programs. The storage unit can store user data, e.g., user preferences and user programs. The computer system in some cases can include one or more additional data storage units that are external to the computer system, such as located on a remote server that is in communication with the computer system through an intranet or the Internet.

[0188] The computer system can communicate with one or more remote computer systems through the network. For instance, the computer system can communicate with a remote computer system of a user (e.g., hand-held device). Examples of remote computer systems include personal computers (e.g., portable PC), slate or tablet PC’s (e.g., Apple®iPad, Samsung® Galaxy Tab), telephones, Smart phones (e.g., Apple® iPhone, Android- enabled device, Blackberry®), or personal digital assistants. The user can access the computer system via the network.

[0189] Methods as described herein can be implemented by way of machine (e.g., computer processor) executable code stored on an electronic storage location of the computer system, such as, for example, on the memory or electronic storage unit. The machine executable or machine readable code can be provided in the form of software. During use, the code can be executed by the processor. In some cases, the code can be retrieved from the storage unit and stored on the memory for ready access by the processor. In some situations, the electronic storage unit can be precluded, and machine-executable instructions are stored on memory.

[0190] The code can be pre-compiled and configured for use with a machine having a processer adapted to execute the code, or can be compiled during runtime. The code can be supplied in a programming language that can be selected to enable the code to execute in a pre-compiled or as-compiled fashion.

[0191] Aspects of the systems and methods provided herein, such as the computer system, can be embodied in programming. Various aspects of the technology may be thought of as “products” or “articles of manufacture” typically in the form of machine (or processor) executable code and / or associated data that is carried on or embodied in a type of machine readable medium. Machine -executable code can be stored on an electronic storage unit, such as memory (e.g., read-only memory, random-access memory, flash memory) or a hard disk. “Storage” type media can include any or all of the tangible memory of the computers, processors or the like, or associated modules thereof, such as various semiconductor memories, tape drives, disk drives and the like, which may provide non -transitory storage at any time for the software programming. All or portions of the software may at times be communicated through the Internet or various other telecommunication networks. Such communications, for example, may enable loading of the software from one computer or processor into another, for example, from a management server or host computer into the computer platform of an application server. Thus, another type of media that may bear the software elements includes optical, electrical and electromagnetic waves, such as used across physical interfaces between local devices, through wired and optical landline networks and over various air-links. The physical elements that carry such waves, such as wired or wireless links, optical links or the like, also may be considered as media bearing the software. As used herein, unless restricted to non-transitory, tangible “storage” media, termssuch as computer or machine “readable medium” refer to any medium that participates in providing instructions to a processor for execution.

[0192] Hence, a machine readable medium, such as computer-executable code, may take many forms, including but not limited to, a tangible storage medium, a carrier wave medium or physical transmission medium. Non-volatile storage media include, for example, optical or magnetic disks, such as any of the storage devices in any computer(s) or the like, such as may be used to implement the databases, etc. shown in the drawings. Volatile storage media include dynamic memory, such as main memory of such a computer platform. Tangible transmission media include coaxial cables; copper wire and fiber optics, including the wires that comprise a bus within a computer system. Carrier-wave transmission media may take the form of electric or electromagnetic signals, or acoustic or light waves such as those generated during radio frequency (RF) and infrared (IR) data communications. Common forms of computer-readable media therefore include for example: a floppy disk, a flexible disk, hard disk, magnetic tape, any other magnetic medium, a CD- ROM, DVD or DVD-ROM, any other optical medium, punch cards paper tape, any other physical storage medium with patterns of holes, a RAM, a ROM, a PROM and EPROM, a FLASH-EPROM, any other memory chip or cartridge, a carrier wave transporting data or instructions, cables or links transporting such a carrier wave, or any other medium from which a computer may read programming code and / or data. Many of these forms of computer readable media may be involved in carrying one or more sequences of one or more instructions to a processor for execution.

[0193] The computer system can include or be in communication with an electronic display that comprises a user interface (UI) for providing, for example, concentration of the analyte of interest. Examples of UFs include, without limitation, a graphical user interface (GUI) and web-based user interface.

[0194] Methods and systems of the present disclosure can be implemented by way of one or more algorithms. An algorithm can be implemented by way of software upon execution by the central processing unit.

[0195] In some aspects, the disclosed provides quality control methods or methods to assess a risk associated with a food, with a hospital, with a clinic, or any other location where the presence of a bacterium poses a certain risk to one or more subjects. In many instances, systems, platforms, software, networks, and methods described herein include a digital processing device, or use of the same. In further embodiments, the digital processing device includes one or more hardware central processing units (CPUs), i.e., processors that carry outthe device’s functions, such as the automated sequencing apparatus disclosed herein or a computer system used in the analyses of a plurality of nucleic acid sequencing reads from samples derived from a food processing facility or from any other facility, such as a hospital a clinical or another. In still further embodiments, the digital processing device further comprises an operating system configured to perform executable instructions. In some embodiments, the digital processing device is optionally connected a computer network. In further embodiments, the digital processing device is optionally connected to the Internet such that it accesses the World Wide Web. In still further embodiments, the digital processing device is optionally connected to a cloud computing infrastructure. In other embodiments, the digital processing device is optionally connected to an intranet. In other embodiments, the digital processing device is optionally connected to a data storage device. In other embodiments, the digital processing device could be deployed on premise or remotely deployed in the cloud.

[0196] In accordance with the description herein, suitable digital processing devices include, by way of non-limiting examples, server computers, desktop computers, laptop computers, notebook computers, sub -notebook computers, netbook computers, netpad computers, set-top computers, handheld computers, Internet appliances, mobile smartphones, tablet computers, personal digital assistants, video game consoles, and vehicles. Those of skill in the art will recognize that many smartphones are suitable for use in the system described herein. Those of skill in the art will also recognize that select televisions, video players, and digital music players with optional computer network connectivity are suitable for use in the system described herein. Suitable tablet computers include those with booklet, slate, and convertible configurations, known to those of skill in the art. In many aspects, the disclosure contemplates any suitable digital processing device that can either be deployed to a food processing facility, or is used within said food processing facility to process and analyze a variety of nucleic acids from a variety of samples.

[0197] In some embodiments, a digital processing device includes an operating system configured to perform executable instructions. The operating system is, for example, software, including programs and data, which manages the device’s hardware and provides services for execution of applications. Those of skill in the art will recognize that suitable server operating systems include, by way of non -limiting examples, FreeBSD, OpenBSD, NetBSD®, Linux, Apple® Mac OS X Server®, Oracle® Solaris®, Windows Server®, and Novell® NetWare®. Those of skill in the art will recognize that suitable personal computer operating systems include, by way of non -limiting examples, Microsoft® Windows®,Apple® Mac OS X®, UNIX®, and UNIX-like operating systems such as GNU / Linux®. In some embodiments, the operating system is provided by cloud computing. Those of skill in the art will also recognize that suitable mobile smart phone operating systems include, by way of non-limiting examples, Nokia® Symbian® OS, Apple® iOS®, Research In Motion® BlackBerry OS®, Google® Android®, Microsoft® Windows Phone® OS, Microsoft® Windows Mobile® OS, Linux®, and Palm® WebOS®.

[0198] In some embodiments, a digital processing device includes a storage and / or memory device. The storage and / or memory device is one or more physical apparatuses used to store data or programs on a temporary or permanent basis. In some embodiments, the device is volatile memory and requires power to maintain stored information. In some embodiments, the device is non-volatile memory and retains stored information when the digital processing device is not powered. In further embodiments, the non-volatile memory comprises flash memory. In some embodiments, the non-volatile memory comprises dynamic random-access memory (DRAM). In some embodiments, the non-volatile memory comprises ferroelectric random access memory (FRAM). In some embodiments, the non-volatile memory comprises phase-change random access memory (PRAM). In other embodiments, the device is a storage device including, by way of non -limiting examples, CD-ROMs, DVDs, flash memory devices, magnetic disk drives, magnetic tapes drives, optical disk drives, and cloud computing based storage. In further embodiments, the storage and / or memory device is a combination of devices such as those disclosed herein.

[0199] In some embodiments, a digital processing device includes a display to send visual information to a user. In some embodiments, the display is a cathode ray tube (CRT). In some embodiments, the display is a liquid crystal display (LCD). In further embodiments, the display is a thin film transistor liquid crystal display (TFT-LCD). In some embodiments, the display is an organic light emitting diode (OLED) display. In various further embodiments, on OLED display is a passive-matrix OLED (PMOLED) or active-matrix OLED (AMOLED) display. In some embodiments, the display is a plasma display. In other embodiments, the display is a video projector. In still further embodiments, the display is a combination of devices such as those disclosed herein.

[0200] In some embodiments, a digital processing device includes an input device to receive information from a user. In some embodiments, the input device is a keyboard. In some embodiments, the input device is a pointing device including, by way of non -limiting examples, a mouse, trackball, track pad, joystick, game controller, or stylus. In some embodiments, the input device is a touch screen or a multi-touch screen. In otherembodiments, the input device is a microphone to capture voice or other sound input. In other embodiments, the input device is a video camera to capture motion or visual input. In still further embodiments, the input device is a combination of devices such as those disclosed herein.

[0201] In some embodiments, a digital processing device includes a digital camera. In some embodiments, a digital camera captures digital images. In some embodiments, the digital camera is an autofocus camera. In some embodiments, a digital camera is a charge - coupled device (CCD) camera. In further embodiments, a digital camera is a CCD video camera. In other embodiments, a digital camera is a complementary metal-oxide- semiconductor (CMOS) camera. In some embodiments, a digital camera captures still images. In other embodiments, a digital camera captures video images. In various embodiments, suitable digital cameras include 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, and higher megapixel cameras, including increments therein. In some embodiments, a digital camera is a standard definition camera. In other embodiments, a digital camera is an HD video camera. In further embodiments, an HD video camera captures images with at least about 1280 x about 720 pixels or at least about 1920 x about 1080 pixels. In some embodiments, a digital camera captures color digital images. In other embodiments, a digital camera captures grayscale digital images. In various embodiments, digital images are stored in any suitable digital image format. Suitable digital image formats include, by way of non-limiting examples, Joint Photographic Experts Group (JPEG), JPEG 2000, Exchangeable image file format (Exif), Tagged Image File Format (TIFF), RAW, Portable Network Graphics (PNG), Graphics Interchange Format (GIF), Windows® bitmap (BMP), portable pixmap (PPM), portable graymap (PGM), portable bitmap file format (PBM), and WebP. In various embodiments, digital images are stored in any suitable digital video format. Suitable digital video formats include, by way of non-limiting examples, AVI, MPEG, Apple® QuickTime®, MP4, AVCHD®, Windows Media®, DivX™, Flash Video, Ogg Theora, WebM, and RealMedia.

[0202] In many aspects, the systems, platforms, software, networks, and methods disclosed herein include one or more non -transitory computer readable storage media encoded with a program including instructions executable by the operating system of an optionally networked digital processing device. For instance, in some aspects, the methods comprise creating data files associated with a plurality of sequencing reads from a plurality of samples associated with a food processing facility. In further embodiments, a computer readable storage medium is a tangible component of a digital processing device. In stillfurther embodiments, a computer readable storage medium is optionally removable from a digital processing device. In some embodiments, a computer readable storage medium includes, by way of non-limiting examples, CD-ROMs, DVDs, flash memory devices, solid state memory, magnetic disk drives, magnetic tape drives, optical disk drives, cloud computing systems and services, and the like. In some cases, the program and instructions are permanently, substantially permanently, semi -permanently, or non-transitorily encoded on the media.

[0203] In some embodiments, the systems, platforms, software, networks, and methods disclosed herein include at least one computer program. A computer program includes a sequence of instructions, executable in the digital processing device’s CPU, written to perform a specified task. In light of the disclosure provided herein, those of skill in the art will recognize that a computer program maybe written in various versions of various languages. In some embodiments, a computer program comprises one sequence of instructions. In some embodiments, a computer program comprises a plurality of sequences of instructions. In some embodiments, a computer program is provided from one location. In other embodiments, a computer program is provided from a plurality of locations. In various embodiments, a computer program includes one or more software modules. In various embodiments, a computer program includes, in part or in whole, one or more web applications, one or more mobile applications, one or more standalone applications, one or more web browser plug-ins, extensions, add-ins, or add-ons, or combinations thereof.

[0204] In some embodiments, a computer program includes a web application. In light of the disclosure provided herein, those of skill in the art will recognize that a web application, in various embodiments, utilizes one or more software frameworks and one or more database systems. In some embodiments, a web application is created upon a software framework such as Microsoft® .NET or Ruby on Rails (RoR). In some embodiments, a web application utilizes one or more database systems including, by way of non-limiting examples, relational, non-relational, object oriented, associative, and XML database systems. In further embodiments, suitable relational database systems include, by way of non -limiting examples, Microsoft® SQL Server, mySQL™, and Oracle®. Those of skill in the art will also recognize that a web application, in various embodiments, is written in one or more versions of one or more languages. A web application may be written in one or more markup languages, presentation definition languages, client-side scripting languages, server-side coding languages, database query languages, or combinations thereof. In some embodiments, a web application is written to some extent in a markup language such as Hypertext MarkupLanguage (HTML), Extensible Hypertext Markup Language (XHTML), or extensible Markup Language (XML). In some embodiments, a web application is written to some extent in a presentation definition language such as Cascading Style Sheets (CSS). In some embodiments, a web application is written to some extent in a client-side scripting language such as Asynchronous Javascript and XML (AJAX), Flash® Actionscript, Javascript, or Silverlight®. In some embodiments, a web application is written to some extent in a serverside coding language such as Active Server Pages (ASP), ColdFusion®, Perl, Java™, JavaServer Pages (JSP), Hypertext Preprocessor (PHP), Python™, Ruby, Tel, Smalltalk, WebDNA®, or Groovy. In some embodiments, a web application is written to some extent in a database query language such as Structured Query Language (SQL). In some embodiments, a web application integrates enterprise server products such as IBM® Lotus Domino®. A web application for providing a career development network for artists that allows artists to upload information and media files, in some embodiments, includes a media player element. In various further embodiments, a media player element utilizes one or more of many suitable multimedia technologies including, by way of non-limiting examples, Adobe® Flash®, HTML 5, Apple® QuickTime®, Microsoft® Silverlight®, Java™, and Unity®.

[0205] In some embodiments, a computer program includes a mobile application provided to a mobile digital processing device. In some embodiments, the mobile application is provided to a mobile digital processing device at the time it is manufactured. In other embodiments, the mobile application is provided to a mobile digital processing device via the computer network described herein.

[0206] In view of the disclosure provided herein, a mobile application is created by techniques known to those of skill in the art using hardware, languages, and development environments known to the art. Those of skill in the art will recognize that mobile applications are written in several languages. Suitable programming languages include, by way of non-limiting examples, C, C++, C#, Objective-C, Java™, Javascript, Pascal, Object Pascal, Python™, Ruby, VB.NET, WML, and XHTML / HTML with or without CSS, or combinations thereof.

[0207] Suitable mobile application development environments are available from several sources. Commercially available development environments include, by way of non- limiting examples, Airplay SDK, alcheMo, Appcelerator®, Celsius, Bedrock, Flash Lite, .NET Compact Framework, Rhomobile, and WorkLight Mobile Platform. Other development environments are available without cost including, by way of non -limiting examples, Lazarus, MobiFlex, MoSync, and Phonegap. Also, mobile device manufacturers distributesoftware developer kits including, by way of non -limiting examples, iPhone and iPad (iOS) SDK, Android™ SDK, BlackBerry® SDK, BREW SDK, Palm® OS SDK, Symbian SDK, webOS SDK, and Windows® Mobile SDK.

[0208] Those of skill in the art will recognize that several commercial forums are available for distribution of mobile applications including, by way of non -limiting examples, Apple® App Store, Android™ Market, BlackBerry® App World, App Store for Palm devices, App Catalog for webOS, Windows® Marketplace for Mobile, Ovi Store for Nokia® devices, Samsung® App s, and Nintendo® DSi Shop.

[0209] In some embodiments, a computer program includes a standalone application, which is a program that is run as an independent computer process, not an add-on to an existing process, e.g., not a plug-in. Those of skill in the art will recognize that standalone applications are often compiled. A compiler is a computer program(s) that transforms source code written in a programming language into binary object code such as assembly language or machine code. Suitable compiled programming languages include, by way of non-limiting examples, C, C++, Objective-C, COBOL, Delphi, Eiffel, Java™, Lisp, Python™, Visual Basic, and VB .NET, or combinations thereof. Compilation is often performed, at least in part, to create an executable program. In some embodiments, a computer program includes one or more executable complied applications.

[0210] In some embodiments, after a first body of the 3D object is produced, the movable stage is removed from the actuator system. The 3D object may then continue to further processing steps, such as a perfusion sequence as described herein. The systems, platforms, software, networks, and methods disclosed herein include, in various embodiments, software, server, and database modules. In view of the disclosure provided herein, software modules are created by techniques known to those of skill in the art using machines, software, and languages known to the art. The software modules disclosed herein are implemented in a multitude of ways. In various embodiments, a software module comprises a file, a section of code, a programming object, a programming structure, or combinations thereof. In further various embodiments, a software module comprises a plurality of files, a plurality of sections of code, a plurality of programming objects, a plurality of programming structures, or combinations thereof. In various embodiments, the one or more software modules comprise, by way of non -limiting examples, a web application, a mobile application, and a standalone application. In some embodiments, software modules are in one computer program or application. In other embodiments, software modules are in more than one computer program or application. In someembodiments, software modules are hosted on one machine. In other embodiments, software modules are hosted on more than one machine. In further embodiments, software modules are hosted on cloud computing platforms. In some embodiments, software modules are hosted on one or more machines in one location. In other embodiments, software modules are hosted on one or more machines in more than one location.

[0211] In some embodiments, the additive manufacturing system features multiple light sources. In some embodiments, each of the light sources comprise different optic assemblies that result in different pixel sizes on the focal plane. In some embodiments, the combination of light from the multiple light sources with multiple optical assemblies allows for a single print sequence to utilize varying spatial resolutions resulting in a superior printing during. In this case, only areas requiring high resolution receive light from the high- resolution light source, while other areas receive light from the lower resolution light source, thereby allowing a layer with large area to be polymerized faster than by only using a single high resolution light source.EXAMPLESPRINT SEQUENCE I

[0212] With regards to FIG. 2A an embodiment of an additive manufacturing system (130) with the internal subsystem (120) positioned in print position is shown. In some embodiments, the removable cartridge (140) is inserted in print position, aligned with print camera (980), linear axis (910) and a projector (950) containing light source and spatial light modulator to initiate photopolymerization.

[0213] With regards to FIG. 2B an embodiment of an additive manufacturing system (130) with the internal subsystem (120) and the removable cartridge (140) positioned in imaging position is shown. In some embodiments, the removable cartridge (140) is moved to imaging position for post-print analysis after a build sequence is complete.

[0214] In some embodiments, the method of additively manufacturing a 3D object comprises the system (130) comprised of a housing (110) that serves as an interface between the internal subsystem (120) and the host spacecraft or module. In some embodiments, the housing (110) comprises an avionics spine (150) that passes all electrical power and data between the host spacecraft or module and the system internals.

[0215] In some embodiments, the internal subsystem (120) comprises a projector (950) containing light source and spatial light modulator, projection optics (940), linear axis (910), and removable cartridge (140). In some embodiments, the removable cartridge (140)contains a print chamber (1250), heaters, sensors, one or more fluid reservoirs (1220) to contain the photopolymerizable material, one or more fluid reservoirs (1220) to contain a wash or perfusion solution, a pump (1270), a microcontroller (1210), and the necessary tubing.

[0216] In some embodiments, the method comprises a local user inserting a prefilled removable cartridge (140) into the internal subsystem (120). In some embodiments, the removable cartridge (1 0) is prefilled with the desired photopolymerizable material in one fluid reservoir (1220), and the desired wash solution in another fluid reservoir (1220). In some embodiments, once inserted, the microcontroller (1210) is connected to the avionics spine (150) via the digital microcontroller connection interface (1240). In some embodiments, after the microcontroller (1210) is connected to the avionics spine (150), the housing (110) door is then closed. In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the host spacecraft and controlled remotely. In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the host spacecraft and controlled manually.

[0217] In some embodiments, a user establishes connection with the machine via the software interface. In some embodiments, the user prompts the software to initiate a heating sequence of the removable cartridge (140). In some embodiments, the heaters in the removable cartridge warm the print chamber (1250) and fluid reservoirs (1220) to the desired temperature. In some embodiments, the method comprises maintaining that temperature via the temperature sensors contained in the removable cartridge. In some embodiments, at any time, the user can load a design file into the software for an object that is intended to be fabricated. In some embodiments, the software receives the design file), slices the file into cross sections, and generates instructions that control the motor of the linear axis. In some embodiments, the deign file is a CAD file. In some embodiments, alternatively, the user may elect to upload their own instructions to control the motors.

[0218] In some embodiments, once the desired temperature has been reached, and the appropriate design file has been uploaded, the user can begin the fabrication sequence.

[0219] In some embodiments, the fabrication sequence begins with the projector (950) containing light source and spatial light modulator turning on, and the spatial light modulator displaying the cross section of the first layer to be printed. In some embodiments, the cross section comprises a patterned light. In some embodiments, the patterned light isfocused through the projection optics (940) and passed through the removable cartridge window (1430) andprint chamber window (1280) and onto the movable stage (1230), which begins with a thin layer of photopolymerizable material between its surface and the print chamber window (1280). In some embodiments, after the proper exposure duration, the linear axis (910) controlling the motion of the movable stage (1230) begins to incrementally move the movable stage (1230) away from the print chamber window (1280). In some embodiments, the proper exposure duration is between about 0.1 seconds to about 10 seconds. In some embodiments, the proper exposure duration is between about 2 to about 5 seconds. In some embodiments, the proper exposure duration is between about 0.1 seconds to about 10 seconds per layer. In some embodiments, the proper exposure duration is between about 2 to about 5 seconds per layer. In some embodiments, as the print chamber (1250) is moved away from the window (1280), more photopolymerizable material is free to flow around the movable stage (1230) and coverthe newly formed space between the previously formed layer and the inner surface of the window (1280).

[0220] In some embodiments, the method further comprises photopolymerizable material may be selected to enter the print chamber (1250) via active or passive flow from the fluid reservoir (1220). In some embodiments, at this time, the wash fluid reservoir (1220) is closed and cannot allow fluid to enter the print chamber (1250).

[0221] In some embodiments, as the movable stage (1230) is incrementally moved away from the transparent window (1280), and fluid flows to replace the space of the previously formed layer, the spatial light modulator displays a sequence of images that correspond to the appropriate cross section of the layer to be formed at that moment.

[0222] In some embodiments, the synchronization of the movable stage’s (1230) motion and the sequence of images displayed by the spatial light modulator allow for the correct cross-sectional layers to be formed at the correct positions along the 3D object that is being created.

[0223] In some embodiments, the motion of the movable stage (1230), the intensity of the light source, and the composition of the photopolymerizable material all play a crucial role in determining the fabrication speed and resolution.

[0224] In some embodiments, after the entire 3D object has been created, the microcontroller (1210) instructs the pump (1270) to actively flow wash solution from the wash fluid reservoir (1220) into the print chamber (1250). In some embodiments, during this time, the photopolymerizable material is displaced from the print chamber (1250) and into thefluid reservoir (1220) wherein the photopolymerizable material originated, turning that fluid reservoir into a waste bag.

[0225] In some embodiments, the wash solution is pumped until it has displaced all the photopolymerizable material from the print chamber (1250), and then allowed to sit for an established duration of time that allows any unpolymerized material to diffuse from the printed object into the surrounding wash fluid. In some embodiments, the washing step is repeated three times to ensure that the printed object is adequately washed of all unpolymerized material before being imaged.

[0226] In some embodiments, once the fabrication and wash sequences are complete, the removable cartridge (140) is moved to the imaging station. In some embodiments, the camera (980) provides real time analysis capability for the user to see the features of the newly printed object. In some embodiments, the imaging station motor (970) allows the user to move the removable cartridge (140) so that different regions of the object can be viewed by the camera (980). In some embodiments, the information from imaging is used for material calibration (increasing or reducing exposure, light intensity, or print speed), and product quality assurance.

[0227] In some embodiments, after imaging, the removable cartridge (140) containing the newly printed object is placed in storage until needed.

[0228] In some embodiments, the entire fabrication sequence can be repeated by replacing the removable cartridge (140) with a new cartridge and selecting the desired design file.PRINT SEQUENCE II

[0229] In some embodiments, the method of additively manufacturing a 3D object comprises a local user inserting a prefilled removable cartridge (140) into the internal subsystem (120).

[0230] In some embodiments, the removable cartridge (140) is prefilled with the desired photopolymerizable material in one fluid reservoir (1220), the desired wash solution in another fluid reservoir (1220), and a cell media solution in another fluid reservoir (1220).

[0231] In some embodiments, once inserted, the microcontroller (1210) is connected to the avionics spine (150) via the digital microcontroller connection interface (1240). In some embodiments, the housing (110) door is then closed. In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the hostspacecraft and controlled remotely. In some embodiments, the power and data connections are established between the avionics spine ( 150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the host spacecraft and controlled manually.

[0232] In some embodiments, a user then establishes connection with the machine via the software interface. In some embodiments, the user begins by prompting the software to initiate a heating sequence of the removable cartridge (140). In some embodiments, the heaters in the removable cartridge begin warming the print chamber (1250) and fluid reservoirs (1220) to the desired temperature and maintains that temperature via the temperature sensors contained in the removable cartridge. In some embodiments, at any time, the user can load a design file into the software for an object that is intended to be fabricated. In some embodiments, the software receives the design file, slices the file into cross sections, and generates instructions that control the motor of the linear axis. In some embodiments, the design file is a CAD file. In some embodiments, alternatively, the user may elect to upload their own instructions to control the motors.

[0233] In some embodiments, once the desired temperature has been reached, and the appropriate design file has been uploaded, the user can begin the fabrication sequence. In some embodiments, the fabrication sequence begins with the light source turning on, and the spatial light modulator displaying the cross section of the first layer to be printed. In some embodiments, the cross section comprises a patterned light. In some embodiments, the patterned light is focused through the projection optics (940) and passed through the removable cartridge window (1430) and print chamber window (1280) and onto the movable stage (1230), which begins with a thin layer of photopolymerizable material between its surface and the window (1280).

[0234] In some embodiments, after the proper exposure duration, the linear axis (910) controlling the motion of the movable stage (1230) begins to incrementally move the movable stage (1230) away from the print chamber window (1280). In some embodiments, as the print chamber (1250) is moved away from the window (1280), more photopolymerizable material is free to flow around the movable stage (1230) and cover the newly formed space between the previously formed layer and the inner surface of the window (1280).

[0235] In some embodiments, the method further comprises photopolymerizable material may be selected to enter the print chamber (1250) via active or passive flow from the fluid reservoir (1220). In some embodiments, at this time, the wash fluid and cell media reservoirs (1220) are closed and cannot allow fluid to enter the print chamber (1250).

[0236] In some embodiments, as the movable stage (1230) is incrementally moved away from the transparent window (1280), and fluid flows to replace the space of the previously formed layer, the spatial light modulator displays a sequence of images that correspond to the appropriate cross section of the layer to be formed at that moment. In some embodiments, the synchronization of the movable stage’s (1230) motion and the sequence of images displayed by the spatial light modulator allow for the correct cross-sectional layers to be formed at the correct positions along the 3D object that is being created. In some embodiments, the motion of the movable stage (1230), the intensity of the light source, and the composition of the photopolymerizable material all play a crucial role in determining the fabrication speed and resolution.

[0237] In some embodiments, after the entire 3D object has been created, the microcontroller (1210) instructs the pump (1270) to actively flow wash solution from the wash fluid reservoir (1220) into the print chamber (1250). In some embodiments, during this time, the photopolymerizable material is displaced from the print chamber (1250) and into the fluid reservoir (1220) that it came from, turning that fluid reservoir into a waste bag. In some embodiments, the wash solutionis pumped until it has displaced all the photopolymerizable material from the print chamber (1250), and then allowed to sit for an established duration of time that allows any unpolymerized material to diffuse from the printed object into the surrounding wash fluid. In some embodiments, the washing step is repeated three times to ensure that the printed object is adequately washed of all unpolymerized material before being imaged.

[0238] In some embodiments, once the fabrication and wash sequences are complete, the removable cartridge (140) is moved to the imaging station. In some embodiments, the camera (980) provides real time analysis capability for the user to see the features of the newly printed object. In some embodiments, the imaging station motor (970) allows the user to move the removable cartridge (140) so that different regions of the object can be viewed by the camera (980). In some embodiments, the information from imaging is used for material calibration and product quality assurance. In some embodiments, the material calibration includes increasing or reducing exposure, light intensity, print speed, or a combination thereof.

[0239] In some embodiments, after imaging, the removable cartridge (140) containing the newly printed object is placed in storage until needed. In some embodiments, the entire fabrication sequence canbe repeated by replacing the removable cartridge (140) with a new cartridge and selecting the desired design file.

[0240] In some embodiments, at any time after the wash sequence is complete, the cell media contained within its respective fluid reservoir (1220) may be pumped into the newly printed construct. In some embodiments, by aligning the lumens of the newly printed construct with the inlet and outlets corresponding to the cell media fluid reservoir (1220), and initiating a pump sequence, the cell media inside of its respective fluid reservoir (1220) can be perfused into and through the newly printed construct.

[0241] In some embodiments, the cell media comprises a particular cell type. In some embodiments, the cell type comprises Human Umbilical-Vein Epithelial Cells (HUVECs), with the intention of lining the interior lumens of the newly printed construct with these cells.

[0242] In some embodiments, the cell media solution may be replaced with another wash solution, with the intention of removing any unpolymerized photopolymerizable material from the print sequence. In some embodiments, the wash solution to be perfused through the newly printed construct may contain electrically conductive materials or particles. In another some embodiments, the wash solution or cell media to be perfused through the lumens of the newly printed construct may contain micro or nanoparticles.

[0243] In some embodiments, these micro or nanoparticles may contain within them a therapeutic agent to be released into the lumens of the newly printed construct.PRINT SEQUENCE III

[0244] In some embodiments, the method comprises a local user inserting a prefilled removable cartridge (140) into the internal subsystem (120). In some embodiments, the removable cartridge (140) is prefilled with a variety of different desired photopolymerizable materials in different fluid reservoirs (1220), and the desired wash solution in another fluid reservoir (1220). In some embodiments, once inserted, the microcontroller (1210) is connected to the avionics spine (150) via the digital microcontroller connection interface (1240). In some embodiments, the housing (110) door is then closed. In some embodiments, the power and data connections are established between the avionics spine ( 150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the host spacecraft and controlled remotely.

[0245] In some embodiments, a user then establishes connection with the machine via the software interface. In some embodiments, the user begins by prompting the software to initiate a heating sequence of the removable cartridge (140). In some embodiments, the heaters in the removable cartridge begin warming the print chamber (1250) and fluid reservoirs (1220) to the desired temperature. In some embodiments, the desired temperatureis between about 37°C to about 42°C. In some embodiments, the method comprises maintaining that temperature via the temperature sensors contained in the removable cartridge.

[0246] In some embodiments, at any time, the user can load a design file into the software for an object that is intended to be fabricated. In some embodiments, the software receives the design file, slices the file into cross sections, determines which photopolymerizable material will correspond to which areas of the object to be printed, and generates instructions that control the motor of the linear axis and fluid pumps (1270). In some embodiments, the design file is a CAD file. In some embodiments, the user may elect to upload their own instructions to control the motors and fluid pumps (1270).

[0247] In some embodiments, once the desired temperature has been reached, and the appropriate design file has been uploaded, the user can begin the fabrication sequence. In some embodiments, the fabrication sequence begins with the light source turning on, and the spatial light modulator displaying the cross section of the first layer to be printed. In some embodiments, the cross section comprises a patterned light. In some embodiments, the patterned light is focused through the projection optics (940) and passed through the removable cartridge window (1430) and print chamber window (1280) and onto the movable stage (1230), which begins with a thin layer of photopolymerizable material between its surface and the window (1280). In some embodiments, after the proper exposure duration, the linear axis (910) controlling the motion of the movable stage (1230) begins to incrementally move the movable stage (1230) away from the print chamber window (1280). In some embodiments, as the print chamber (1250) is moved away from the window (1280), a selected photopolymerizable material may be actively pumped to flow around the movable stage (1230) and cover the newly formed space between the previously formed layer and the inner surface of the window (1280). In some embodiments, the proper exposure duration is between about 0.1 seconds to about 10 seconds. In some embodiments, the proper exposure duration is between about 2 to about 5 seconds. In some embodiments, the proper exposure duration is between about 0.1 seconds to about 10 seconds per layer. In some embodiments, the proper exposure duration is between about 2 to about 5 seconds per layer.

[0248] In some embodiments, by predetermining which photopolymerizable material is pumped into the newly formed space between the movable stage (1230) and the window (1280), the user has control over the composition of the layers of the construct being printed. In some embodiments, at this time, the wash fluid reservoir (1220) is closed and cannot allow fluid to enter the print chamber (1250).

[0249] In some embodiments, as the movable stage (1230) is incrementally moved away from the transparent window (1280), and fluid flows to replace the space of the previously formed layer, the spatial light modulator displays a sequence of images that correspond to the appropriate cross section of the layer to be formed at that moment. In some embodiments, the synchronization of the movable stage’s (1230) motion, the injection of new photopolymerizable material, and the sequence of images displayed by the spatial light modulator allow for the correct cross-sectional layers to be formed with the correct materials at the correct positions alongthe 3D object that is being created. In some embodiments, the motion of the movable stage (1230), the intensity of the light source, and the composition of the photopolymerizable material all play a crucial role in determining the fabrication speed and resolution.

[0250] In some embodiments, after the entire 3D object has been created, the microcontroller (1210) instructs the pump (1270) to actively flow wash solution from the wash fluid reservoir (1220) into the print chamber (1250). In some embodiments, during this time, the photopolymerizable material is displaced from the print chamber (1250) and into the fluid reservoir (1220) that it came from, turning that fluid reservoir into a waste bag. In some embodiments, the wash solutionis pumped until it has displaced all the photopolymerizable material from the print chamber (1250), and then allowed to sit for an established duration of time that allows any unpolymerized material to diffuse from the printed object into the surrounding wash fluid. In some embodiments, the washing step is repeated three times to ensure that the printed object is adequately washed of all unpolymerized material before being imaged.

[0251] In some embodiments, once the fabrication and wash sequences are complete, the removable cartridge (140) is moved to the imaging station. In some embodiments, the camera (980) provides real time analysis capability for the user to see the features of the newly printed object. In some embodiments, the imaging station motor (970) allows the user to move the removable cartridge (140) so that different regions of the object can be viewed by the camera (980). In some embodiments, the information from imaging is used for material calibration (increasing or reducing exposure, light intensity, or print speed), and product quality assurance.

[0252] In some embodiments, after imaging, the removable cartridge (140) containing the newly printed object is placed in storage until needed. In some embodiments, the entire fabrication sequence canbe repeated by replacing the removable cartridge (140) with a new cartridge and selecting the desired design file.PRINT SEQUENCE IV

[0253] In some embodiments, the method comprises a local user inserting a prefilled removable cartridge (140) into the internal subsystem (120). In some embodiments, the removable cartridge (1 0) is prefilled with the desired photopolymerizable material in one fluid reservoir (1220), and the desired wash solution in another fluid reservoir (1220). In some embodiments, once inserted, the microcontroller (1210) is connected to the avionics spine (150) via the digital microcontroller connection interface (1240). In some embodiments, the housing (110) door is then closed. In some embodiments, the power and data connections are established between the avionics spine (150) and the host spacecraft via the connectors on the housing (110), allowing the system (130) to be powered by the host spacecraft and controlled remotely.

[0254] In some embodiments, the user then establishes connection with the machine via the software interface. In some embodiments, the user begins by prompting the software to initiate a heating sequence of the removable cartridge (140). In some embodiments, the heaters in the removable cartridge begin warming the print chamber (1250) and fluid reservoirs (1220) to the desired temperature. In some embodiments, the method comprises maintaining that temperature via the temperature sensors contained in the removable cartridge. In some embodiments, at any time, the user can load a design file into the software for an object that is intended to be fabricated. In some embodiments, the software receives the design file, slices the file into cross sections, determines the number of fields of view required to span a particular cross-sectional layer, and generates instructions that control the motors of the linear axes that move / scroll the light source(s) relative to the movable stage in the XY direction as required by the cross sections and their constituent fields of view, and the motor of the linear axis that moves the movable stage away from the light source in the Z direction. In some embodiments, the user may elect to upload their own instructions to control the motors.

[0255] In some embodiments, once the desired temperature has been reached, and the appropriate design file has been uploaded, the user can begin the fabrication sequence. In some embodiments, the fabrication sequence begins with the light source turning on, and the spatial light modulator displaying the first field of view of the first cross section of the first layer to be printed. In some embodiments, the patterned light is focused through the projection optics (940) andpassed through the removable cartridge window (1430) and print chamber window (1280) and onto the movable stage (1230), which begins with a thin layer ofphotopolymerizable material between its surface and the window (1280). In some embodiments, after the proper exposure duration, the linear axes controlling the motion of the light source(s) relative to the movable stage in the XY direction begin to incrementally move along the movable stage (1230). In some embodiments, as the light source(s) is moved relative to the movable stage, the image(s) displayed by the spatial light modulator(s) begin to change as required by the new area of the cross section being exposed by the light source. In some embodiments, the synchronization of the light source(s) motion relative to the movable stage (1230) alongthe XY axes, and the sequence of images displayed by the spatial light modulator allow for the correct pattern to be formed along the current cross-sectional layer of the 3D object being created. In some embodiments, the motion of the light source(s) relative to the movable stage (1230) alongthe XY axes, the intensity of the light source, and the composition of the photopolymerizable material all play a crucial role in determining the fabrication speed and resolution of the particular layer being formed.

[0256] In some embodiments, after the first cross-sectional layer had been formed, the linear axis (910) controlling the motion of the movable stage (1230) in the Z direction begins to incrementally move the movable stage (1230) away from the print chamber window (1280). In some embodiments, as the print chamber (1250) is moved away from the window (1280), more photopolymerizable material is free to flow around the movable stage (1230) and cover the newly formed space between the previously formed lay er and the inner surface of the window (1280). In some embodiments, the photopolymerizable material may be selected to enter the print chamber (1250) via active or passive flow from the fluid reservoir (1220). In some embodiments, at this time, the wash fluid reservoir (1220) is closed and cannot allow fluid to enter the print chamber (1250).

[0257] In some embodiments, once the movable stage (1230) has moved away from the transparent window (1280) by one increment equal to the thickness of the next layer to be formed, and fluid flows to replace the space of the previously formed layer, the spatial light modulator(s), in synchronization with the linear axes controlling the motion of the light source(s) relative to the movable stage (1230) displays the next sequence of images that correspond to the appropriate cross section of the layer to be formed. In some embodiments, the synchronization of the light source(s) motion relative to the movable stage (1230) along the XY axes, and the sequence of images displayed by the spatial light modulator allow for the correct pattern to be formed alongthe current cross-sectional layer of the 3D object being created. In some embodiments, the motion of the light source(s) relative to the movable stage (1230) along the XY axes, the intensity of the light source, and the composition of thephotopolymerizable material all play a crucial role in determining the fabrication speed and resolution of the next layer to be formed. In some embodiments, the process may repeat for as many layers as required until the entire 3D object has been created.

[0258] In some embodiments, after the entire 3D object has been created, the microcontroller (1210) instructs the pump (1270) to actively flow wash solution from the wash fluid reservoir (1220) into the print chamber (1250). In some embodiments, during this time, the photopolymerizable material is displaced from the print chamber (1250) and into the fluid reservoir (1220) that it came from, turning that fluid reservoir into a waste bag. In some embodiments, the wash solutionis pumped until it has displaced all the photopolymerizable material from the print chamber (1250), and then allowed to sit for an established duration of time that allows any unpolymerized material to diffuse from the printed object into the surrounding wash fluid. In some embodiments, the washing step is repeated three times to ensure that the printed object is adequately washed of all unpolymerized material before being imaged.

[0259] In some embodiments, once the fabrication and wash sequences are complete, the removable cartridge (140) is moved to the imaging station. In some embodiments, the camera (980) provides real time analysis capability for the user to see the features of the newly printed object. In some embodiments, the imaging station motor (970) allows the user to move the removable cartridge (140) so that different regions of the object can be viewed by the camera (980). In some embodiments, the information from imaging is used for material calibration (increasing or reducing exposure, light intensity, or print speed), and product quality assurance.

[0260] In some embodiments, after imaging, the removable cartridge (140) containing the newly printed object is placed in storage until needed. In some embodiments, the entire fabrication sequence can be repeated by replacing the removable cartridge (140) with a new cartridge and selecting the desired design file.

[0261] In some embodiments, at any time after the wash sequence is complete, the cell media contained within its respective fluid reservoir (1220) may be pumped into the newly printed construct. In some embodiments, by aligning the lumens of the newly printed construct with the inlet and outlets corresponding to the cell media fluid reservoir (1220), and initiating a pump sequence, the cell media inside of its respective fluid reservoir (1220) can be perfused into and through the newly printed construct.

[0262] In some embodiments, the cell media may contain a particular choice of cell type. In some embodiments, the cell type is Human Umbilical-Vein Epithelial Cells(HUVECs). In some embodiments, the interior lumens of the newly printed construct are lined with these cells.

[0263] In some embodiments, the cell media solution may be replaced with another wash solution, with the intention of removing any unpolymerized photopolymerizable material from the print sequence. In some embodiments, the wash solution to be perfused through the newly printed construct comprises electrically conductive materials or particles.

[0264] In some embodiments, the wash solution or cell media to be perfused through the lumens of the newly printed construct comprises micro or nanoparticles. In some embodiments, the micro or nanoparticles comprises a therapeutic agent. In some embodiments, the therapeutic agents comprise growth factors, pharmaceuticals, a small molecule drug, antibodies, large biomolecules, or any combination thereof. In some embodiments, the therapeutic agent is released into the lumens of the newly printed construct.

[0265] In some embodiments, after a first body of the 3D object is produced, the movable stage is removed from the actuator system. The 3D object may then continue to further processing steps, such as a perfusion sequence as described herein.DEFINITIONS

[0266] Unless defined otherwise, all terms of art, notations and other technical and scientific terms or terminology used herein are intended to have the same meaning as is commonly understood by one of ordinary skill in the art to which the claimed subject matter pertains. In some cases, terms with commonly understood meanings are defined herein for clarity and / or for ready reference, and the inclusion of such definitions herein should not necessarily be construed to represent a substantial difference over what is generally understood in the art.

[0267] Throughout this application, various embodiments may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 6, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 6, and 6. This applies regardless of the breadth of the range.

[0268] As used in the specification and claims, the singular forms “a”, “an” and “the” include plural references unless the context clearly dictates otherwise. For example, the term “a sample” includes a plurality of samples, including mixtures thereof.

[0269] As used herein, the term “about” a number refers to that number plus or minus 10% of that number. The term “about” a range refers to that range minus 10% of its lowest value and plus 10% of its greatest value.

[0270] The section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0271] While preferred embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that various alternatives to the embodiments of the invention described herein may be employed in practicing the invention. It is intended that the following claims define the scope of the invention and that methods and structures within the scope of these claims and their equivalents be covered thereby.

Claims

CLAIMS1. An additive manufacturing system comprising: a housing defining a first sealable compartment, the housing being configurable from a first position in which the housing is sealed, to a second position in which the housing is unsealed, the housing comprising: a door providing access to the compartment when the housing is in the second position; one or more light sources; a removable cartridge defining a second sealable compartment comprising: a. a plurality of fluid reservoirs; b. a print chamber defining a third sealable compartment; c. a movable stage positioned within the print chamber; d. one or more transparent walls permitting for transmission of light into the removable cartridge.

2. The system of claim 1, wherein the removable cartridge is enclosed within the first compartment, wherein the print chamber is enclosed within the second compartment.

3. The system of claim 1, wherein the print chamber is hermetically sealed from the second compartment.

4. The system of claim 1, wherein the removable cartridge is hermetically sealed from the first compartment.

5. The system of claim 1, wherein the print chamber comprises one or more transparent walls permitting for transmission of light into the print chamber.

6. The system of claim 1, wherein the print chamber is fully filled with a liquid and comprises essentially no gas.

7. The system of claim 1, wherein the one or more transparent walls are gas permeable.

8. The system of claim 7, wherein the gas permeability permits an active or passive flow of oxygen.

9. The system of claim 1, wherein the one or more transparent walls comprise a gas inlet.

10. The system of claim 1, further comprising a pressurized oxygen source coupled to an oxygen line, wherein the oxygen line is directly coupled to the one or more transparent walls.

11. The system of claim 1, further comprising one or more fans configured to direct an active flow of oxygen to the one or more transparent walls.

12. The system of claim 1, wherein the removable cartridge comprises a gas inlet directing gas proximally to the one or more transparent walls.

13. The system of claim 1 , wherein the removable cartridge is interchangeable within the system.

14. The system of claim 1, wherein the plurality of fluid reservoirs are removable and interchangeable within the removable cartridge.

15. The system of the preceding claim, wherein the removable cartridge is thereby modular.

16. The system of claim 1, wherein the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the plurality of fluid reservoirs, and provides an outlet of fluid to at least one of the plurality of fluid reservoirs.

17. The system of the preceding claim, wherein the plurality of fluid reservoirs comprise a wash fluid and a photopolymerizable fluid, wherein the outlet of fluid to at least one of the plurality of fluid reservoirs is to the fluid reservoir comprising the photopolymerizable fluid.

18. The system of claim 1, wherein the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage.

19. The system of claim 16 or 18, wherein the perfusion path comprises one or more pumps coupled to the fluid reservoirs and the print chamber.

20. The system of claim 1, wherein the removable cartridge is moveable within the system along at least 2 of an x-axis, a y-axis, and a z-axis.21 . The system of the preceding claim, wherein the removable cartridge is movable relative to at least one light source, thereby allowing a larger exposure area than a single field of view of light form the at least one light source onto the movable stage.

22. The system of claim 1, wherein the moveable stage comprises a patterned surface configured to enhance adhesion between the patterned surface and a fluid or a solid in contact with the patterned surface.

23. The system of claim 22, wherein the patterned surface comprises a plurality of raised regions and recessed regions arranged in a predetermined pattern.

24. The system of claim 23, wherein the raised regions have a height from 0.1 mm to 1 mm.

25. The system of claim 1, wherein the one or more transparent walls comprise an anti- adhesion surface configured to reduce adhesion between a fluid or a solid in contact with the anti-adhesion surface.

26. The system of claim 25, wherein the anti-adhesion surface comprises a coating of a hydrophobic or a superhydrophobic material.

27. The system of claim 25, wherein the anti-adhesion surface comprises a micropattern or nanopattern to induce hydrophobic or superhydrophobic properties.

28. The system of claim 27, wherein the micropattem or nanopattem comprises dimensions less than 500 nm.

29. The system of claim 28, wherein the micropattern or nanopattern comprises dimensions less than 400 nm.

30. The system of claim 1, wherein the removable cartridge further comprises a heating element.

31. The system of claim 1, further comprising one or more imaging devices configured to image the first compartment, or the second compartment.

32. The system of claim 1, further comprising one or more imaging devices configured to image one or more components of the system.

33. The system of claim 1, further comprising one or more imaging devices configured to image an object manufactured within the system.

34. The system of claim 1, further comprising one or more imaging devices configured to image the internals of the system before, during, and after a fabrication sequence.

35. The system of claim 1, further comprising one or more imaging devices configured to image the removable cartridge during a fabrication sequence.

36. The system of claim 1, wherein one or more imaging devices are oriented to image a completed 3D object after the fabrication steps.

37. The system as in one of claims 31-36, wherein the one or more imaging devices comprise one or more optical cameras.

38. The system of claim 1, wherein the system can be placed in any orientation and maintains functionality without regard to a gravitational field.

39. The system of claim 1, wherein the system is configured to operate in microgravity, e.g., low earth orbit, or any orbit around a celestial object.

40. The system of claim 1, wherein the system can be placed in any orientation and maintains functionality without regard to a gravitational field, wherein the system is configured to operate in microgravity.

41. The system of claim 1, wherein the removable cartridge further comprises a thermometer.

42. The system of claim 1, wherein the system is configured to maintain the temperature within the removable cartridge below a critical level.

43. The system of claim 1, further comprising an avionics spine comprising a plurality of electrical and data connections electrically coupling the system to a spacecraft.

44. The system of claim 1, wherein the one or more transparent walls comprises glass, quartz, transparent plastic, silicon, silica, polydimethylsiloxane (PDMS), transparent ceramic, fluorinated ethylene propylene (FEP), or combinations thereof.

45. The system of claim 1, wherein at least one of the plurality of fluid reservoirs comprise a photopolymerizable material, a photo initiator, a chemical quencher, (2, 2, 6, 6- Tetramethylpiperidin-l-yl)oxyl, (2,2,6,6-tetramethylpiperidin-l-yl)oxidanyl (TEMPO), a physical quencher, a wash solution, a biologic solution, cell media, suspension of cells, microparticles, nanoparticles, therapeutic loaded nanoparticles, therapeutic loaded microparticles, electrolytic or electrically conductive materials, or combinations thereof.

46. The system of claim 45, wherein the wash solution comprises water, a buffer, an organic material, a solution containing antibiotics, an antimicrobial agent, or combinations thereof.

47. The system of claim 45, wherein the biologic solution comprises nutrients and / or growth factors suitable for cell culture and proliferation.

48. The system of claim 45, wherein the biologic solution comprises a suspension of cells.

49. The system of claim 1, wherein the one or more light sources are movable within the first compartment.

50. The system of claim 1, wherein the removable cartridge is moveable with the first compartment.

51. The system of claim 1, further comprising a plurality of spatial light modulators configured to direct light from the one or more light sources.

52. The system of claim 51, wherein the plurality of spatial light modulators are configured to direct light along an x-axis, and a y-axis.

53. The system of claim 51, wherein the plurality of spatial light modulators are configured to direct light along an x-axis, a y-axis, and a z-axis.

54. The system of claim 51 , wherein the plurality of spatial light modulators comprise one or more digital micro-mirror devices.

55. The system of claim 51 , wherein the plurality of spatial light modulators comprise one or more liquid crystal devices.

56. The system of claim 1, wherein at least one of the one or more light sources comprises a light emitting diode.

57. The system of claim 1, wherein at least one of the one or more light sources comprises a laser.

58. The system of claim 1, wherein at least one of the one or more light sources is movable along an x-axis, and a y-axis.

59. The system of claim 1, wherein at least one of the one or more light sources is movable along an x-axis, a y-axis, and a z-axis.

60. The system of claim 1, wherein the housing is configured to interface with a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform.

61. The system of claim 1, wherein the removable cartridge comprises a plurality of movable stages, each stage in line with at least one of the one or more transparent walls.

62. The system of claim 61, wherein at least one of the one or more light sources is movable along two or more of an x-axis, a y-axis, and a z-axis relative to the removable cartridge such that multiple optical field-of-views can be exposed onto at least one of the plurality of movable stages, thereby allowing a larger print area than a single field of view.

63. The system of claim 1, further comprising one or more computer processors comprising a computer readable medium configured to implement a method of additively manufacturing a 3D object comprising: a) sealing one or more of the first compartment, the second compartment, or the third compartment from one another; b) filling the print chamber with a photopolymerizable fluid comprised by the plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from the one or more light sources through the one or more transparent walls onto the movable stage and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid, wherein the method takes place in a reduced gravity environment.

64. The system of claim 63, wherein the computer readable medium further comprises instructions for producing a scaffold, a biological tissue, or a microfluidic device.

65. The system of claim 63, wherein the instructions comprise a 3D model of the 3D object in discrete layers which correspond to a printing resolution of the system about a z axis defined by the movement of the movable stage.

66. The system of claim 65, wherein the method further comprises monitoring the 3D object during any of c.-e. with one or more imaging devices.

67. The system of claim 66, wherein the monitoring occurs before, during, and after a fabrication sequence.

68. The system of claim 65, wherein the method further comprises monitoring a component of the system, the first compartment, or the second compartment during any of c. - e. with one or more imaging devices.

69. The system of claim 68, wherein the monitoring occurs before, during, and after a fabrication sequence.

70. The system according to claims 66 or 68, wherein the one or more imaging devices comprise one or more optical cameras.71 . The system of claim 63, wherein the monitoring 3D object comprises monitoring a dimensionality of the solid for compliance with the 3D model.

72. The system of claim 71, wherein the monitoring the dimensionality of the solid for compliance with the 3D model is within 0.6 microns.

73. The system of claim 63, wherein the computer readable medium further comprises instructions for producing the 3D object, the 3D object comprising a plurality of channels.

74. The system of claim 73, wherein the plurality of fluid reservoirs comprise a wash fluid and a photopolymerizable fluid, where the removable cartridge comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber with the inlet aligning with an opening in the moveable stage, wherein the channels align with the inlet, wherein the method further comprise flushing the plurality of channels with the wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels.

75. The system of claim 63, wherein the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising flushing the print chamber with the wash fluid to remove the photopolymerizable fluid, and directing the washfluid and the removed photopolymerizable fluid into one of the fluid reservoirs initially holding the photopolymerizable fluid.

76. The system of claim 63, wherein the directing the light comprises directing two- dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage.

77. The system of claim 76, wherein the method further comprises moving the one or more light sources relative to the movable stage and exposing multiple optical field -of-views onto the moveable stage, thereby allowing a larger print area than a single field of view.

78. The system of claim 63, wherein the method further comprises moving the removeable cartridge relative to the movable stage and exposing multiple optical field -of- views onto the moveable stage, thereby allowing a larger print area than a single field of view.

79. The system of claim 63, wherein the movingthe movable stage comprises movingthe movable stage away from the optical plane such that the projected two-dimensional crosssections of the three-dimensional object are incident on photopolymerizable fluid in contact with and adjacent to previously formed layers of the object.

80. The system of claim 63, wherein the movingthe movable stage comprises movingthe movable stage continuously.

81. The system of claim 63, wherein the movingthe movable stage comprises movingthe movable stage incrementally.

82. The system of claim 63, wherein the directing light comprises intermittently activating the one or more light sources during the formation of the three-dimensional object.

83. The system of claim 63, wherein the method further comprises monitoring a temperature within the removable cartridge with a thermometer and maintaining the temperature below a critical level.

84. The system of claim 63, wherein the method further comprises monitoring a temperature within the removable cartridge with a thermometer, and ceasing operation of the system if a critical temperature is reached.

85. The system of claim 63, wherein the one or more transparent walls are gas permeable, the method further comprising providing an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring against the one or more transparent walls.

86. The system of claim 85, the method further comprising contactingthe top layer of the photopolymerizable fluid with the one or more transparent walls.

87. The system of claim 85, wherein the top layer comprises a thickness of between about 1 to about 200 micrometers.

88. The system of claim 63, wherein the plurality of fluid reservoirs comprise a chemical quencher, physical quencher, or a combination thereof that quenches free-radicals and prevents polymerization, or over polymerization.

89. The system of claim 88, wherein the method further comprises filling the print chamber with the chemical quencher and preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber.

90. The system of claim 89, wherein the target polymerization depth is between about 1 to about 200 micrometers.

91. The system of claim 90, wherein the target polymerization depth is between about 10 to about 50 micrometers.

92. The system of claim 88, wherein a concentration of the physical quencher in the print chamber is between about 1 to about 5% v / v.

93. The system of claim 92, wherein a concentration of the physical quencher in the print chamber is about 4% v / v.

94. The system of claim 63, further comprising mounting the system in an orientation without regard to a gravitational field.

95. The system of claim 63, further comprising operating the system in an orientation without regard to a gravitational field.

96. The system of any of the preceding claims, wherein a printed construct is created.

97. The system of claim 63, wherein the plurality of fluid reservoirs comprises a cell media fluid reservoir.

98. The system of claim 97, wherein the removable cartridge further comprises a perfusion path between the fluid reservoirs and the print chamber which provides an inlet of fluid to the print chamber from at least one of the fluid reservoirs, and provides an outlet of fluid to at least one of the fluid reservoirs, the method further comprising perfusing the cell media through the newly printed construct.

99. The system of claim 63, wherein the moving the stage occurs at a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

100. The system of claim 63, wherein the method further comprises additively manufacturing the 3D objectat a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

101. The system of claim 63, wherein the sealing one or more of the first compartment, the second compartment, or the third compartment from one another comprising hermetically sealing one or more of the first compartment, the second compartment, or the third compartment from one another.

102. A method of additively manufacturing a 3D object comprising: a) sealing one or more of a first compartment, a second compartment, or a third compartment from one another, wherein the second compartment and third compartment comprise one or more transparent walls; b) filling a print chamber with a photopolymerizable fluid comprised by a plurality of fluid reservoirs completely such that there is essentially no gas within the print chamber; c) directing light from a light source through one or more transparent walls of the second compartment or the third compartment onto a movable stage within the print chamber and photopolymerizing a first portion of the photopolymerizable fluid in contact with the movable stage into a solid; d) moving the movable stage within the print chamber away from the one or more transparent walls; and e) photopolymerizing a second portion of the photopolymerizable fluid in contact with the solid, wherein the method takes place in a reduced gravity environment.

103. The method of claim 102, further comprising interfacing the first compartment with a host module in a spacecraft comprising a shelf, a locker, or standardized organizational platform.

104. The method of claim 102, wherein the 3D object is a scaffold, a biological tissue, or a microfluidic device.

105. The method of claim 102, wherein the photopolymerizing comprises printing the 3D object in discrete layers which correspond to a printing resolution of the method about a z axis defined by the movement of the movable stage.

106. The method of claim 102, wherein the method further comprises monitoring the 3D object during any of c.-e. with one or more imaging devices.

107. The method of claim 106, wherein the one or more imaging devices comprise one or more optical cameras.

108. The method of claim 106, wherein the monitoring occurs before, during, and after a fabrication sequence comprising performance of a.-e.

109. The method of claim 108, wherein the monitoring the 3D object comprises monitoring a dimensionality of the solid for compliance with a 3D model.

110. The method of claim 109, wherein the monitoring the dimensionality of the solid for compliance with the 3D model is within 0.6 microns.

111. The method of claim 102, further comprising flushing a plurality of channels of the 3D object with a wash fluid to remove unpolymerized material by directing the wash fluid from the inlet through the channels.

112. The method of claim 111, further comprising directing the wash fluid and the removed photopolymerizable fluid into a fluid reservoir initially holding the photopolymerizable fluid.

113. The method of claim 102, wherein the directing light from a light source comprises directing two-dimensional cross-sections of the three-dimensional objects onto a focused optical plane coincident or above the movable stage.

114. The method of claim 102, further comprising moving the light source relative to the movable stage and exposing multiple optical field -of-views onto the moveable stage, thereby allowing a larger print area than a single field of view.

115. The method of claim 102, further comprising moving a removeable cartridge defining the second compartment relative to the movable stage and exposing multiple optical field-of- views onto the moveable stage, thereby allowing a larger print area than a single field of view.

116. The method of claim 115, wherein the moving the movable stage comprises moving the movable stage away from the optical plane such that the projected two-dimensional crosssections of the three-dimensional object are incident on photopolymerizable fluid in contact with and adjacent to previously formed layers of the object.

117. The method of claim 115, wherein the moving the movable stage comprises moving the movable stage continuously.

118. The method of claim 115, wherein the moving the movable stage comprises moving the movable stage incrementally.

119. The method of claim 102, wherein the directing light from a light source comprises intermittently activating the light source during the formation of the three-dimensional object.

120. The method of claim 102, further comprising monitoring a temperature within the removable cartridge with a thermometer, and maintaining the temperature below a critical level.

121. The method of claim 102, further comprising monitoring a temperature within the removable cartridge with a thermometer, and ceasing operation of the method if a critical temperature is reached.

122. The method of claim 102, further comprising providing an active flow or passive flow of oxygen into a top layer of the photopolymerizable fluid, thereby preventing photopolymerization from occurring in the top layer of the photopolymerizable fluid.

123. The method of claim 102, further comprising contacting a top layer of the photopolymerizable fluid with one or more transparent walls comprising an anti -adhesive surface, a hydrophobic surface, or combinations thereof.

124. The method of claim 102, wherein a top layer of the photopolymerizable fluid comprises a thickness of about 1 to about 200 microns.

125. The method of claim 102, further comprising introducing a chemical quencher into the photopolymerizable fluid, and preventing polymerization, or over polymerization, by quenching a free radical.

126. The method of claim 102, further comprising at least partially filling the print chamber with the chemical quencher and preventing polymerization of the photopolymerizable material beyond a target polymerization depth within the print chamber.

127. The method of claim 122, wherein the top layer is between about 1 to about 200 mm.

128. The method of claim 127, wherein the top layer is between about 10 to about 50 mm.

129. The method of 102, wherein the moving the movable stage occurs at a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

130. The method of claim 102, further comprising additively manufacturing the 3D object at a speed of at least 5, 4, 3, 2, 1, 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, or 0.1 mm / min.

131. The method of claim 102, wherein the sealing one or more of a first compartment, a second compartment, or a third compartment from one another comprises hermetically sealing one or more of a first compartment, a second compartment, or a third compartment from one another.

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