Waste reduction for automatic dispense of density gradients

Abstract

A system for dispensing a set of density gradients. The system calculates a dispense queue including volumes of gradient materials for dispensing the set of density gradients. The set of density gradients including at least a first density gradient and a second density gradient for dispensing into at least a first container and a second container, respectively. The system dispenses the first density gradient into the first container by dispensing one or more sections composed of mixtures of the gradient materials through a probe positioned to have a distal end at a bottom of the first container. The system purges the probe by displacing a mixture of gradient materials used for dispensing a last section of the first density gradient with a material included in the dispense queue before the dispense of the first density gradient is completed.

Description

WASTE REDUCTION FOR AUTOMATIC DISPENSE OF DENSITY GRADIENTSCROSS-REFERENCE TO RELATED APPLICATION

[0001] This application is being filed on October 17, 2024, as a PCT International application and claims the benefit of and priority to U.S. Provisional Application No. 63 / 593,295, filed October 26, 2023; the disclosure of which is hereby incorporated by reference in its entirety.BACKGROUND

[0002] Proteins have a variety of cellular functions, structures, and mechanisms of action. Routinely, proteins bind with other biomolecules, called ligands, in order to complete a task. Researchers can gain valuable knowledge on how7proteins work in a cellular environment by purification of proteins bound to ligands, also called proteinligand complexes. For example, purified protein-ligand complexes are often used in downstream analyses such as high-resolution imaging, sequencing, or crystallography for discovery of protein-based therapeutics.

[0003] Protein-ligand complexes can be purified using density gradients, which are typically layered in steps using an underlay or overlay approach, where solutions are added in order of increasing or decreasing density. A step gradient may be used directly or allowed to diffuse in a controlled manner to create a continuous or linear gradient. A step gradient has sharp interfaces between layers having different densities, whereas a continuous or linear gradient has layers that gradually increase in density moving from top to bottom. In some instances, a step gradient is spun in a centrifuge which causes the layers to diffuse into a continuous or linear gradient.

[0004] The underlay technique for generating a density7gradient includes adding a first layer having a low- density into a tube, and then adding successively denser layers to the bottom of the tube by a long syringe so as to not disturb the previous layers. The overlay technique for generating a density gradient includes using a pipette to dispense decreasingly dense layers sequentially on top of denser layers. These layering techniques for generating density7gradients can be tedious and time-consuming, and are difficult to replicate among researchers.SUMMARY

[0005] The present disclosure generally relates to a system that dispenses density gradients. In one configuration, the system reduces waste when automatically- dispensing density gradients by generating a queue of material for dispensing through a probe. Various aspects are described in this disclosure, which include, but are not limited to, the following aspects.

[0006] One aspect relates to a method of dispensing a set of density gradients, the method comprising: calculating a dispense queue including volumes of gradient materials for dispensing the set of density gradients, the set of density gradients including at least a first density gradient and a second density- gradient for dispensing into at least a first container and a second container, respectively; dispensing the first density gradient into the first container by dispensing one or more sections composed of mixtures of the gradient materials through a probe; and purging the probe by displacing a mixture of gradient materials used for dispensing a last section of the first density gradient with a material included in the dispense queue before the dispense of the first density- gradient is completed.

[0007] Another aspect relates to a system for dispensing a set of density gradients, the system comprising: a processing circuitry having a memory for storing instructions which, when executed by the processing circuitry-, cause the system to: calculate a dispense queue including volumes of gradient materials for dispensing the set of density gradients, the set of density gradients including at least a first density gradient and a second density gradient for dispensing into at least a first container and a second container, respectively; dispense the first density gradient into the first container by dispensing one or more sections composed of mixtures of the gradient materials through a probe; and purge the probe by displacing a mixture of gradient materials used for dispensing a last section of the first density gradient with a material included in the dispense queue before the dispense of the first density- gradient is completed.

[0008] Another aspect relates to a system for dispensing a set of density- gradients, the system comprising: a processing circuitry having a memory for storing instructions which, when executed by the processing circuitry, cause the system to: calculate a dispense queue including volumes of gradient materials for dispensing the set of density- gradients, the set of density gradients including at least a first density' gradient and a second density gradient for dispensing into at least a first container and a second container, respectively; dispense a first section of the first density gradient into the firstcontainer by dispensing a first mixture of the gradient materials through a probe; purge the probe by displacing the first mixture of gradient materials used for dispensing the first section of the first density gradient with a second mixture of gradient materials used for dispensing a second section of the first density gradient; dispense the second section of the first density gradient into the first container; and purge the probe by displacing the second mixture of gradient materials used for dispensing the second section of the first density gradient with a material included in the dispense queue before the dispense of the first density gradient is completed.

[0009] A variety of additional aspects will be set forth in the description that follows. The aspects can relate to individual features and to combinations of features. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the broad inventive concepts upon which the embodiments disclosed herein are based.DESCRIPTION OF THE FIGURES

[0010] The following drawing figures, which form a part of this application, are illustrative of the described technology and are not meant to limit the scope of the disclosure in any manner.

[0011] FIG. 1 schematically illustrates an example of a system for generating density gradients for centrifugation.

[0012] FIG. 2 illustrates an example of a mixer housed inside a manifold and mixing chamber of the system of FIG. 1 .

[0013] FIG. 3 is an isometric view of a probe of the system of FIG. 1, the probe having a distal end inserted into a container, and a proximal end connected to the manifold and mixing chamber.

[0014] FIG. 4 illustrates an example of the proximal end of the probe of FIG. 3 connected to the manifold and mixing chamber.

[0015] FIG. 5 schematically illustrates an example of a method of generating a density gradient inside a container using the system of FIG. 1.

[0016] FIG. 6 schematically illustrates an example of a density gradient formed inside the container after completion of the method of FIG. 5.

[0017] FIG. 7 is an isometric view of a measurement apparatus in the system of FIG. 1 for measuring a density gradient.

[0018] FIG. 8 is a detailed isometric view of the measurement apparatus of FIG. 7.

[0019] FIG. 9 is a front view of the measurement apparatus of FIG. 7.

[0020] FIG. 10 schematically illustrates an example of an electrical configuration for a sensor assembly mounted on the measurement apparatus of FIG. 7.

[0021] FIG. 11 schematically illustrates an example of a method of dispensing a plurality of density gradients that can be performed by the system of FIG. 1.

[0022] FIG. 12 illustrates an example of a graphical user interface that can be displayed on a display of a workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0023] FIG. 13 illustrates another example of a graphical user interface that can be displayed on the display of the workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0024] FIG. 14 illustrates another example of a graphical user interface that can be displayed on the display of the workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0025] FIG. 15 illustrates another example of a graphical user interface that can be displayed on the display of the workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0026] FIG. 16 illustrates another example of a graphical user interface that can be displayed on the display of the workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0027] FIG. 17 illustrates another example of a graphical user interface that can be displayed on the display of the workstation, the graphical user interface including parameters that can be selected by a user of the system of FIG. 1 to define a dispense project.

[0028] FIG. 18 schematically illustrates another example of the system of FIG. 1 showing volumes used for calculating a dispense queue.

[0029] FIG. 19 illustrates another example of a density gradient formed inside a container during performance of the method of FIG. 11.

[0030] FIG. 20 illustrates an example of the probe of the system of FIG. 1 removed from the density gradient of FIG. 19.

[0031] FIG. 21 schematically illustrates an example of a method of dispensing a sample after a density gradient is dispensed in accordance with the method of FIG. 11.

[0032] FIG. 22 illustrates an example of a sample dispensed above a density gradient formed inside a container during performance of the method of FIG. 21.

[0033] FIG. 23 illustrates another example of the sample dispensed above the density gradient formed inside the container during performance of the method of FIG. 21.

[0034] FIG. 24 schematically illustrates an example of a dispense queue that can be calculated by the system of FIG. 1.

[0035] FIG. 25 illustrates example computing hardware of the system of FIG. 1.

[0036] FIG. 26 schematically illustrates an example of a method of improving liquid level sensing after dispensing a density gradient in accordance with the method of FIG. 11 or after dispensing a sample in accordance with the method of FIG. 21.

[0037] FIG. 27 schematically illustrates another example of a method of improving liquid level sensing after dispensing a density gradient in accordance with the method of FIG. 11 or after dispensing a sample in accordance with the method of FIG. 21.

[0038] FIG. 28 provides an illustrative example of a droplet formed on the distal end of the probe of the system of FIG. 1.

[0039] FIG. 29 is a plot illustrating liquid level measurements obtained by the system of FIG. 1 before and after implementation of the method of FIG. 27.

[0040] FIG. 30 shows an isometric view of internal components of an example of the system of FIG. 1 that includes a first portion for dispensing a density gradient and sample, and a second portion for sealing a container after the density gradient and sample are dispensed.

[0041] FIG. 31 shows a front view of the first and second portions of FIG. 30.

[0042] FIG. 32 shows a top isometric view of a carousel that can be mounted on a platform of the system of FIGS. 30 and 31.

[0043] FIG. 33 shows an example of a container held by a holder on the carousel of FIG. 32 that is aligned with a seal former of the second portion of the system of FIG. 30.

[0044] FIG. 34 shows a bottom view of an example of the seal former of FIG. 33.

[0045] FIG. 35 shows an example of a container sealed by the seal former of FIG. 33.

[0046] FIG. 36 schematically illustrates an example of a method of sealing a container that can be performed by the second portion of the system of FIG. 30.

[0047] FIG. 37 schematically illustrates an example of a method of preparing container for centrifugation that can be performed by the system of FIG. 30.DETAILED DESCRIPTION

[0048] FIG. 1 schematically illustrates an example of a system 100 that generates density gradients for centrifugation. The system 100 is computer controlled to precisely dispense a gradient of any type, slope, or shape. The system 100 can be used to generate linear density gradients, which have densities that gradually increase from top to bottom, and step density gradients, which have at least two discrete steps of different densities.

[0049] Additionally, the system 100 can measure a density gradient dispensed inside a container 110 without touching or disturbing the density gradient. These measurements can be used by the system 100 to replicate the density gradient inside another container. In some examples, the container 110 is a tube for use in a centrifuge rotor for centrifugation.

[0050] The system 100 can include features and provide functionalities similar to those described in U.S. Provisional Patent Application No. 63 / 369.299, filed July 25, 2022; U.S. Provisional Patent Application No. 63 / 375,558, filed September 14, 2022; U.S. Provisional Patent Application No. 63 / 369,306, filed July 25, 2022; U.S. Provisional Patent Application No. 63 / 375,563, filed September 14, 2022; U.S. Provisional Patent Application No. 63 / 369,318, filed July 25. 2022; U.S. Provisional Patent Application No. 63 / 375,564, filed September 14, 2022; and PCT Patent Application No. PCT / US2023 / 028403, filed July 21, 2023, the entire disclosures of which are herein incorporated by reference in their entireties.

[0051] The system 100 includes reservoirs 102 that each hold a separate gradient material for generating a density gradient inside the container 110. Each reservoir 102 is connected to a pump 104 for pumping the gradient material held in the reservoir 102 into a mixing manifold 106. Each of the pumps 104 is programmed to pump the gradient materials from the reservoirs 102 at a given volume and speed for mixinginside the mixing manifold 106. The gradient materials housed inside the reservoirs are liquids that can have different densities.

[0052] In the example of FIG. 1, the system 100 includes four reservoirs such as a first reservoir 102a connected to a first pump 104a for pumping a first gradient material into the mixing manifold 106, a second reservoir 102b connected to a second pump 104b for pumping a second gradient material into the mixing manifold 106, a third reservoir 102c connected to a third pump 104c for pumping a third gradient material into the mixing manifold 106, and a fourth reservoir 102d connected to a fourth pump 104d for pumping a fourth gradient material into the mixing manifold 106. The system 100 can include more than four reservoirs for holding more than four gradient materials, or can include fewer than four reservoirs for holding fewer than four gradient materials for generating a density gradient in the container 110.

[0053] The gradient materials held in the reservoirs 102 are liquids pumped into the mixing manifold 106 for dispensing homogenous streams of fluid into the container 110. As an illustrative example, the first reservoir 102a can hold deionized (DI) water, the second reservoir 102b can hold a density modifier such as sucrose, glycerol, or iodixanol, the third reservoir 102c can hold a buffer solution, and the fourth reservoir 102d can hold additives such as nucleic acids, amino acids, proteins, chelators, stabilizers, detergents, salts, virus particles, gene therapy vectors, lipid-based particles such as lipid nanoparticles (LNPs), liposomes, exosomes. extracellular vesicles, other biological nanoparticles, and biological sample material. Illustrative examples of the buffer solutions can include, without limitation, a phosphate-buffered saline (PBS), a tris buffer concentration (e.g., tris (hydroxymethyl) aminomethane, also known as tromethamine or THAM), and HEPES (4-(2-hydroxyethyl)-l -piperazineethanesulfonic acid).

[0054] All four gradient materials can be introduced into a single stream that goes through the mixing manifold 106. The DI water and density modifier make up a majority of the volume in the stream, while the buffer solution and additives have smaller concentrations.

[0055] As an illustrative example, the DI water and density modifier are pumped into the mixing manifold 106 from the respective first and second reservoirs 102a, 102b using the first and second pumps 104a, 104b, respectively. The first and second pumps 104a, 104b can include peristaltic pumps for providing a smooth pumping flow for theDI water and density modifier gradient materials. The buffer solution and additives from the respective third and fourth reservoirs 102c, 102d are pumped by the third and fourth pumps 104c, 104d, respectively. In some examples, the third and fourth pumps 104c, 104d include peristaltic pumps. In other examples, the third and fourth pumps 104c, 104d can include syringe pumps, which can be used when higher precision pumping is desirable for the buffer solution and additives.

[0056] FIG. 2 illustrates an example of a mixer 200 housed inside the mixing manifold 106 of the system 100. Referring now to FIGS. 1 and 2, the DI water, density modifier, buffer solution, and additives are introduced into a single stream that goes through the mixing manifold 106. Inside the mixing manifold 106, the mixer 200 includes mixing elements 202a-202f that mix the gradient materials together as they pass through the mixing elements. The mixer 200 mixes the gradient materials together to generate a homogenous stream of fluid for a probe 108 to dispense a density gradient into the container 110. The gradient materials are mixed together to have a predetermined density based on their relative concentrations.

[0057] In some examples, the mixer 200 is a static mixer and the mixing elements 202a-202f include alternating helical elements. In some examples, each helical element is set 90° to an adjacent helical element to provide thorough blending of the gradient materials over a length L of the mixer 200 inside the mixing manifold 106. The mixing elements 202a-202f mix the gradient materials together to eliminate pockets of low and / or high-density material. The mixing elements 202a-202f slice and rotate the DI water and density modifier multiple times together to produce a substantially homogenous stream for the probe 108 to dispense the density gradient into the container 110. In alternative examples, the mixing manifold 106 can include alternative types of mixers, mixing elements, and the like.

[0058] FIG. 3 is an isometric view of the probe 108 having a distal end 112 inserted into the container 110, and a proximal end 114 that connects to tubing of the mixing manifold 106. As shown in FIG. 3, the distal end 112 is positioned toward a bottom of an interior volume 122 of the container 110 such that the probe 108 is ready for dispensing a density gradient inside the interior volume of the container. In some examples, the probe 108 remains fixed in the same position when dispensing the density gradient inside the interior volume 122 of the container.

[0059] As show n in FIG. 3, the container 110 is fixedly positioned by a holder 116 relative to the probe 108 during dispensing of the density gradient. In the example of FIG. 3, the holder 116 includes a clamp for securely fixing the container 110 to a frame 118 of the system 100.

[0060] FIG. 4 illustrates an example of the proximal end 114 of the probe 108 connected to the mixing manifold 106. The mixing manifold 106 includes a manifold portion 402 having inputs 404a-404d that each receive a gradient material pumped from a reservoir 102a-102d by a pump 104a-104d, respectively. The mixing manifold 106 further includes a mixing portion 406 housing the mixer 200 for mixing the gradient materials pumped from the reservoirs together before they reach the proximal end 114 of the probe 108.

[0061] In FIG. 4, the proximal end 114 of the probe 108 is shown fixed by a set screw that can be tightened or loosened around the proximal end 114 by using a rotatable handle 410. The mixing manifold 106 is attached to a motor driven mechanism that moves the probe 108 up and down to a desired position inside the container 1 10. In other examples, the probe 108 can be manually lowered into a desired position inside the container 110.

[0062] The probe 108 is coated with a non-stick material. In some examples, the probe 108 is coated with anon-stick material such as Teflon, or similar materials. The non-stick material coating on the probe 108 minimizes the density gradient dispensed into the container 1 10 from sticking to or building-up on the probe 108. Thus, the nonstick material coating allows the probe 108 to be removed while mitigating mixture in the dispensed density gradient.

[0063] Referring back to FIG. 1, the system 100 can include a workstation 130 for receiving inputs from a user to generate one or more density gradients. In some examples, the workstation 130 includes a display 132 that can be used by the user to select and enter parameters that define one or more density gradients for generation by the system 100. In some examples, the display 132 operates as both an output and an input device such as a touchscreen display. The workstation 130 can also include additional input devices such as a keyboard and a mouse.

[0064] In some examples, the workstation 130 is separate from an instrument that houses the mixing manifold 106, the pumps 104, and / or the reservoirs 102. In such examples, the workstation 130 is communicatively connected to the instrument such asthrough a wired or wireless connection. Alternatively, the workstation 130 can be integrated with an instrument that houses the mixing manifold 106. the pumps 104, and / or the reservoirs 102.

[0065] FIG. 5 schematically illustrates an example of a method 500 of generating a density gradient inside the container 110 using the system 100. The method 500 includes an operation 502 of lowering the probe 108 close to a bottom of the container 110. An example of the probe 108 positioned close to the bottom of the container 110 is shown in FIGS. 1 and 3. In some examples, the distal end 112 of the probe 108 will remain positioned close to the bottom of the container 110 while the probe 108 dispenses the density gradient.

[0066] The method includes an operation 504 of dispensing into the container 110 a first step made of the gradient materials held in the reservoirs 102 (e.g., a first step of the DI water, density7modifier, buffer solution, and additives). The first step has a first density based on the relative concentrations of the gradient materials. For example, increasing an amount of the density modifier mixed by the mixing manifold 106 increases the density of the first step dispensed by the probe 108. while decreasing the amount of the density modifier mixed by the mixing manifold 106 decreases the density of the first step dispensed by the probe 108.

[0067] Next, the method 500 includes an operation 506 of dispensing into the container 110 a second step made of the gradient materials held in the reservoirs 102 (e g., a second step of the DI water, density modifier, buffer solution, and additives). The second step has a second density based on the relative concentrations of the gradient materials pumped from the reservoirs 102. The second density is heavier than the first density’ such that the second step pushes up the first step, and the second step remains below the first step at the bottom of the container 1 10.

[0068] Next, the method 500 includes an operation 508 of determining whether the density gradient includes an additional step. When the density' gradient includes an additional step (i.e.,L‘Yes” in operation 508), the method 500 repeats the operation 506 to dispense an additional step made of the gradient materials held in the reservoirs 102 (e g., an additional step of the DI water, density modifier, buffer solution, and additives). The additional step has a density’ based on the relative concentrations of the gradient materials that is heavier than the densities of the previously dispensed steps such that the additional step pushes up the previously dispensed steps and remainsbelow the previously dispensed steps. The operation 506 can be repeated based on the desired number of steps for the density gradient. Each time the operation 506 is performed, the probe 108 remains in the same position (i.e., close to the bottom of the container 110).

[0069] When the density gradient does not include an additional step (i.e., “No” in operation 508), the method 500 proceeds to an operation 510 of removing the probe 108 from the container 110. Operation 510 can include removing the probe 108 slowly to not disturb the steps of the density gradient. As discussed above, the probe 108 can be coated with a non-stick material to minimize the steps in the density gradient from sticking to the probe 108 during its removal.

[0070] In the method 500. each of the pumps 104a-104d is programmed to control the flow of each gradient material into the mixing manifold 106 to have a given volume and / or speed for generating each step of the density gradient. This allows the system 100 to precisely control the concentration of each gradient material in each step of the density gradient dispensed by the probe 108 into the container 110 for generating the density gradient.

[0071] FIG. 6 schematically illustrates an example of a density gradient 300 formed by the system 100 in the container 110 after completion of the method 500. The density gradient 300 is a medium created for the separation of particles in ultracentrifugation. In this example, the density gradient 300 is a step gradient, such that the density gradient 300 includes discrete steps having different densities. In this example, the density gradient includes five discrete steps. In alternative examples, the method 500 can be performed to form a continuous or linear gradient.

[0072] In the example of FIG. 6, the density gradient 300 includes steps 302a-302e. Each of the steps 302a-302e has a unique density based on the relative proportions of the gradient materials mixed in the mixing manifold 106. In this illustrative example, the density gradient 300 includes a first step 302a having a first density, a second step 302b having a second density, a third step 302c having a third density, a fourth step 302d having a fourth density, and a fifth step 302e having a fifth density. In accordance with the above description, the fifth density of the fifth step 302e is the heaviest and the first density of the first step 302a is the lightest, such that the densities of the steps 302a-302e increase from top to bottom inside the container 110. Each of the steps that form part of the density gradient 300 is shown separated by boundaries 310.

[0073] The system 100 and / or the method 500 can form density gradients having more than five separate steps, and / or to form density gradients having fewer than five separate steps. Also, the system 100 can form step gradients, such as the one shown in FIG. 6, and also continuous or linear gradients having layers of gradually increasing density from top to bottom. Thus, the density7gradient 300 is shown by way of illustrative example only.

[0074] As further shown in FIG. 6, the container 110 includes several features. For example, the container 110 includes a bottom portion 304, a cylindrical portion 312 that extends from the bottom portion, and an opening 314 that allows liquid to be dispensed into an interior volume of the container 110. The container 110 includes a centerline CL that runs down the middle of the container 110. In some examples, the container 110 can include a seam between the bottom portion 304 of the container 110 and the cylindrical portion 312 of the container 110. In some examples, the boundary7between the bottom portion 304 and the cylindrical portion 312 is used to establish an origin for positioning a sensor to start measuring the density gradient 300.

[0075] In FIG. 6, the density gradient 300 exhibits characteristics such as a meniscus 308, which is located at the top of the density gradient 300. The meniscus 308 is caused by surface tension between the density7gradient 300 and an interior surface of the walls of the container 110. The meniscus 308 includes a bottom edge 309 where the centerline CL of the container 110 is located. The meniscus 308 further includes top edges 31 1 near the walls of the container 110. The meniscus 308 can interfere with measurements of the density7gradient 300 such that measurements are not obtained or are filtered from this portion of the density7gradient 300.

[0076] The container 110 includes a volume 306 above the meniscus 308 of the density gradient 300 and below the opening 314. The volume 306 can be filled with air or an inert gas. Measurement data can be obtained from the volume 306 to determine a ty pe of material from which the container 110 is made. As an illustrative example, the container 110 can be made from polypropylene, polycarbonate, co-polyester resins such as polyethylene terephthalate glycol (PETG), and other materials. Each type of material can exhibit unique characteristics when light is transmitted through an empty portion of the container (e.g., the volume 306). The measurement data from the volume 306 can be used to standardize the measurements of the density gradient 300 for different types of containers made from different types of materials.

[0077] As further show n in FIG. 6, the container 110 has a length L that extends from the bottom portion 304 to the opening 314 of the container. The density gradient 300 has a length LD. In this illustrative example, the length LD of the density gradient 300 is less than the length L of the container 110 such that the density gradient 300 occupies a portion of the length of the container. In some examples, the length LD of the density gradient 300 can be about 80 mm.

[0078] The system 100 can perform a non-destructive density gradient measurement over the length L of the container 110. The density gradient measurement can be used to verify that the densify gradient 300 conforms to a desired profile or meets a desired qualify control. Additionally, the densify gradient measurement can be stored in a memory for the system 100 or another system to replicate the densify gradient 300 inside another container.

[0079] FIG. 7 is an isometric view of an example of a measurement apparatus 700 in the system 100 for performing a non-destructive densify gradient measurement over the length L of the container 110. For example, the measurement apparatus 700 can measure the densify gradient 300 without touching or disturbing the densify gradient. FIG. 8 is a detailed isometric view of the measurement apparatus 700. FIG. 9 is a front view of the measurement apparatus 700.

[0080] Referring now to FIGS. 7-9, in this example, the measurement apparatus 700 includes a platform 702 that supports a frame 704 that includes rails 706. In some examples, the rails 706 include a screw rail. The measurement apparatus 700 further includes a sensor assembly 712 mounted on a carriage 708 powered by a motor 710 to move up and down the rails 706 while the container 110 remains in a fixed position. In some examples, the motor 710 is a step motor or other similar type of electric motor. The rails 706 and the motor 710 provide precise vertical movement of the sensor assembly 712 relative to the container 110. The container 110 can be held relative to the measurement apparatus 700 by a holder (see FIG. 27) such as a clamp. Also, in some examples, the rails 706 and the motor 710 can be used to insert the probe 108 into the container 110, and can also be used to remove the probe 108 from the container 110.

[0081] Alternative examples for moving the sensor assembly 712 and / or the probe 108 along the length L of the container 110 and / or the length LD of a density gradient are possible. For example, gantry and pully system could be used to move the sensorassembly 712 and / or the probe 108 along the length L of the container 110 and / or the length LD of a density gradient. Additional structures for moving the sensor assembly 712 and / or the probe 108 along the length L of the container 110 and / or the length LD of a density gradient are contemplated such that the structure show n in FIGS. 7-9 is provided by way of illustrative example.

[0082] In some examples, the sensor assembly 712 is used for measuring density across the length LD of the density gradient 300 dispensed in the container 110. As shown in FIGS. 8 and 9, the sensor assembly 712 includes an emitter 714 that emits a signal such as light, and a detector 716 that measures the signal from the emitter 714 after transmission through the density gradient dispensed in the container 110. The emitter 714 and the detector 716 are fixed in relationship to each other when mounted on the carriage 708, which allows these components of the sensor assembly 712 to be moved up and down the rails 706 together. The carriage 708 allows coordinated movement of the emitter 714 and the detector 716, and provides a precise alignment of the emitter 714, the detector 716, and the centerline CL of the container 110.

[0083] In the example illustrated in FIGS. 7-9, the emitter 714 is mounted on one side of the carriage 708, and the detector 716 is mounted on an opposite side of the carriage 708. The container 110 with a density gradient dispensed therein is fixedly positioned relative to a center axis of the carriage 708 such that the signal (e.g., light) emitted by the emitter 714 passes through the container 110, and is received by the detector 71 on an opposite side of the container 110.

[0084] In alternative examples, the emitter 714 and detector 716 can be mounted on the same side of the carriage 708. For example, the emitter 714 can emit the signal from a first side of the camage 708 that passes through the density gradient dispensed in the container 110 and that is reflected by a mirror mounted on a second side of the carriage 708 for reflection back tow ard the first side of the carriage 708 where the detector 716 is mounted together with the detector 716. Further alternative arrangements for the sensor assembly 712 are contemplated.

[0085] In one example embodiment, the emitter 714 emits light, and the detector 716 includes a photodiode that detects a current that results from the transmission of the light through the container 110. In some examples, the emitter 714 emits light within the infrared spectrum (e.g., light having a wavelength of about 700 nm to about 1000 nm). In some further examples, the emitter 714 emits light having a wavelengthof about 880 nm. In alternative examples, the emitter 714 emits light within the visible spectrum (e.g.. from about 380 nm to about 750 nm).

[0086] A current is generated on the detector 716 when the light from the emitter 714 that passes through the container 110 strikes the detector 716. The sensor assembly 712 can further includes an amplifier circuit that converts the current into a voltage. Thus, the sensor assembly 712 measures and records voltages at multiple points along the length L of the container 1 10 for measuring a density gradient dispensed in the container 110. As an example, the sensor assembly 712 can measure and record voltages at 320 points over a length of about 80 mm.

[0087] The voltage measurements recorded by the sensor assembly 712 correlate to refractive indices along the length of a density’ gradient dispensed in the container 110, and can be used to compute densities along the length of the density gradient. This is because density affects the transmission of the light from the emitter 714 through the density gradient. Thus, the voltage measurements recorded by the sensor assembly 712 can be used to measure density’ values at given points along the length of the density gradient dispensed in the container 110.

[0088] As shown in FIG. 8, the carriage 708 includes a slot 718 that is positioned in front of the detector 716. The slot 718 focuses light emitted from the emitter 714 that passes through a narrow slice of the density gradient dispensed in the container 110. The slot 718 allows the measurement apparatus 700 to measure narrow slices along the length of the density gradient dispensed in the container 1 10. Additionally, multiple measurements can be taken for each slice of the density’ gradient (e.g., 100 measurements per slice), and the measurements can be averaged to reduce variation in the measurements made along the entire length of the density gradient.

[0089] To further reduce sensitivity due to positional errors between the container 110, the emitter 714, and the detector 716, the detector 716 is provided with a large surface area. In some examples, the detector 716 includes a photodiode having a surface area of about 8.5 mm2.

[0090] FIG. 10 schematically illustrates an example of an electrical configuration for the sensor assembly 712. As shown in FIG. 10, a resistor 720 sets the current for the emitter 714. As an illustrative example, the resistor 720 can set the current for the emitter 714 to be in a range of about 19 mA to about 33mA. In some examples, the resistor 720 can have an electrical resistance of about 90 . The emitter 714 can bepowered by a stable, high precision DC power supply to maintain a steady level of infrared (IR) radiance. In an alternative example, the DC power supply for the emitter 714 can be replaced with a constant current source, which can reduce variation due to power supply drift. Additional examples for powering the emitter 714 are possible.

[0091] A transimpedance amplifier design is used to convert a detected current (Id) of the detector 716 into a voltage across a feedback resistor 722. In some examples, the feedback resistor 722 has an electrical resistance of about 47kQ. An operational amplifier 724 having precision input current is used for its ability to operate with low current. A second DC power supply can provide + / - 6VDC for the operational amplifier 724.

[0092] When the density gradient is dispensed in the container 110, the container becomes a cylindrical lens such that the spacing between the emitter 714, the container 110, and the detector 716 can affect the voltage measurements obtained from the sensor assembly 712. For example, a spacing of about 1.10 inches (28 mm) between the emitter 714 and the detector 716 can be used for a container having a 9 / 16 inch diameter, and a spacing of about 2.44 inches (62mm) between the emitter 714 and the detector 716 can be used for a container having a 1 inch diameter.

[0093] The system 100 is programmable to dispense a plurality of density gradients into a number of containers 110 as a set defined by the user. The process of dispensing the plurality of density gradients typically requires dispensing greatly different concentrations of gradient materials in succession. The system 100 is programmable to dispense the set of density gradients using a single probe such that the system 100 does not replace the probe between steps of a density gradient or between different density gradients in the set of density gradients. Purging the gradient materials from the mixing manifold 106 and the probe 108 into a waste container between steps of a density gradient or each time a new density gradient is generated in the set of density’ gradients can lead to undesirable waste of the gradient materials and downtime, which can reduce efficiency and lead to increased operating costs of the system 100.

[0094] FIG. 11 schematically illustrates an example of a method 1100 of dispensing a plurality of density gradients into a number of containers 110. In some examples, the method 1100 can be performed by the system 100. As will be described in more detail, the method 1100 can be performed by the system 100 to reduce wastewhen dispensing a set of density gradients into separate containers as part of a single density gradient dispensing process.

[0095] As shown in FIG. 11 , the method 1100 includes an operation 1102 of generating a dispense project. The dispense project is generated based on parameters that are selected and entered by a user via the workstation 130 and the display 132 shown in FIG. 1.

[0096] FIGS. 12-17 illustrate examples of graphical user interfaces that can be displayed on the display 132 of the workstation 130. The graphical user interfaces of FIGS. 12-17 display parameters that can be selected and entered by the user of the system 100 to define the dispense project. FIG. 12 illustrates a graphical user interface 1200 that includes a first selectable option 1202 that when selected allows the user to enter in a field 1206 a quantity of containers for filling with predetermined density gradients dispensed by the system 100. For example, the user can select any quantity of containers between one container and twelve containers.

[0097] The graphical user interface 1200 includes a second selectable option 1204 that when selected allows the user to enter in a field 1208 a volume of sample, and the system 100 calculates a quantity7of containers for filling with the predetermined density gradients based on the volume of sample. As an illustrative example, the system 100 can calculate any quantity of containers between one container and twelve containers.

[0098] The graphical user interface 1200 further displays an area 1210 that identifies the gradient materials such as a sample of adeno-associated virus (AAV) held in the fourth reservoir 102d, a PBS buffer solution held in the third reservoir 102c, an iodixanol density modifier held in the second reservoir 102b, and DI water held in the first reservoir 102a. The area 1210 can further display a volume of each gradient material that is to be dispensed in each container, and a total run volume of each gradient material. Once the user has selected the desired parameters on the graphical user interface 1200, the user can select a “Next” icon 1212 to enter the parameters. Under either the first selectable option 1202 or the second selectable option 1204, the dispense project is defined by the quantity of containers for filling with density gradients dispensed by the system 100, and by the volume of each gradient material that is dispensed in each container and the total run volume of each gradient material displayed in the area 1210.

[0099] FIG. 13 illustrates an example of a graphical user interface 1300 displaying a plurality of density gradients 1302 that can be selected by the user for automatic generation by the system 100. Also, each density gradient in the plurality of density gradients 1302 is selectable to view and adjust one or more parameters that define the density gradient.

[0100] In FIG. 13, a density gradient titled “‘AAV Gradient 4” is shown as selected, and parameters such as type of container, number of sections or steps, gradient materials used, and whether the container is a heat seal tube or not are displayed. Also, a window 1304 allows a user to adjust a quantity of containers to be filled by the system 100 with the selected density gradient (e.g., “AAV Gradient 4”). When the user is satisfied with the parameters for a given density gradient, the user can select a "Run” icon 1306. This causes the dispense project to be generated, and the system 100 automatically generates the selected density gradient into a number of containers 110 set by the user without requiring any further input from the user.

[0101] FIG. 14 illustrates an example of a graphical user interface 1400 that allows a user to edit the density gradient selected in the graphical user interface 1300 (e.g., “AAV Gradient 4”), and thereby edit the dispense project. In this example, the graphical user interface 1400 allows the user to select one or more characteristics of the containers 110 to be filled with the selected density gradient. The graphical user interface 1400 includes a first section 1402 to select a shape of the containers 110 (e.g., “Quick-Seal”, “Open-Top”, or “Optiseal”); a second section 1404 to select a material of the containers 110 (e.g., “Ultra-Clear” or “Polypropylene”); a third section 1406 to identity' a volume of the containers 110; a fourth section 1408 to select an additional parameter (e.g., “None”. “Sterile”, or “Certified Free”); and a fifth section 1410 to select whether the containers 110 are heat seal tubes. Also, the graphical user interface 1400 includes an option 1412 to select a quantity of containers to be filled with the selected density7gradient.

[0102] FIGS. 15 and 16 illustrate examples of a graphical user interface 1500 that includes drop down menus 1502a-1502d each allowing a user to identify the reagents pumped from the reservoirs 102a-102d by the pumps 104a-104d. For example, as shown in FIG. 16, when the user selects the drop down menu 1502a for “Pump 1”, a plurality of reagents 1504 are displayed for selection by the user. The plurality of reagents 1504 that can be selected for “Pump 1” can include DI water, one or moretypes of buffer solutions (e.g., phosphate-buffered saline (PBS)), one or more types of density modifiers (e.g., caesium chloride (CsCl), iodixanol) and one or more types of sample material (e.g., ‘'AAV Sample Type 1”, “AAV Sample Type 2”).

[0103] FIG. 17 illustrates an example of a graphical user interface 1700 that allows the user to define each section or step of the selected density gradient. In this example, a first section 1702 toward the top of the container (e.g., “Section 1”) is defined by target volumes for gradient materials within a total section volume of 10 mL. The first section 1702 includes the following gradient materials: DI water, a density modifier (e.g., CsCl), a buffer solution, and sample.

[0104] A second section 1704, which is below the first section, is defined as a continuous section that includes a first set of target concentrations 1706 at a top of the second section, and a second set of target concentrations 1708 at a bottom of the second section. The first and second sets of target concentrations 1706, 1708 include a density modifier (e.g., CsCl) and a buffer solution, and the second section 1704 is defined as having a total section volume of 29 mL.

[0105] Each of the foregoing parameters of the first and second sections 1702. 1704 such as selected gradient materials and their associated target concentration can be modified to adjust the density7gradient. Additionally, the user can select an “Add Section’" icon 1710 to define an additional section to add to the density gradient. The graphical user interface 1700 includes up and down arrows 1714. 1716 to switch between the different sections of the density’ gradient. Once the user is satisfied with the sections of the density gradient and their associated parameters, the user can select the “Save’" icon 1712 to save the density’ gradient. Once the density gradient is saved, the system 100 generated the dispense project based on the parameters of the densitygradient, w hether or not a sample is to be overtaxed on top of the density gradient, and a quantity of containers that are to be filled w ith the density’ gradient.

[0106] Referring back to FIG. 11, after the dispense project is generated, the method 1100 includes an operation 1104 of calculating a dispense queue. The dispense queue is calculated based on the dispense project, which as described above, is generated based on the parameters of the density gradient, whether or not a sample is to be overlayed on top of the density’ gradient, and a quantity of containers that are to be filled w ith the density gradient. For example, the dispense queue is calculated based onat least the volumes of gradient materials that are needed to complete each section of the density gradient in each container 110.

[0107] FIG. 18 is a cross-sectional view showing internal volumes of the system 100 that are used for calculating the dispense queue in operation 1104 of the method 1100. As shown in FIG. 18, a first volume VI is defined by an internal mixing chamber 124 of the mixing manifold 106. a second volume V2 is defined by tubing 126 that connects the probe 108 to the mixing manifold 106. and a third volume V3 is defined by the probe 108. The gradient materials are pumped into the internal mixing chamber 124 where they are mixed together, and leave the internal mixing chamber 124 via the tubing 126 which is connected to the proximal end of the probe 108. The gradient materials exit the distal end of the probe for dispense inside a container 110. The system 100 considers each of the first, second, and third volumes VI, V2, V3 when calculating the dispense queue in operation 1104 as will be described further below. An illustrative example of a dispense queue 2400 is describe further below with reference to FIG. 24.

[0108] Referring back to FIG. 11. the method 1100 includes an operation 1106 of dispensing the density gradient in a first container based on the dispense queue. Operation 1106 can be performed as described above with respect to the method 500 illustrated in FIG. 5. Operation 1106 can include dispensing successively denser steps or sections that push up previously dispensed steps or sections while the probe 108 remains at the bottom of a container 1 10.

[0109] FIG. 19 illustrates an example of a density gradient 1900 generated inside a container 110 during performance of operation 1106. As shown in FIG. 19, the probe 108 remains at the bottom of the container 110 during performance of operation 1106. In FIG. 19, the density gradient 1900 is illustrated as a step gradient having steps 1902a-1902d. In other examples, the density gradient generated in operation 1106 can include a continuous or linear gradient. Also, FIG. 19 illustrates the container 110 as a quick-seal tube for illustration purposes. Additional types of containers and tubes can be used for dispensing the density gradient.

[0110] The method 1100 includes an operation 1108 of determining whether the dispense project requires a sample to be overlay ed on top of the density' gradient dispensed in the first container. When a sample is to be overlayed on top of the density gradient (i.e. , "Yes’' in operation 1108), the method 1100 proceeds to an operation 1110of purging the mixture of gradient materials from the probe 108 by displacing the mixture of gradient materials with the sample. As described above, the density gradient is dispensed in operation 1 106 by dispensing successively denser steps that push up previously dispensed steps while the probe 108 remains at the bottom of the container. Thus, before the density gradient is completed, the volume of at least the probe 108 (see volume V3 in FIG. 18) is filled with a mixture of gradient materials used for generating the last step or section of the density gradient. This mixture of gradient materials will have a higher density than the previously dispensed steps or sections of the density gradient.[OHl] The dispense queue is calculated such that before the last step or section of the density gradient is completed, the dispense queue includes a predetermined volume of sample that is pumped into the internal mixing chamber 124 and through the tubing 126 to displace the mixture of gradient materials used for dispensing last step or section of the density7gradient from the probe 108 while the probe 108 remains at the bottom of the container 110. As an example, the system 100 can pump about 0.1 - 0.2 mL of sample to purge from the probe 108 the mixture of gradient materials used for dispensing last step or section of the density7gradient.

[0112] The sample has a lower density7than the mixture of gradient materials used for dispensing last step or section of the density gradient. The sample at least partially fills the volume of the probe 108 (i.e.. the third volume V3 in FIG. 18). The sample does not exit the probe 108 due to differential pressure inside the probe 108.

[0113] In view of the foregoing, operation 1110 includes pumping the sample into the probe 108 before the density gradient is completed to prepare the probe 108 for dispensing the sample after completion of the density gradient. Advantageously, this eliminates the need to purge the gradient materials from the mixing manifold 106, the tubing 126, and the probe 108 into a waste bin or container, which saves resources such as the gradient materials. This also reduces or eliminates downtime which would ty pi cal ly be required to purge the gradient materials into the waste bin or container before the sample can be overlay ed on top of the density gradient. Also, operation 1110 allows the first volume VI of the internal mixing chamber 124, the second volume V2 of the tubing 126, and the third volume V3 of the probe 108 to be continuously filled with liquid, and thereby prevents air bubbles from forming inside the probe 108. Also,operation 1110 eliminates the need to prime the probe 108 for dispensing the sample, which can further reduce downtime and improve efficiency of the system 100.

[0114] Referring back to FIG. 11, when a sample is not required to be overlayed on top of the density gradient (i.e., “No” in operation 1108), the method 1100 proceeds to an operation 1112 of purging from the probe 108 the mixture of gradient materials used for dispensing the last step or section of the density gradient using a mixture of gradient materials used for dispensing the first step or section of the next density gradient in the dispense project. Operation 1112 is similar to operation 1 110. In operation 1112, the dispense queue is calculated such that before the last step or section of the density gradient is completed, the dispense queue includes a volume of the mixture of gradient materials used for dispensing the first step or section of the next density gradient for pumping into the internal mixing chamber 124 and through the tubing 126 to displace from the probe 108 the mixture of gradient materials used for dispensing last step or section of the density gradient while the probe 108 remains at the bottom of the container 110.

[0115] The mixture of gradient materials used for dispensing the first step or section of the next density gradient has a lower density than the mixture of gradient materials used for dispensing last step or section of the current density gradient. The mixture of gradient materials used for dispensing the first step or section of the next density gradient fills the volume of the probe 108 (i.e., the third volume V3 in FIG. 18) to displace the mixture of gradient materials used for dispensing last step or section of the current density gradient. The mixture of gradient materials used for dispensing the first step or section of the next density gradient does not exit the probe 108 due to differential pressure inside the probe 108.

[0116] Operation 11 12 includes pumping the mixture of gradient materials used for dispensing the first step or section of the next density gradient into the probe 108 before the density gradient is completed to prepare the probe 108 for dispensing the next density gradient before completion of the current density gradient. Advantageously, this eliminates the need to purge the gradient materials from the mixing manifold 106. the tubing 126, and the probe 108 into a waste bin or container, which saves resources such as the gradient materials. Also, this reduces or eliminates downtime which would typically be required to purge the gradient materials into the waste bin or container before dispensing of the next density gradient can begin. Also, operation 1112 allowsthe first volume VI of the internal mixing chamber 124, the second volume V2 of the tubing 126, and the third volume V3 of the probe 108 to be continuously filled with liquid, and thereby prevents air bubbles from forming inside the probe 108. Operation 1112 also eliminates the need to prime the probe 108 for dispensing the first step or section of the next density gradient, which can further reduce downtime and improve efficiency of the system 100.

[0117] After completion of operation 1110 or operation 1 112, the method 1100 includes an operation 11 14 of removing the probe 108 from the density gradient. In examples when the dispense project includes overlaying sample on top of the density gradient, operation 1114 includes positioning the probe 108 above the meniscus of the density gradient. Alternatively, when dispense project does not include overlaying sample on top of the density gradient, operation 1 114 includes removing the probe 108 from the container such that the probe can be positioned inside another container for dispensing a next density gradient in the dispense project.

[0118] In addition to operations 1110.1112. the method 1100 can also include purging the probe 108 between sections of the same density gradient. For example, operation 1106 of dispensing the density gradient in the first container based on the dispense queue can include a sub-operation of purging the probe 108 by displacing a first mixture of gradient materials used for dispensing a first section of the first density gradient with a second mixture of gradient materials used for dispensing a second section of the first density gradient.

[0119] FIG. 20 illustrates an example of when the dispense project includes overlaying sample on top of the density gradient. Referring now' to FIGS. 19 and 20, after the mixture of gradient materials for dispensing the last step 1902d of the density gradient 1900 is purged from the probe 108, the probe 108 is raised above the density gradient 1900 such that the probe 108 can be used to overlay the sample on top of the density gradient. The sensor assembly 712 can detect the location of the meniscus of the density gradient 1900 dispensed in the container 110, and the rails 706 and the motor 710 shown in FIGS. 7-9, or other mechanisms such as a gantry and pully system powered by the motor 710 can be used to raise the probe 108 such that the distal end of the probe 108 is moved from the bottom of the container 110 to a position above the meniscus of the density gradient 1900, as show n in FIG. 20. When in this position, the probe 108 is ready to overlay the sample on top of the density gradient.

[0120] FIG. 21 schematically illustrates an example of a method 2100 of dispensing the sample after the density gradient 1900 is dispensed in accordance with the method 1100. The method 2100 includes an operation 2102 of dispensing the sample on top of the density gradient. During operation 2102, the sample is dispensed slowly to not disturb the density gradient below.

[0121] The method 2100 further includes an operation 2104 of raising the probe 108. The operations 2102. 2104 can occur simultaneously such that the probe 108 is raised while dispensing the sample. As described above, the rails 706 and the motor 710 shown in FIGS. 7-9, or other mechanisms such as a gantry and pully system powered by the motor 710 can be used to raise the probe 108 while the probe dispenses the sample on top of the density gradient.

[0122] FIGS. 22 and 23 illustrate the probe 108 dispensing a sample 1916 on top of the density gradient 1900. As shown in FIGS. 22 and 23, the probe 108 is raised while dispensing the sample 1916 until the probe 108 reaches the top of the container 110.

[0123] Referring back to FIG. 21, the method 2100 includes an operation of determining whether a threshold volume of sample is dispensed above the density gradient 1900. The threshold volume is determined from the dispense project, which as described above, is generated based on the parameters selected and entered by the user of the system 100 such as through the graphical user interfaces illustrated in FIGS. 12- 17. When the threshold volume is not reached (i.e., “No” in operation 2106). the method 2100 continues to perform operations 2102-2106. Otherwise, when the threshold volume is reached (i.e., “Yes” in operation 2106), the method 2100 proceeds to an operation 2108 of purging the sample from the probe 108 using DI water or the mixture of gradient materials used for dispensing the first step or section of the next density gradient. Thus, before the dispense of the sample is completed, the volume of at least the probe 108 (see volume V3 in FIG. 18) is filled with DI water or the mixture of gradient materials used for dispensing the first step or section of the next densitygradient.

[0124] The dispense queue is calculated such that before the dispense of the sample is completed, the dispense queue includes a predetermined volume of DI w ater or the mixture of gradient materials used for dispensing the first step or section of the next density gradient. The predetermined volume of DI water or the mixture of gradient materials is pumped into the internal mixing chamber 124 and through the tubing 126to purge from the probe 108 the sample while the probe 108 is raised inside the container 110. The DI water or the mixture of gradient materials used for dispensing the first step or section of the next density gradient do not exit the probe 108 due to differential pressure inside the probe 108.

[0125] In instances when the DI water is used to purge the sample from the probe 108, the DI water is dispensed to top-off the container 110 such as when the container 110 is a quick-seal tube that needs to be filled to the neck for sealing, as is shown in the illustrative example provided in FIG. 23. In such examples, the dispense queue can further include the mixture of gradient materials used for dispensing the first step or section of the next density gradient to purge the DI water from the probe 108 before the container 110 is filled to the neck.

[0126] The method 2100 includes pumping the DI water or the mixture of gradient materials for dispensing the first step or section of the next density gradient into the probe 108 before the dispense of the sample is completed. Advantageously, this eliminates the need to purge the sample from the mixing manifold 106. the tubing 126, and the probe 108 into a waste bin or container, which means that the dispense project can be completed with less sample because the sample is not wasted. This also reduces or eliminates downtime that is typically required to purge the sample into the w aste bin or container before dispensing of the next density gradient begins. Also, this allows the first volume VI of the internal mixing chamber 124, the second volume V2 of the tubing 126, and the third volume V3 of the probe 108 to be continuously filled with liquid, and thereby prevents air bubbles from forming inside the probe 108. This also eliminates the need to prime the probe 108 for dispensing the first step or section of the next density gradient, which can further reduce downtime and improve efficiency of the system 100.

[0127] FIG. 24 schematically illustrates an example of a dispense queue 2400 that can be calculated by the system 100, such as during operation 1104 of the method 1100. The dispense queue 2400 includes a first dispense volume DV 1 for a first liquid 2402, a second dispense volume DV2 for a second liquid 2404. a third dispense volume DV3 for a third liquid 2406, and so on. It is noted that the dispense queue 2400 can contain additional dispense volumes DV, such that the dispense volumes shown in FIG. 24 are provided for illustrative purposes only.

[0128] In FIG. 24, the first dispense volume DV1, the second dispense volume DV2, and the third dispense volume DV3 are displayed relative to the first volume VI of the internal mixing chamber 124, the second volume V2 of the tubing 126, and the third volume V3 of the probe 108. The first dispense volume DV1, the second dispense volume DV2, and the third dispense volume DV3 do not have to necessarily align with the first, second, and third volumes VI, V2, V3. Instead, the system 100 calculates the first dispense volume DV1, the second dispense volume DV2, and the third dispense volume DV3 based on the dispense project. For example, the system 100 calculates the first dispense volume DV1, the second dispense volume DV2, and the third dispense volume DV3 based on the parameters of the density gradient such as the volumes and the relative compositions of each gradient material in each step or section of the density gradient, whether or not a sample is to be overlayed on top of the density gradient, and a quantity of containers for filling with the density gradient.

[0129] As an illustrative example, the first liquid 2402 is a mixture of gradient materials for dispensing a last step or section of a current density gradient, the second liquid 2404 is a sample of particles or cells, and the third liquid 2406 is a mixture of gradient materials for dispensing a first step or section of a next density gradient. In this example, the second dispense volume DV2 is calculated to purge the first liquid 2402 from the probe 108 while the probe 108 remains at the bottom of a container 110, and to then dispense the sample on top of the density gradient when the probe is positioned above the meniscus of the density gradient. Further, the third dispense volume DV3 is calculated to purge the second liquid 2404 from the probe 108 while the probe 108 is positioned above the density gradient such that the probe 108 is ready to dispense the next density gradient in another container according to the dispense project.

[0130] FIG. 25 schematically illustrates an example of computing hardware 2500 of the system 100 for implementing aspects of the present disclosure. The computing hardware 2500 includes a processing circuitry having a memory for storing instructions which, when executed by the processing circuitry, cause the processing circuitry to perform the functions described herein. For example, as shown in the example provided in FIG. 25, the system 100 includes one or more processing devices 2502, a memory storage device 2504, and a system bus 2506 that couples the memory storage device 2504 to the one or more processing devices 2502. The one or more processing devices 2502 can include central processing units (CPU).

[0131] As further shown in FIG. 25. the memory storage device 2504 can include a random-access memory ("RAM") 2508 and a read-only memory ("ROM") 2510. Basic input and output logic having basic routines that help to transfer information between elements within the system 100, such as during startup, can be stored in the ROM 2510.

[0132] The system 100 can also include a mass storage device 2512 that can include an operating system 2514 and store software instructions 2516 and data. The mass storage device 2512 is connected to the processing device 2502 through the system bus 2506. The mass storage device 2512 and associated computer-readable data storage media provide non-volatile, non-transitory storage for the system 100.

[0133] Although the description of computer-readable data storage media contained herein refers to the mass storage device 2512. it should be appreciated by those skilled in the art that computer-readable data storage media can be any available non- transitory', physical device or article of manufacture from which the system 100 can read data and / or instructions. The computer-readable storage media can be comprised of entirely non-transitory’ media. The mass storage device 2512 is an example of a computer-readable storage device.

[0134] Computer-readable data storage media include volatile and non-volatile, removable and non-removable media implemented in any method or technology7for storage of information such as computer-readable softw are instructions, data structures, program modules or other data. Example types of computer-readable data storage media include, but are not limited to, RAM, ROM, EPROM, EEPROM, flash memory or other solid-state memory7technology7, or any other medium w7hich can be used to store information, and which can be accessed by the device.

[0135] The system 100 can operate in a networked environment using logical connections to the other devices through the communications network 2520. The system 100 connects to the communications netw ork 2520 through a network interface unit 2518 connected to the system bus 2506. The netw ork interface unit 2518 can connect to additional types of communications networks and devices, including through Bluetooth, Wi-Fi, and cellular telecommunications networks including 4G and 5G networks. The network interface unit 2518 can connect the system 100 to additional networks, systems, and devices. The system 100 also includes an input / output unit 2522 for receiving and processing inputs and outputs from peripheral devices.

[0136] The mass storage device 2512 and the RAM 2508 store software instructions and data. The software instructions include an operating system 2514 for operating the system 100. The mass storage device 2512 and / or the RAM 2508 store software instructions 2516, which when executed by the processing device 2502, provide the functionality of the system 100 discussed herein. The mass storage device 2512 and / or the RAM 2508 can store the profile of the density gradient measured by the measurement apparatus 700. as described above.

[0137] FIG. 26 schematically illustrates an example of a method 2600 of improving liquid level sensing after dispensing a density gradient in accordance with the method 1100 of FIG. 11 , or alternatively, after dispensing a sample on top of a density gradient in accordance with the method 2100 of FIG. 21. The method 2700 can be performed by the system 100. Liquid level sensing is a technology used to measure the height of a liquid within the container 110. Liquid level sensing uses capacitive sensors that measure changes in capacitance caused by the presence of the liquid when the liquid touches the distal end of the probe 108.

[0138] The liquid level sensing can be performed by the system 100 to calibrate the pumps 104 and to verify dispense volumes. The liquid level sensing uses the probe 108 to locate the surface height of the dispensed liquid within the container 110. The liquid level sensing is typically performed after a volume of liquid has been dispensed through the probe 108. either with the probe 108 fully submerged (e.g.. such as when performing the underlay technique) or with the probe 108 just over the surface of the liquid (e.g., such as when performing the overlay technique). During either the underlay or overlay techniques, droplets can form on the distal end of the probe 108 which can interfere with the accuracy of the liquid level measurement.

[0139] It is desirable for the system 100 to be able to reliably calibrate and to verify volumes of liquids dispensed within the container 110. Additionally, it is desirable to mitigate air bubbles in the liquid dispensed within the container 110, especially before centrifugation. However, the droplets at the distal end of the probe 108 interfere with the liquid level sensing due to the droplets triggering a measurement before the distal end of the probe 108 reaches the meniscus of the liquid. The method 2600 is directed to removing the droplets from the distal end of the probe 108 to improve the accuracy of the liquid level measurements captured by the system 100.

[0140] As shown in FIG. 26, the method 2600 includes an operation 2602 of lowering the probe 108 to contact the meniscus of the liquid dispensed in the container 110. In some examples, the meniscus of the liquid includes the density gradient dispensed in accordance with the method 1100 of FIG. 11. Alternatively, the meniscus of the liquid includes the sample dispensed on top of the density gradient in accordance with the method 2100 of FIG. 21. Operation 2602 causes the distal end of the probe 108 to contact the meniscus of the liquid to deposit the droplets formed on the probe 108 back into the liquid within the container 110.

[0141] The method 2600 includes an operation 2604 of raising the probe 108 after contacting the meniscus of the liquid in operation 2602. Then, the method 2600 includes an operation 2606 of lowering the probe 108 to contact the meniscus of the liquid dispensed in the container 110 to obtain a first liquid level measurement. Operation 2606 can occur following a pause after operation 2604 to allow a ripple caused by the initial contact between the probe 108 and the meniscus to dissipate. In some examples, the pause is about 3 seconds.

[0142] The method 2600 includes an operation 2608 of raising the probe 108 after contacting the meniscus of the liquid in operation 2606. Then, the method 2600 includes an operation 2610 of lowering the probe 108 to contact the meniscus of the liquid dispensed in the container 110 to obtain a second liquid level measurement. Operation 2610 occurs following a pause after operation 2608 to allow a ripple caused by the contact between the probe 108 and the meniscus to dissipate. In some examples, the pause is about 3 seconds.

[0143] The method includes an operation 2612 of determining whether the second liquid level measurement is within a predefined tolerance relative to the first liquid level measurement. For example, operation 2612 can include calculating an average of the first and second liquid level measurements, and then determining whether the second liquid level measurement is within a predefined tolerance of the average. In some examples, the operation 2612 includes determining whether the second liquid level measurement is within 0. 15 mm of the average of the first and second liquid level measurements.

[0144] When the second liquid level measurement is not within the predefined tolerance relative to the first liquid level measurement (i.e. , “No’' in operation 2612), the method 2600 repeats the operations 2608-2612 until at least two liquid levelmeasurements are within the predefined tolerance relative to each other. In some examples, operations 2608-2612 are repeated up to six times. When the method is unable to detect at least two liquid level measurements that are within the predefined tolerance relative to each other, the method 2600 includes displaying an error message on the display 132, and thereafter the method 2600 terminates.

[0145] In some alternative examples, operation 2612 can include determining whether at least three liquid level measurements are within a predefined tolerance relative to each other. In some examples, operation 2612 can include determining whether first, second, and third liquid level measurements are each within a predefined tolerance of an average of the three liquid level measurements. In some examples, operation 2612 can include determining whether the first, second, and third liquid level measurements are each within 0.07 mm of the average of the three liquid level measurements. The increased quantity of liquid level measurements that must be within the lower predefined tolerance can be performed during calibration of the system 100.

[0146] When the second liquid level measurement is within the predefined tolerance relative to the first liquid level measurement, or alternatively, the first, second, and third liquid level measurements are each within the predefined tolerance of the average of the three liquid level measurements (i.e. , “Yes” in operation 2612), the method 2600 proceeds to an operation 2614 of outputting a liquid level measurement. In some examples, operation 2614 includes outputting the average of the first and second liquid level measurements (or the first, second, and third liquid level measurements). Alternatively, operation 2614 can include outputting the last liquid level measurement such as the second liquid level measurement when two liquid level measurements are determined to be within the predefined tolerance, or the third liquid level measurement when three liquid level measurements are determined to be within the predefined tolerance.

[0147] FIG. 27 schematically illustrates another example of a method 2700 of improving liquid level sensing after dispensing a density gradient in accordance with the method 1100 of FIG. 11, or alternatively, after dispensing a sample on top of a density gradient in accordance with the method 2100 of FIG. 21. The method 2700 can be performed by the system 100. The method 2700 is directed to an alternative technique to remove droplets from the distal end of the probe 108 to improve the accuracy of the liquid level measurements captured by the system 100.

[0148] The method 2700 includes an operation 2702 of aspirating a volume of liquid inside the probe 108 prior to obtaining a liquid level measurement. Operation 2702 can include controlling one or more of the pumps 104 (see FIG. 1) to run in reverse to aspirate the volume of liquid. This causes a droplet on the distal end of the probe 108 to be pulled within the probe 108.

[0149] FIG. 28 provides an illustrate example where after dispensing a liquid L into the container 1 10, a droplet D is formed on the distal end of the probe 108. After operation 2702 is completed, the volume of liquid inside the probe 108 is aspirated such that the droplet D is pulled within the probe 108 and is no longer present on the distal end of the probe 108.

[0150] The volume of liquid aspirated in operation 2702 should be minimized to reduce risk of contamination in the lines running to the mixing manifold 106 where mixtures of liquids in the dispense queue are present. Operation 2702 can include aspirating a volume of liquid of about 10 uL which balances between reducing noise in the liquid level measurements and aspirating a minimum volume of liquid to avoid contamination. It is contemplated that alternative volumes of liquid can be aspirated in operation 2702 such as 27 uL, 5 uL, and 3 uL.

[0151] When there is no droplet present on the distal end of the probe 108 (see right side of FIG. 8), operation 2702 causes a small volume of air A to be pulled into the probe 108. The volume of air A can be pushed out of the probe 108 prior to dispensing the next liquid to ensure that there is no volume error accumulation over time due to air pockets inside the probe 108.

[0152] Referring back to FIG. 27, the method 2700 includes an operation 2704 of lowering the probe 108 to contact the meniscus M of the liquid L dispensed in the container 1 10 (see FIG. 28) to obtain a liquid level measurement. As discussed above, the liquid level measurement is obtained by detecting a change in capacitance when the distal end of the probe 108 touches the meniscus M of the liquid L within the container 110. By removing the droplet in operation 2702, the accuracy of the liquid level measurement obtained in operation 2704 is improved.

[0153] The method 2700 can further include an operation 2706 of raising the probe 108 after the liquid level measurement is obtained in operation 2704. For example, after the distal end of the probe 108 touches the meniscus M of the liquid L, the system 100 raises the probe 108.

[0154] The method 2700 can further include an operation 2708 of dispensing the volume of the liquid back into the probe 108. Alternatively, operation 2708 can include pushing out the air that was pulled into the probe 108 such as when there was no droplet present on the distal end of the probe 108. Operation 2708 can ensure that there is no volume error accumulation over time.

[0155] FIG. 29 is a plot 2900 illustrating liquid level measurements obtained by the system 100 before and after implementation of the method 2700. The plot 2900 displays differences in liquid level sensing (y-axis) between sequential measurements (x-axis). The data shown in the plot 2900 was obtained from dispensing a constant volume of liquid, and then measuring the liquid level using the liquid level sensing of the system 100. The data was repeated on the same types of containers for different volumes of liquid aspirated in operation 2702 of the method 2700. For example, the plot 2900 shows liquid level sensing data when 0 uL, 3 uL, 5 uL, 10 uL, and 27 uL are aspirated within the probe 108 before taking the liquid level measurements. Each of the volumes of liquid that were aspirated before each individual liquid level measurement, were pushed back out before the next dispense to avoid error accumulation.

[0156] As shown in the plot 2900, before implementation of the method 2700 (i.e., 0 uL is aspirated), significant noise is present. For example, the noise (i.e., difference in liquid level measurements between sequential measurements) ranges from about +2 mm to about - 1 mm.

[0157] The plot 2900 shows a significant noise reduction when droplet retraction is performed in accordance with the method 2700. For example, apart from an outlier in one run of aspirating a volume of 27uL of liquid from within the probe 108, the difference in liquid level measurements is reasonably flat. Similarly, the data from aspirating a volume of 10 uL of liquid from within the probe 108 also provides a relatively flat curve for the differences in liquid level sensing between sequential measurements. Given that it is desirable to minimize the volume of liquid aspirated in operation 2702, a volume of about lOuL appears to be sufficient for there to be a significant reduction in noise for the liquid level sensing by the system 100.

[0158] FIG. 30 show's an isometric view' of internal components of an example of the system 100 that includes a first portion 3000 for dispensing a density gradient and a sample on top of the density gradient, and a second portion 3002 for sealing a container after the density gradient and sample are dispensed. FIG. 31 illustrates a front view ofthe first and second portions 3000, 3002. As shown in the example of FIGS. 30 and 31, the first portion 3000 includes a first motor 3004 for raising and lowering the probe 108 along the height of a container. For example, the first motor 3004 can lower the probe 108 to the bottom of the container such as to dispense a density gradient using the underlay technique such as in the method 1100 of FIG. 11.

[0159] Once the density gradient is dispensed into the container, the first motor 3004 can raise the probe 108 to remove the probe 108 from the density gradient. The first motor 3004 can lower the probe 108 to dispense a sample on top of the density gradient while gradually raising the probe 108 in accordance with the overlay technique such as in the method 2100 of FIG. 21.

[0160] The second portion 3002 includes a second motor 3006 for raising and lowering a heating element 3012 along rails 3014 that are parallel to the height of the container. As will be described in more detail further below, the heating element 3012 is made of a heat conductive material and is configured to be electrically powered to generate heat for transferring to a seal former 3020 (see FIGS. 33 and 34) that is used to form a seal on the container.

[0161] The second portion 3002 eliminates the need for a user of the system 100 to position the heating element 3012 relative to the container 110, and to remove the heating element 3012 from the container 110. Advantageously, this can mitigate errors that result from manually positioning and removing the heating element 3012 from the container 1 10.

[0162] In some examples, the second portion 3002 is programmed to control the second motor 3006 to lower the heating element 3012 to automatically seal a container 110 upon detecting that the container 110 is positioned under the second portion 3002. For example, the system 100 can include a sensor 3024 that detects when a container 110 is positioned under the second portion 3002. Upon detection of the container 110 by the sensor 3024, the system 100 instructs the second motor 3006 to lower the heating element 3012 to seal the container 110.

[0163] In further examples, the sensor 3024 can detect presence of a container 110 under the first portion 3000 such that the system 100 is programmed to automatically dispense a density gradient into the container 110 when the sensor 3024 detects that the container 110 is positioned under the first portion 3000 of the system 100. In some examples, the sensor 3024 is a motion sensor, a time-of-flight (ToF) sensor, or othertype of sensor that can detect a presence of a container 110 under the first and second portions 3000, 3002 of the system 100.

[0164] The system 100 is programmed to include a timer for controlling the second motor 3006 to position the heating element 3012 and the seal former 3020 on top of the container 110 for a predetermined period of time for sealing the container 110. After expiration of the predetermined period of time, the second motor 3006 is triggered to raise the heating element 3012 and the seal former 3020 away from the container 110 to prevent damage to the container 110 that can be caused by prolonged exposure to the heat from the heating element 3012 and / or from the seal former 3020 pressing down on the shoulder of the container 110.

[0165] In alternative examples, the system 100 includes a switch that allows a user to manually trigger the second motor 3006 to lower the heating element 3012 to seal the container 110 when the container 110 is positioned under the second portion 3002. Thereafter, upon expiration of the predetermined period of time, the second motor 3006 is triggered to raise the heating element 3012 away from the container 110 to prevent damage to the container 110.

[0166] In yet further examples, the system 100 is configured to allow' the user to manually low er the heating element 3012 and the seal former 3020 along the rails 3014 to seal a container 110 positioned under the second portion 3002. The system 100 can include an alarm that illuminates a light switch and / or emits a sound to alert the user to raise the heating element 3012 along the rails 3014 away from the container 1 10 to prevent damage to the container.

[0167] The system 100 further includes a platform 3008 and a third motor 3010 that rotates the platform 3008 in at least one direction about a central axis of the platform 3008. For example, the third motor 3010 can rotate the platform 3008 in a clockwise direction. Alternatively, or additionally, the third motor 3010 that can rotate the platform 3008 in a second direction such as in a counterclockwise direction about the central axis of the platform 3008.

[0168] FIG. 32 show s a top isometric view of a carousel 3016 that can be mounted on the platform 3008. As shown in FIG. 32, the carousel 3016 includes a plurality of holders 3018 that can each hold a container 110. The third motor 3010 can rotate the platform 3008 to position a container 110 held by a holder 3018 on the carousel 3016 under the first portion 3000 for dispensing a density gradient and in some instances asample on top of the density gradient into the container 110. The third motor 3010 can then rotate the platform 3008 to position the container 110 under the second portion 3002 for sealing the container 110 after the density gradient and the sample are dispensed in the container 110 by the first portion 3000.

[0169] In some examples, the container 110 is detected as being positioned under the first portion 3000 or the second portion 3002 based on where the container 110 is held on the carousel 3016. and the position of the carousel 3016 relative to the first and second portions 3000, 3002.

[0170] FIG. 33 shows an example of a container 110 held by a holder 3018 on the carousel 3016 that is aligned with the seal former 3020 of the second portion 3002 of the system 100. FIG. 34 shows a bottom view of an example of the seal former 3020 and a seal guide 3022. The heating element 3012 heats the seal former 3020, and the second motor 3006 of the second portion lowers the seal former 3020 such that it is pressed onto a neck 111 of the container 110. The heat from the seal former 3020 causes the neck 111 of the container 110 to melt and deform, and eventually causing the neck 111 to seal. The seal guide 3022 can be made of an insulated material that does not transfer heat such that the heat is isolated to the seal former 3020 and does not cause damage to the container 110 or to other portions of the system 100.

[0171] FIG. 35 shows an example of a container 110 having the neck 111 sealed by the seal former 3020. Before being sealed, the neck 111 includes an opening 113 (see FIG. 33) allowing the probe 108 to be lowered into the container 1 10 for dispensing the density gradient. As shown in FIG. 35, the heat applied by the seal former 3020 causes the opening 113 in the neck 111 to close such that the neck 111 has a nipple shape formed by the seal former 3020.

[0172] FIG. 36 schematically illustrates an example of a method 3600 of sealing a container 110 that can be performed by the second portion 3002 of the system 100 of FIG. 30. As shown in FIG. 36, the method 3600 includes an operation 3602 of detecting a container 110 is positioned under the second portion 3002. As described above, the system 100 can include a sensor 3024 for detecting the container 110 is positioned under the second portion 3002. Alternatively, operation 3602 can include detecting that the container 110 is positioned under the second portion 3002 based on where the container 110 is held on the carousel 3016, and the position of the carousel 3016 relative to the first and second portions 3000, 3002 of the system 100.

[0173] The method 3600 includes an operation 3604 of sealing the container 110. Operation 3604 can include several stages that are part of the sealing process including an engagement phase 3606, a heating element contact phase 3608, a preheat phase 3610, a seal forming phase 3612, a holding phase 3614, a cooling phase 3616, and a retraction phase 3618. In some examples, the system 100 is configured to automatically perform all of the stages of operation 3604 without user involvement. This can eliminate human errors, and can also improve efficiency especially when sealing a plurality of the containers 110 having density gradients dispensed by the first portion 3000 of the system 100. The performance of the phases in operation 3604 that is automated by the system 100 can increase the throughput of the system.

[0174] During the engagement phase 3606, the heating element 3012 and the seal former 3020 start out separated and move down together along the rails 3014 by control of the second motor 3006. During the heating element contact phase 3608, as the neck 111 of the container 110 feeds into the seal former 3020, the seal former 3020 (which is mounted on a spring-loaded plate) will stop moving while the heating element 3012 continues to move down to contact the seal former 3020. During the preheat phase 3610, the heating element 3012 begins to transfer heat into the seal former 3020. This allows the seal former 3020 to come up to a predetermined temperature to begin melting the material on the neck 111 of the container 110.

[0175] The seal forming phase 3612 has the slowest speed for moving the heating element 3012 and the seal former 3020 downwards on the container 110. The slower speed of the seal forming phase 3612 gives time for the material in the neck 111 to melt and flow. This speed is limited by how quickly heat can transfer into the material of the neck 111 to make it soft enough to flow. In scenarios where the material in the neck 111 does not melt fast enough, the seal former 3020 will start to collapse the container 110 which can cause the liquid dispensed into the container 110 to be pushed out of the neck 111. This can cause the neck 111 to cool before the seal has been formed, which can result in an incomplete or poorly shaped seal.

[0176] In some instances, such as when the container 110 made of polypropylene material that is prone to compression during the seal forming phase 3612, the holding phase 3614 occurs at the end of travel in the forming phase. In the holding phase 3614, the heating element 3012 and the seal former 3020 remain in contact with the container 110 for a short period of time to ensure that all of the material in the neck 111 hasflowed together to form the seal. When sealing a container 110 that is made of a rigid material, such as Ultra-Clear™ available from Beckman Coulter. Inc., the holding phase 3614 can be skipped at the end of the seal forming phase 3612.

[0177] During the cooling phase 3616, the heating element 3012 backs off while the seal former 3020 stays in place due to the springs pushing downwards on an ejector plate that holds the seal former 3020. After the seal former 3020 has had time to cool, the retraction phase 3618 occurs which includes controlling the second motor 3006 to move the seal former 3020 upwards to clear a sealed nub on the neck 1 11 (e.g., see FIG. 35) such that the sealed container can be removed away from the second portion 3002 and an incoming unsealed container can be positioned under the second portion 3002. In instances where the container 110 becomes stuck to the seal former 3020 during the retraction phase, the system 100 can include a state foot to shuck the container 110 off the seal former 3020.

[0178] Table 1 summarizes exemplary parameters for the phases of operation 3604 for sealing different types of containers 110 by the second portion 3002 of the system 100. The specific settings for each type of container may van'.

[0179] As show n in Table 1, for containers 110 that are made of polypropylene material and that have an inner diameter of 16 mm, the heating element 3012 is controlled to have a lower sealing temperature (175 °C) to avoid burning holes throughthe tops of the containers 110 where the shoulder and the neck of the containers 110 meet. This issue can be avoided by lowering the sealing temperature applied by the heating element 3012 during the seal forming phase 3612.

[0180] FIG. 37 schematically illustrates an example of a method 3700 of preparing a container 110 for centrifugation that can be performed by the system 100 of FIG. 30. The method 3700 includes an operation 3702 of receiving a container 110 on the carousel 3016. For example, the container 110 can be received on one of the holders 3018 on the carousel 3016. In some examples, operation 3702 can include receiving a plurality of containers 110.

[0181] The method 3700 includes an operation 3704 of rotating the carousel 3016 to position the container 110 under the first portion 3000 of the system 100. Operation 3704 can include controlling the third motor 3010 to rotate the platform 3008 about the central axis.

[0182] The method 3700 includes an operation 3706 of dispensing liquid into the container 110 using the first portion 3000. Operation 3706 can include dispensing a density gradient into the container 1 10 accordance with the method 1100 of FIG. 11. In some examples, operation 3706 can also include dispensing a sample over the density gradient in accordance with the method 2100 of FIG. 21. In some further examples, operation 3706 can also include measuring a liquid level inside the container 110 in accordance with the method 2600 of FIG. 26, or in accordance with the method 2700 of FIG. 27 to verify a dispensed volume in the container 1 10.

[0183] The method 3700 includes an operation 3708 of rotating the carousel 3016 to position the container 110 under the second portion 3002 of the system 100. Operation 3708 can include controlling the third motor 3010 to rotate the platform 3008 about the central axis.

[0184] The method 3700 includes an operation 3710 of sealing the container 110. Operation 3710 can include sealing the container 110 accordance with the method 3600 of FIG. 36. For example, operation 3710 can include controlling the second motor 3006 to lower the heating element 3012 to gradually deform and melt the neck 111 on the container 110 for a predetermined period of time causing the neck 111 to seal the opening 113 as show n in the example of FIG. 35.

[0185] The method 3700 can include an operation 3712 of rotating the carousel 3016 such that the container 110 is positioned in an output location allowing a user toremove the container 110 from the carousel 3016 when the container 110 is no longer under the second portion 3002.

[0186] The various embodiments described above are provided by way of illustration only and should not be construed to be limiting in any way. Various modifications can be made to the embodiments described above without departing from the true spirit and scope of the disclosure.

Claims

What is claimed is:

1. A method of dispensing a set of density gradients, the method comprising: calculating a dispense queue including volumes of gradient materials for dispensing the set of density gradients, the set of density gradients including at least a first density gradient and a second density gradient for dispensing into at least a first container and a second container, respectively; dispensing the first density gradient into the first container by dispensing one or more sections composed of mixtures of the gradient materials through a probe; and purging the probe by displacing a mixture of gradient materials used for dispensing a last section of the first density gradient with a material included in the dispense queue before the dispense of the first density gradient is completed.

2. The method of claim 1, further comprising: positioning a distal end of the probe above a meniscus of the density gradient dispensed in the first container; and dispensing the material through the probe, wherein the material is a sample that is overlayed on top of the first density gradient.

3. The method of claim 2, further comprising: purging the probe by displacing the sample with a mixture of gradient materials for dispensing a first section of the second density gradient before the dispense of the sample in the first container is completed.

4. The method of claim 3, further comprising: dispensing the second density gradient in the second container by at least dispensing the first section of the second density gradient through the probe positioned to have the distal end at a bottom of the second container.

5. The method of claim 1, wherein the material is a mixture of gradient materials for dispensing a first section of the second density gradient in the second container.

6. The method of claim 5, further comprising:raising the probe such that the distal end of the probe is removed from the first container; positioning the distal end of the probe at a bottom of the second container; and dispensing the second density gradient in the second container by at least dispensing the first section of the second density gradient through the probe.

7. A system for dispensing a set of density gradients, the system comprising: a processing circuitry having a memory for storing instructions which, when executed by the processing circuitry, cause the system to: calculate a dispense queue including volumes of gradient materials for dispensing the set of density gradients, the set of density gradients including at least a first density gradient and a second density gradient for dispensing into at least a first container and a second container, respectively; dispense the first density gradient into the first container by dispensing one or more sections composed of mixtures of the gradient materials through a probe; and purge the probe by displacing a mixture of gradient materials used for dispensing a last section of the first density gradient with a material included in the dispense queue before the dispense of the first density gradient is completed.

8. The system of claim 7, wherein the instructions, when executed by the processing circuitry, further cause the system to: position a distal end of the probe above a meniscus of the density gradient dispensed in the first container; and dispense the material through the probe, wherein the material is a sample that is overlay ed on top of the first density gradient.

9. The system of claim 8, further comprising: a sensor assembly for detecting a location of the meniscus of the density gradient dispensed in the first container.

10. The system of claim 8 or 9, wherein the instructions, when executed by the processing circuitry, further cause the processing circuitry to:purge the probe by displacing the sample with a mixture of gradient materials for dispensing a first section of the second density gradient before the dispense of the sample in the first container is completed.

11. The system of claim 10, wherein the instructions, when executed by the processing circuitry, further cause the system to: dispense the second density gradient in the second container by at least dispensing the first section of the second density gradient through the probe positioned to have the distal end at a bottom of the second container.

12. The system of claim 7. wherein the material is a mixture of gradient materials for dispensing a first section of the second density gradient in the second container.

13. The system of claim 7, wherein the instructions, when executed by the processing circuitry, further cause the system to: raise the probe such that the distal end of the probe is removed from the first container; position the distal end of the probe at a bottom of the second container; and dispense the second density gradient in the second container by at least dispensing the first section of the second density gradient through the probe.

14. The system as in any one of the preceding claims, further comprising: pumps that are configured to pump the gradient materials from separate reservoirs at a predetermined volume and a predetermined speed for mixing inside a mixing manifold.

15. The system of claim 14, wherein the pumps include at least one pump selected from the group consisting of peristaltic pumps and syringe pumps.

16. The system of claim 14 or 15, further comprising: a mixer housed inside the mixing manifold, the mixer configured to mix the gradient materials together to generate a homogenous stream of fluid having a predetermined density based on relative concentrations of the gradient materials.

17. A system for dispensing a set of density gradients, the system comprising: a processing circuitry having a memory for storing instructions which, when executed by the processing circuitry, cause the system to: calculate a dispense queue including volumes of gradient materials for dispensing the set of density gradients, the set of density gradients including at least a first density gradient and a second density gradient for dispensing into at least a first container and a second container, respectively; dispense a first section of the first density gradient into the first container by dispensing a first mixture of the gradient materials through a probe; purge the probe by displacing the first mixture of gradient materials used for dispensing the first section of the first density gradient with a second mixture of gradient materials used for dispensing a second section of the first density gradient; dispense the second section of the first density gradient into the first container; and purge the probe by displacing the second mixture of gradient materials used for dispensing the second section of the first density gradient with a material included in the dispense queue before the dispense of the first density gradient is completed.

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