Methods and apparatus for processing tissues and cells

Abstract

This invention provides a system and method for processing biological samples. [Solution] The system comprises a support plate and at least one carriage assembly, the support plate having a central portion configured to reversibly interact with a motor for providing rotational force and at least one transverse arm, each of the at least one transverse arm including a first opening and at least one fixed opening, the at least one carriage assembly configured to reversibly interact with one of the at least one transverse arm, each of the at least one carriage assembly comprising a base portion and a tip assembly, the tip assembly comprising a microfluidic tip, a first sample chamber and a second sample chamber.

Description

[Technical Field]

[0001] [Incorporation by reference to priority application] All applications in which foreign or domestic priority claims are identified in the application data sheet filed with this application are incorporated herein by reference pursuant to U.S. Federal Regulation 37 1.57.

[0002] [Statement regarding federally funded research or development] This invention was made with the support of the U.S. Government under Contract No. 1R43DK116389-01, granted by the National Institute of Diabetes, Gastroenterology and Kidney Disease (NIDDK) of the National Institutes of Health (NIH). The U.S. Government may have certain rights in this invention.

[0003] [Areas of Disclosure] This application relates, in general, to apparatus and methods for applying shear stress to living cells, tissues, reagents, particles, and fluids, as well as to the use of cells obtained in the treatment of damaged tissue. In particular, the art relates to the use of rotational force in conjunction with microfluidic-based apparatus to apply shear stress to living cells and / or tissues, reagents, particles, and fluids. In some embodiments, the methods and apparatus described herein are applicable to the treatment of wounds such as diabetic ulcers, venous stasis ulcers, arterial ulcers, and pressure ulcers (i.e., bedsores). The methods and apparatus described herein are applicable to treatments related to surgical specialties such as orthopedics, arthroscopic surgery, neurosurgery, gastrointestinal and related organ surgery, urological surgery, general surgery, gynecological surgery, thoracic surgery, laparoscopic surgery, and plastic and reconstructive surgery when cosmetic body contouring is desired, when transplantation of treated and harvested adipose tissue is desired. [Background technology]

[0004] Diabetic foot ulcers (DFUs) are a major complication of diabetes mellitus (DM) and a leading cause of non-traumatic lower limb amputation. Lower limb amputation in diabetes is 2 to 1 in prevalence in men compared to women. The lifetime incidence of DFU formation in individuals with DM can be as high as 25%. In 2013, an estimated 384 million people had DM, and this number is projected to increase to 592 million by 2035. The pathogenesis of DFUs is multifactorial, primarily involving a combination of neuropathy, reduced vascular distribution, and isolated or recurrent trauma. Once a DFU forms, the wound microenvironment is characterized by poor healing due to continuous pressure and / or trauma, chronic infection, and displacement of the plantar fat pad. Current methods of treating DFUs are costly and have unpredictable outcomes. However, emerging evidence suggests that autologous stem cell therapy may be a safe and effective alternative to current treatment options.

[0005] Various techniques and procedures can be used to process tissues. In some applications, chemicals or enzymes are added to the tissue to break down large clumps or aggregates into increasingly smaller fragments. For example, digestive enzymes such as collagenase, trypsin, or dispase are used to digest tissues such as adipose tissue. Such enzymatic processing typically involves washing, followed by enzymatic digestion and centrifugation. This enzymatic approach can be variable due to differing activity levels of digestive enzymes. Furthermore, these methods require additional costs for reagents, including expensive enzymes derived from bacteria, and can take considerable time to complete. Similarly, such methods may require additional processing and / or washing steps to minimize the effects of enzymatic contamination.

[0006] Non-enzymatic approaches have also been developed for processing tissues including adipose tissue. For example, ultrasonic cavitation has been proposed for separating the interstitial vascular fraction from adipose tissue. (See Patent Document 1 (U.S. Patent No. 8,440,440), which is incorporated herein by reference in its entirety.) Yet another method involves using beads to homogenize adipose tissue, which is disclosed, for example, in Patent Document 2 (International Publication No. 2014 / 036094). (Patent Document 2 is incorporated herein by reference in its entirety.) Patent Document 3 (U.S. Patent No. 9,580,678) (which is incorporated herein by reference in its entirety) discloses a microfluidic tumor separation device used to destroy tumor tissue, which utilizes a plurality of sequentially arranged channels or stages having expansion and contraction regions. A syringe pump is used to allow tumor tissue to pass back and forth through the microfluidic device.

[0007] The processing of tissues such as adipose tissue is particularly important in the fields of plastic and reconstructive surgery, where adipose tissue is moved from one location to another to fill soft tissue defects (i.e., fat grafting). Cell-assisted fat grafting (CAL) is a technique that adds stromal vascular fraction (SVF) to fat grafts, which has greatly improved the maintenance of the fat graft. Typically, SVF is harvested from adipose tissue by a short digestion step using collagenase enzymes. More recently, a technique called "nanofat grafting" has been developed. This involves homogenizing standard liposuction material by manually and forcefully passing it between two connected syringes, and then reinjecting the homogenized liposuction material into a human patient to correct surface wrinkles and pigmentation. The nanofat processing method also serves as a means of mechanically separating SVF, and at the same time, it can stress cells to generate multipotent or pluripotent populations. For example, nano-adipose-derived SVF is known to have a high proportion of mesenchymal stem cells (MSCs), adipose-derived stem cells (ADSCs), endothelial progenitor cells (EPCs), and Muse cells. It was hypothesized that the amount of stress applied to the cells directly correlates with their stem-like properties.

[0008] For example, MSCs could be used to treat diabetic ulcers. Current treatments for diabetic foot ulcers, such as allogeneic transplantation, are expensive and may not be effective due to potential rejection by the patient. If such ulcers are left untreated, the patient may have to undergo limb amputation, which can lead to further health complications. One innovative solution to treat these ulcers is to use MSCs for direct treatment of these ulcers. However, current approaches to obtaining such cells are time-consuming, complex, and result in variable outcomes in terms of cell yield, volume, and reproducibility. A rapid and cost-effective method for obtaining processed tissue is needed. [Prior art documents] [Patent Documents]

[0009] [Patent Document 1] U.S. Patent No. 8440440 [Patent Document 2] International Publication No. 2014 / 036094 [Patent Document 3] U.S. Patent No. 9580678 [Patent Document 4] International Publication No. 2017 / 214323 [Overview of the project]

[0010] In consideration of the need for apparatus, systems, and methods for processing tissues efficiently, effectively, and reproducibly, various embodiments of such apparatus, methods, and systems, and their use in the development, production, or preparation of cells for processing (treatment) or administration to a subject, are provided herein. Various details relating to the apparatus for generating activated stem cells disclosed herein can be found in Patent Document 4 (International Publication No. 2017 / 214323), which is incorporated herein in its entirety by reference.

[0011] In some embodiments, a system for processing a biological sample with a support plate is provided. The support plate includes a central portion configured to reversibly interact with a motor for providing a rotational force, and at least one lateral arm. Each of the at least one lateral arm includes a first opening and at least one fixed opening. The system includes at least one carriage assembly configured to reversibly interact with one of the at least one lateral arms. The at least one carriage assembly of the system includes a base portion, a spring platform, and a fixing element. The base portion of the at least one carriage assembly includes an upper surface including at least one circular groove, a bottom surface including a column portion configured to extend through the first opening in one of the at least one lateral arms, a first arm, and a second arm. The spring platform includes at least one circular groove disposed on a bottom surface of the spring platform, a first engaging portion, and a second engaging portion. The spring platform is disposed adjacent to the upper surface of the base portion. The fixing element is configured to fix a chip assembly on the at least one carriage assembly.

[0012] The at least one carriage assembly may include at least one spring held between the base portion and the spring platform. The at least one spring is fixed in the at least one circular groove of the base portion at its first end and fixed in the at least one circular groove of the spring platform at its upper end. The at least one spring is configured to provide an upward force to the spring platform. Each of the plurality of carriage assemblies may be rotatable by at least about 180 degrees.

[0013] In other embodiments, the spring platform may be configured to move between a first position and a second position in response to insertion or removal of the chip assembly. In other embodiments, the spring platform may further include a tab, the fixed element further includes a channel configured to receive the tab of the spring platform, and the tab is configured to move within the channel of the fixed element when the spring platform moves between the first position and the second position. In other embodiments, the fixed element includes at least one guide rail configured to guide and position the chip assembly. In other embodiments, the first engaging portion of the spring platform is configured to hold the first arm of the base, and the second engaging portion of the spring platform is configured to hold the second arm of the base portion. In other embodiments, the spring platform further includes a platform configured to guide the chip assembly to the at least one carriage assembly. In other embodiments, the spring platform includes a splash guard on the bottom surface of the spring platform, and the splash guard is configured to prevent fluid from being introduced into the at least one spring.

[0014] In other embodiments, the chip assembly comprises a microfluidic chip, a first sample chamber, and a second sample chamber, wherein the microfluidic chip has a fluid path extending from a first end to a second end, the first sample chamber is fluidly connected to the first end of the microfluidic chip, and the second sample chamber is fluidly connected to the second end of the microfluidic chip. In other embodiments, the fluid path has at least one expansion region and a plurality of compression regions. In other embodiments, the at least one expansion region increases its radius along a first axis, a second axis, and a third axis, each perpendicular to the central axis of the fluid path, the at least one compression region has a diameter smaller than the diameter of the at least one expansion region, and the at least one compression region has no change in diameter. In other embodiments, the first axis, the second axis, and the third axis are perpendicular to each other. In other embodiments, the at least one expansion region increases in three or more dimensions (increasing beyond two dimensions). In other embodiments, the fluid path includes a plurality of teardrop-shaped expansion regions. In other embodiments, the fluid path includes a spherical or elliptical expansion region. In other embodiments, the fluid path includes a plurality of semi-teardrop-shaped expansion regions. In other embodiments, the fluid path includes a hemispherical or semi-elliptical expansion region. In other embodiments, the fluid path includes a D-shaped expansion region. In other embodiments, the fluid path includes an hourglass-shaped portion. In other embodiments, the fluid path comprises at least two expansion regions and at least one compression region located between the at least two expansion regions. In other embodiments, the fluid path comprises at least three expansion regions and at least two compression regions, each of the at least two compression regions located between adjacent expansion regions.

[0015] In other embodiments, a filter is located between the microfluidic chip and the first sample chamber, and between the microfluidic chip and the second sample chamber, at least one of the two. In other embodiments, the microfluidic chip of any of the systems disclosed herein is provided with Luer locks at the first and second ends of the microfluidic chip.

[0016] In some embodiments, a system is provided for processing a biological sample, comprising a support plate including a central portion configured to reversibly interact with a motor for providing rotational force, and at least one lateral arm, each of the at least one lateral arm having an interaction region. The system may also comprise at least one carriage assembly, the at least one carriage assembly configured to reversibly interact with one of the at least one lateral arm. In some embodiments, each of the at least one carriage assembly comprises a base portion, a spring platform positioned on the base portion, at least one spring held between the base portion and the spring platform and configured to provide an upward force to the spring platform, and a fixing element. The system may also comprise at least one chip assembly having a microfluidic chip, the microfluidic chip having a fluid path extending from a first end to a second end of the microfluidic chip. The fluid path may include at least one expansion region and at least one compression region. The at least one expansion region may have an increased diameter along a first axis, a second axis, and a third axis, each perpendicular to the central axis of the fluid path. The at least one compression region may have a smaller diameter than the at least one expansion region, and the at least one compression region is configured to maintain a constant diameter. The chip assembly may comprise a first sample chamber fluidly connected to a first end of the microfluidic chip and a second sample chamber fluidly connected to a second end of the microfluidic chip. The at least one chip assembly may be housed within the at least one carriage assembly, each of which is rotatable at least about 180 degrees.

[0017] In other embodiments, the first, second, and third axes are perpendicular to each other. In other embodiments, at least one expansion region increases in three or more dimensions (increasing beyond two dimensions). In other embodiments, the fluid path includes a plurality of teardrop-shaped expansion regions. In other embodiments, the fluid path includes a spherical or elliptical expansion region. In other embodiments, the fluid path includes a plurality of semi-teardrop-shaped expansion regions. In other embodiments, the fluid path includes a hemispherical or semi-elliptical expansion region. In other embodiments, the fluid path includes a D-shaped expansion region. In other embodiments, the fluid path includes an hourglass-shaped portion. In other embodiments, the fluid path comprises at least two expansion regions and at least one compression region located between the at least two expansion regions. In other embodiments, the fluid path comprises at least three expansion regions and at least two compression regions, each of the at least two compression regions located between adjacent expansion regions. In other embodiments, a filter is located between the microfluidic chip and the first sample chamber, and between the microfluidic chip and the second sample chamber, at least one of the two. In another embodiment, the microfluidic chip is provided with Luer locks at a first end and a second end of the microfluidic chip.

[0018] In some embodiments, a system for processing biological samples is provided. The system may comprise a support plate having a central portion configured to reversibly interact with a motor for providing rotational force, and a plurality of lateral arms, each having an interaction region. The support plate may comprise a plurality of retaining arms, each of which is positioned between each of the plurality of lateral arms, and each of which is configured to hold a syringe. The system may comprise at least one carriage assembly, the at least one carriage assembly configured to reversibly interact with one of the at least one lateral arms, and the at least one tip assembly is received within the carriage assembly. In some embodiments, each of the at least one carriage assembly is rotatable at least about 180 degrees.

[0019] In some embodiments, methods for treating damaged tissue are disclosed. This method involves administering a population of activated adipose stem cells to a subject having damaged or diseased tissue, at a rate of approximately 1 × 10⁶ per gram. 4 ~1 × 10 10 This may include administering in amounts within a range of 1. The activated adipose stem cells may result in one or more of the following: upregulation of the regenerating phenotype (e.g., CD34, CD13, CD73, or CD146), reduced inflammation, rapid tissue proliferation, rapid tissue remodeling, increased angiogenesis, or a combination thereof.

[0020] In other embodiments, the administration of the method is performed by subcutaneous injection. In other embodiments, the administration of the method is performed by intravenous injection. In other embodiments, the injured tissue is the result of an ulcer. In other embodiments, the ulcer is selected from the group consisting of diabetic foot ulcers, pressure ulcers (i.e., bedsores), venous stasis ulcers, and arterial ulcers. In other embodiments, the injured tissue is the result of a wound. In other embodiments, the wound is a burn-related wound, abrasion (e.g., road rash), laceration (e.g., knife wound), puncture wound, or delamination (e.g., bullet wound or wound from another weapon). In other embodiments, the administration results in a reduction of the time to heal the injured tissue by at least 10% compared to injured tissue not exposed to the activated adipose stem cells. In other embodiments, the administration results in a reduction of the time to heal the injured tissue by at least 50% compared to injured tissue not exposed to the activated adipose stem cells.

[0021] In some embodiments, methods for treating damaged tissue are disclosed. These methods may include administering to a subject having damaged or diseased tissue a population of activated adipose stem cells in an amount sufficient to increase angiogenesis at the site of injury.

[0022] In some embodiments, the use of activated adipose stem cells for treating damaged tissue is provided. In this case, the population of activated adipose stem cells is about 1 × 10⁶ per gram of the subject having damaged tissue. 4 ~1 × 10 10 Provided in quantities within a range of individual values, exposure of the damaged tissue to the activated adipose stem cells reduces the time it takes for the damaged tissue to heal by at least 10% compared to damaged tissue that has not been exposed to the activated adipose stem cells.

[0023] In other embodiments, the activated adipose-derived stem cells result in one or more of the following: upregulation of the regenerating phenotype (e.g., CD34, CD13, CD73, or CD146), reduced inflammation, rapid tissue proliferation, rapid tissue remodeling, increased angiogenesis, or a combination thereof. In other embodiments, administration reduces the time to healing of the damaged tissue by at least 50% compared to damaged tissue not exposed to the activated adipose-derived stem cells.

[0024] In some embodiments, a method for activating adipose stem cells for use in tissue repair is disclosed. The method may include the step of extracting a sample of adipose tissue from a patient in an amount sufficient to produce a sufficient amount of mechanically processed adipose-derived stem cells. The method may include the step of inserting the adipose tissue sample into a first sample chamber located at one end of a microfluidic chip. The method may include the step of rotating the microfluidic chip using a motor-driven rotary chuck so that the adipose tissue sample is subjected to shear forces. In the method, while the microfluidic chip is rotating, the adipose tissue sample reciprocates along a plurality of microfluidic channels from the first sample chamber to a second sample chamber located at the opposite end of the microfluidic chip. The method may include the step of removing the mechanically processed adipose-derived stem cells from the microfluidic chip. The method may include the step of administering the mechanically processed adipose-derived stem cells to the injured site of the patient. [Brief explanation of the drawing]

[0025] The features, aspects, and advantages described herein are described with reference to drawings of specific embodiments, which are intended to schematically illustrate specific embodiments and are not intended to limit the herein. [Figure 1] This document describes one embodiment of a system for processing biological samples. [Figure 1A]This shows an exploded view of one embodiment of a system for processing biological samples. [Figure 1B] An exploded view of the same embodiment is shown. [Figure 2A] An exploded view of another embodiment of a system for processing biological samples is shown. [Figure 2B] An exploded view of the same embodiment is shown. [Figure 3A] A diagram of the carriage assembly is shown. [Figure 3B] A diagram of the carriage assembly is shown. [Figure 4A] This figure shows one embodiment of a chip assembly. [Figure 4B] This figure shows the same embodiment. [Figure 5A] Figures 4A and 4B show one embodiment of a microfluidic chip configured to be fixed to the chip assembly. [Figure 5B] The same embodiment is shown. [Figure 5C] The same embodiment is shown. [Figure 6A] Figures 5A to 5C show enlarged views of the microfluidic chips configured to be fixed to the chip assemblies shown in Figures 4A to 4B. [Figure 6B] A magnified view of the microfluidic chip is shown. [Figure 6C] A magnified view of the microfluidic chip is shown. [Figure 6D] A magnified view of the microfluidic chip is shown. [Figure 7A] Another embodiment of a microfluidic chip configured to be fixed to the chip assembly shown in Figures 4A and 4B is shown. [Figure 7B] The same embodiment is shown. [Figure 7C] The same embodiment is shown. [Figure 8A] Another embodiment of a microfluidic chip configured to be fixed to the chip assembly shown in Figures 4A and 4B is shown. [Figure 8B] The same embodiment is shown. [Figure 9]Another embodiment of a microfluidic chip configured to be fixed to the chip assembly shown in Figures 4A and 4B is shown. [Figure 10] Another embodiment of a microfluidic chip configured to be fixed to the chip assembly shown in Figures 4A and 4B is shown. [Figure 11A] This document illustrates one embodiment of a three-dimensional channel in a microfluidic chip having an expanded region and a compressed region. [Figure 11B] The same embodiment is shown. [Figure 12A] Another embodiment of a three-dimensional channel in a microfluidic chip having an expanded region and a compressed region is shown. [Figure 12B] The same embodiment is shown. [Figure 13A] Another embodiment of a three-dimensional channel in a microfluidic chip having an expanded region and a compressed region is shown. [Figure 13B] The same embodiment is shown. [Figure 14A] This figure shows one embodiment of a sample chamber configured to be fixed to a microfluidic chip. [Figure 14B] This figure shows the same embodiment. [Figure 14C] This figure shows the same embodiment. [Figure 14D] This figure shows the same embodiment. [Figure 14E] This figure shows the same embodiment. [Figure 14F] This figure shows the same embodiment. [Figure 15A] This figure shows one embodiment of the filter. [Figure 15B] This figure shows the same embodiment. [Figure 16A] Figures 15A and 15B show the filters configured to be fixed to the chip assembly. [Figure 16B] This figure shows the filter. [Figure 17A] Figures 3A and 3B show one embodiment of the base platform of the carriage assembly. [Figure 17B] The same embodiment is shown. [Figure 18A] Figures 3A and 3B show one embodiment of the spring platform of the carriage assembly. [Figure 18B] The same embodiment is shown. [Figure 19A] Figures 3A and 3B show one embodiment of the fixing mechanism of the carriage assembly. [Figure 19B] The same embodiment is shown. [Figure 20A] This figure shows one embodiment of a screw cap. [Figure 20B] This figure shows the same embodiment. [Figure 21A] This figure shows one embodiment of a rotor foot insert. [Figure 21B] This figure shows the same embodiment. [Figure 22] This figure shows one embodiment of the base part of a system for processing biological systems. [Figure 23A] This figure shows another embodiment of a system for processing biological systems. [Figure 23B] This figure shows the same embodiment. [Figure 24] This figure shows another embodiment of a system for processing biological systems. [Figure 25A] This figure shows one embodiment of a system for processing a biological system, including one embodiment of a motor. [Figure 25B] This figure shows the same embodiment. [Figure 26A] This figure shows another embodiment of a system for processing biological systems, including another embodiment of the motor. [Figure 26B] This figure shows the same embodiment. [Figure 27A] This figure shows one embodiment of a system for processing a biological system having a structure for holding multiple syringes. [Figure 27B] This figure shows the same embodiment. [Figure 27C] This figure shows the same embodiment. [Figure 27D]This figure shows the same embodiment. [Figure 27E] Figures 27A to 27D show the tip of a structure for holding multiple syringes in an embodiment of a system for processing a biological system. [Figure 27F] The tip of the same structure is shown. [Figure 28A] A flowchart of a method for processing biological materials is shown. [Figure 29] This is a schematic diagram of one non-limiting embodiment of the treatment paradigm. [Figure 30A] This specification relates to phenotypic data of cells obtained from adipose tissue using the systems and methods disclosed herein, showing the proportion of CD26+ / CD55+ cells derived from adipose samples from healthy patients and diabetic patients. [Figure 30B] This specification relates to phenotypic data of cells obtained from adipose tissue using the systems and methods disclosed herein, showing the degree of CD34+ cells derived from adipose samples from healthy patients and diabetic patients. [Figure 31A] This specification relates to cell count and viability data for cells obtained from fat samples using the systems and methods disclosed herein, showing cell density obtained from processing fat samples from healthy patients and diabetic patients. [Figure 31B] This specification relates to cell count and viability data for cells obtained from fat samples using the systems and methods disclosed herein, showing cell viability obtained from processing fat samples from healthy patients and diabetic patients. [Figure 32A] This relates to the types of cells present after processing fat samples using the systems and methods disclosed herein, and shows the percentage of endothelial progenitor cells (EPCs) in cell populations derived from processing fat samples from healthy patients and diabetic patients. [Figure 32B]This relates to the types of cells present after processing fat samples using the systems and methods disclosed herein, and shows the percentage of mesenchymal stem cells (MSCs) in cell populations derived from processing fat samples from healthy patients and diabetic patients. [Figure 32C] This relates to the types of cells present after processing fat samples using the systems and methods disclosed herein, and shows the percentage of Muse cells in cell populations derived from processing fat samples from healthy patients and diabetic patients. [Figure 33A] This specification relates to the characteristics of cells and cell types after processing adipose samples using the systems and methods disclosed herein, and shows the cell viability of cells derived from adipose samples under the indicated conditions. [Figure 33B] This specification relates to the characteristics of cells and cell types after processing adipose samples using the systems and methods disclosed herein, and indicates the cell count of cells derived from the adipose samples under the indicated conditions. [Figure 33C] This specification relates to the characteristics of cells and cell types after processing adipose samples using the systems and methods disclosed herein, and shows subpopulations of cell types derived from adipose samples under the indicated conditions. [Figure 34A] This specification presents data related to wound healing using cells obtained by processing fat samples using the systems and methods disclosed herein, and in particular, histological data regarding the presence of specific markers and tissue regeneration. [Figure 34B] This specification presents data related to wound healing using cells obtained by processing fat samples using the systems and methods disclosed herein, and in particular, photographs of the progression of wound healing over time in a mouse injury model. [Figure 34C] This specification presents data related to wound healing using cells obtained by processing fat samples using the systems and methods disclosed herein, and in particular provides summary data on wound healing for the indicated groups. [Modes for carrying out the invention]

[0026] While some examples are given below, those skilled in the art will understand that this disclosure extends beyond the examples and / or uses specifically disclosed, as well as obvious modifications and their equivalents. Therefore, the scope of the disclosure disclosed herein is not intended to be limited by the specific examples given below.

[0027] In some embodiments, a system for processing biological samples comprises a support plate and a plurality of carriages, the support plate having a central portion containing a receptor element and a lateral portion containing a plurality of interaction regions, each interaction region configured to reversibly interact with the plurality of carriages, and each of the plurality of carriages configured to be operably coupled to the lateral portion of the support plate.

[0028] In some embodiments, the receiving element is configured to reversibly interact with the drive shaft of a motor configured to impart centrifugal motion to a support plate. In one embodiment, the central portion lies in a plane perpendicular to the axis of rotation of the motor's drive shaft, and the lateral portion extends radially from the central portion and lies in a plane at least partially parallel to the plane of the central portion.

[0029] In some embodiments, each of the multiple carriages comprises a first end, a second end, a base portion extending between the first and second ends, and a receiving region configured to reversibly interact with a microfluidic chip. The microfluidic chip is fluid-coupled to at least one sample chamber configured to receive a sample for processing. In some embodiments, each of the multiple carriages comprises a column, rod, shaft, or other extension extending substantially perpendicularly from the base portion and configured to interact with (e.g., by connecting, attaching, or otherwise) one of the multiple interaction regions of the lateral portion. In some embodiments, each of the multiple carriages is coaxially arranged around one of the multiple axes, and during operation, each axis extends substantially parallel to the axis of rotation of the motor's drive shaft, and each of the multiple carriages is at least intermittently rotatable about one of the multiple axes. Depending on the embodiment, the carriages can rotate to a variety of angles. For example, in some embodiments, the carriages are guided to rotate over an arc of about 180 degrees.

[0030] In some embodiments, the system further comprises at least one microfluidic tip that plays a role in holding and processing a sample according to the system. In some embodiments, each microfluidic tip comprises a central body portion located between a first end and a second end, and at least one microfluidic channel extending between the first end and the second end, the at least one channel having various dimensions and configured to allow the passage of a sample from the first end to the second end. In some embodiments, each of the first and second ends is configured to fluidically interact with the sample chamber. For ease of use, each microfluidic tip is sized to fit within the corresponding receiving area of ​​the corresponding carriage. In some embodiments, each microfluidic tip is reversibly fluidically coupled to the sample chamber at each of the first and second ends.

[0031] In some embodiments, the sample chamber comprises a vent and a vent channel fluidically connected to the interior of the sample chamber. In some embodiments, each sample chamber is reversibly fluidically coupled to a microfluidic chip via an adapter.

[0032] In some embodiments, each carriage is provided with a capture element at first and second ends of the carriage, the capture element being configured to communicate with a release element on the lateral portion of a support plate, and the communication between the capture and release elements allows the release element to enable the intermittent rotation of each of the multiple carriages. In other words, the capture element plays a role in holding the carriage in a desired position until there is a signal (or force, or lack thereof) that allows the capture element to disengage from the release element or otherwise stop. The carriage is then allowed to rotate, followed by re-engagement of the capture element to stop the movement of the carriage (in some embodiments, this allows the carriage to rotate over a 180-degree arc at a desired time in the tissue processing protocol). In some embodiments, the capture element includes a magnet of first polarity, and the release element includes a magnet of opposite polarity.

[0033] In some embodiments, the lateral portion of the support plate includes a disk, and multiple interaction regions are circumferentially spaced around the disk. In such embodiments, the lateral portion and the central portion are integrally constructed, but in other embodiments, the support plate may consist of multiple parts that are connected or integrated before use.

[0034] In some embodiments, the lateral portion of the support plate includes a plurality of arms, each arm including a corresponding interaction region. In one embodiment, the arms and the central portion are a single integrated structure. In another embodiment, the arms and the central portion are separate structures coupled to one another. In some embodiments, the arms are hinged to the central portion. In such embodiments, the hinges allow the arms to move in a plane of an axis substantially parallel to the rotation axis of the motor's drive shaft during operation. In some embodiments, this allows for a gentle start and stop process so that abrupt application or removal of centrifugal force does not damage the cell / tissue sample.

[0035] In some embodiments, the interaction region of the lateral portion comprises a through-hole that receives a column (or other structure) from the corresponding carriage. In some embodiments, the receiving region is located on the upper surface of the base portion of the carriage. In some embodiments, the column extends from the bottom surface of the base portion of the carriage. In such embodiments, the column (or other structure) extends from the bottom of the carriage, passes through a hole (receiving region) in the lateral portion (e.g., an arm), and is secured (to allow rotation relative to the lateral portion) by, for example, a nut, pin, clamp, or other mechanism. In some embodiments, intermittent rotation of each carriage is achieved by the interaction of a fixed tooth that induces rotation of each carriage with a gear located in the lateral portion.

[0036] In some embodiments, the lateral portion includes at least three arms, each of which includes an interaction region configured to interact with one of at least three carriages, each of which is configured to reversibly interact with one of at least three microfluidic chips, each of which includes a first end, a second end, and a body between them, each end of the microfluidic chip being fluid-coupled to a sample chamber, the body of the microfluidic chip including a plurality of microfluidic paths extending between the first end and the second end, the carriage being configured to rotate intermittently between a first position and a second position, the first position being such that the first end is positioned at a first location at a first distance from the central receiving element, and the second position being such that the first end is positioned at a second location at a second distance from the central receiving region, the first distance being greater than the second distance.

[0037] Depending on the embodiment, the system may optionally include a housing, which isolates the system from the external environment.

[0038] Depending on the embodiment, the system may further include a motor operably connected to the drive shaft. In some embodiments, the motor is controlled by a controller unit that enables control of the motor's rotational speed, and the controller unit includes an interface that enables a user to program (or select from pre-programmed) protocols for processing tissue.

[0039] This specification also provides methods for processing biological samples. For example, in some embodiments, the method includes introducing a biological sample into a first sample chamber configured to be fluidly coupled to a microfluidic chip. The microfluidic chip has a central body portion located between a first end and a second end. The first end is configured to be fluidly coupled to a first sample chamber, and the second end is configured to be fluidly coupled to a second sample chamber, with at least one microfluidic channel extending between the first and second ends. The at least one channel includes various dimensions and is configured to allow the passage of a sample from the first end to the second end. The method includes reversibly coupling the microfluidic chip to a receiving region of one of a plurality of carriages which are part of a centrifuge. The centrifuge comprises a support plate having a central portion and lateral portions. The lateral portions of the centrifuge extend radially from the central portion and are located in a plane parallel to the plane of the central portion. Each carriage is operably coupled to the side of a support plate and comprises a first end, a second end, and a base portion extending between the first and second ends. The base portion of the carriage includes a receiving region, and each carriage is configured to be rotatable about an axis substantially perpendicular to the plane of the central portion. The carriage is rotatable from a first position to a second position. In the first position, the first end is located at a first distance from the receiving region, and in the second position, the second end is located at the first distance from the central portion of the support plate. The method includes applying a rotational force to a centrifuge. By applying a rotational force to the centrifuge, the sample passes from a first sample chamber coupled to the first end of the microfluidic chip through at least one microfluidic channel extending between the first and second ends into a second sample chamber. The method includes enabling the carriage to rotate between the first and second positions.

[0040] The method includes applying a further rotational force to cause the sample to return from the second sample chamber to the first sample chamber through at least one microfluidic channel extending between the first and second ends. In some embodiments, the biological sample includes adipose tissue, but other tissue types may be processed using the systems and methods disclosed herein. For example, adipose tissue, tumor tissue, cell preparations, liposuctions, cultured cells, etc., can be easily processed.

[0041] Furthermore, in some embodiments, a system is provided for processing a sample, the system comprising a support plate and at least one microfluidic chip, the support plate having a plurality of rotatable carriages arranged radially around the support plate, the at least one microfluidic chip positioned on one of the rotatable carriages, the at least one microfluidic chip defining a fluid path formed by one or more microfluidic channels arranged therein, the fluid path extending from a first port of the microfluidic chip to a second port located at the opposite end of the microfluidic chip.

[0042] In some embodiments, the support plate comprises a plurality of arms, each of which holds a rotatable carriage. In such embodiments, the plurality of arms are fixed to a separate central hub. In some embodiments, the support plate includes a first magnetic element located in or on the support plate and adjacent to the end of the rotatable carriage, and the rotatable carriage further includes a second magnetic element located in or on the carriage.

[0043] Alternatively, in some embodiments, a rotatable carriage is coupled to a gear set arranged in a gear assembly mounted on a support plate, the gear set including gears exposed on the radially outer portion of the gear assembly. In some embodiments, the gear assembly or support plate further comprises fixed magnets disposed therein, and the rotatable carriage includes a pair of magnetic elements positioned at both ends thereof.

[0044] In some embodiments, the multiple rotatable carriages are rotatable in a plane substantially parallel to the plane of rotation of the support plate. In some embodiments, the multiple rotatable carriages are rotatable in a plane substantially perpendicular to the plane of rotation of the support plate.

[0045] In some embodiments, the system further comprises electromagnets positioned on support plates beneath each of a plurality of rotatable carriages, the rotatable carriages comprising magnetic post elements (magnetic column elements) extending through openings formed in the support plates.

[0046] In some embodiments, at least one of the first and second sample holding chambers comprises a syringe barrel, e.g., a standard 2 mL, 5 mL, 10 mL, 20 mL, or 60 mL syringe barrel. In some embodiments, at least one microfluidic tip is further included, interposed between the first and second sample holding chambers. In some embodiments, the filter is positioned upstream or before the microfluidic tip and is configured to filter the sample to prevent clogging of the microfluidic tip. In some examples, the upstream filter may include a mesh configured to cut or pulverize tissue or tissue fragments so that the sample can pass through the microfluidic tip without clogging. The cutting or pulverization of the sample is configured to generate macroscopic aggregates for the purpose of microfluidic shearing in the microfluidic tip. In some embodiments, the filter is positioned downstream or behind the microfluidic tip and allows only samples of a specific size to pass out of the device for collection.

[0047] In some embodiments, the system also includes a sample holding chamber housed in a rotatable carriage and coupled to a first port of the microfluidic tip, and a syringe coupled to a second port of the microfluidic tip, the syringe mounted substantially perpendicular to the plane of rotation of the microfluidic tip. In some embodiments, the system disclosed herein may optionally include a vertically movable plate or ring coupled to the plunger of the syringe. In some embodiments, the vertically movable plate or ring includes a female threaded bearing mounted on a rotatable threaded rod. Optionally, certain embodiments further include a second motor coupled to the threaded rod.

[0048] In some embodiments, the system includes at least one of a first sample holding chamber and a second sample holding chamber, each containing an inlet with a one-way valve located inside.

[0049] In some embodiments, a method is provided using the system disclosed herein, the method comprising: rotating a support plate to move a sample through a first port into one or more microfluidic channels of at least one microfluidic chip and out through a second port; rotating a rotatable carriage containing at least one microfluidic chip about 180 degrees; rotating the support plate to move a sample through the second port into one or more microfluidic channels and out through the first port; and repeating the above steps multiple times until the sample is processed to a desired extent.

[0050] In such a method, the sample moves between a first sample holding chamber fluidly connected to a first port and a second sample holding chamber fluidly connected to a second port. In some embodiments, at least one of the first and second sample holding chambers is equipped with a syringe barrel.

[0051] In some embodiments, the sample includes tumor tissue. In some embodiments, the sample includes adipose tissue. In some embodiments, the sample includes a fluid containing one or more reagents. In some embodiments, the sample includes particles (e.g., nanoparticles, magnetic particles, particles coated with reagents or antibodies). In some embodiments, the sample includes a cell-containing fluid.

[0052] In some embodiments, after processing the tissue, the methods disclosed herein further include injecting the processed tissue (e.g., adipose tissue) into a subject (subject).

[0053] In addition to the systems, apparatus, and methods described above, the following system for processing samples is also provided herein. The system comprises a support plate, a plurality of arms extending radially from the support plate, a motor coupled to the support plate and configured to rotate the support plate, and a plurality of carriages, each of which carriages is positioned on one of the plurality of arms on the support plate, and each of which carriages is positioned coaxially around one of a plurality of axes, each axis extending perpendicularly from the arm on which the carriage is positioned, and each of which carriages is configured to receive a microfluidic tip and at least one sample chamber for receiving a sample for processing, the at least one sample chamber having an opening fluidly connected to the microfluidic tip, and each of which carriages is rotatable around one of the plurality of axes.

[0054] In some embodiments, the system further comprises a controller configured to drive a motor, the controller configured to adjust the motor's rotational speed or revolutions per minute (RPM). In some embodiments, the controller is adjustable or programmable by a predetermined spin program or series of operations. In some embodiments, the controller is configured (or configurable) to increase the motor's spin speed to an RPM speed such that a sample is configured to flow from a first end of the microfluidic chip to a second end of the microfluidic chip. In some embodiments, the controller is configured to accelerate or decelerate the motor's RPM such that each of a plurality of carriages is configured to rotate around one of a plurality of axes.

[0055] In some embodiments, each of the multiple carriages is configured to rotate 180 degrees around one of the multiple axes. Furthermore, in some embodiments, each of the multiple carriages is configured to receive a first sample chamber and a second sample chamber, the first sample chamber being located at the first end of the microfluidic chamber and the second sample chamber being located at the second end of the microfluidic chamber.

[0056] Some embodiments include at least one sample chamber attached to the end of a microfluidic tip using an adapter. The adapter may include a Luer slip, a slip tip connector, a Luer lock, and a rotating collar. Depending on the embodiment and whether the microfluidic tip is disposable or reusable (e.g., sterilizable), the adapter may be made of metal or polymer material.

[0057] In some embodiments, the support plate is housed within a case (enclosure), which is configured to protect the user from the system for processing the sample. The case is made of any material, such as plastic or metal, and is provided to be thick enough to prevent penetration or rupture of the case by fragments caused by a drop in vacuum pressure, low temperatures, thermal changes, or centrifugal rotation. In some embodiments, the case (enclosure) is configured to be opened and closed as needed to place a sample, remove a sample, or manually rotate one or more carriages. In some embodiments, the case is optically transparent and configured to allow monitoring of the operation of the system for processing the sample.

[0058] In embodiments including multiple arms, each of the multiple arms further includes a first engagement structure and a second engagement structure located at a distance from the first engagement structure, wherein the first and second engagement structures are configured to engage with one of a first structure located at a first end of the carriage and a second structure located at a second end of the carriage, respectively. In some such embodiments, the first and second engagement structures are configured to release and engage the first and second structures interchangeably so that the carriage moves in multiple orientations around one of the multiple axes.

[0059] In some embodiments, the first and second engagement structures are magnets, and the first and second structures include a magnetically responsive material. In some embodiments, the first engagement structure is positioned distal to the second engagement structure along the length of each of the multiple arms. To move the carriage, in some embodiments, an accelerating or decelerating force is configured to move the carriage between multiple orientations. In some embodiments, the carriage includes a gear assembly, and the gear assembly is configured to move the carriage between multiple orientations. In some embodiments, the carriage includes a centering ratchet, and the centering ratchet is configured to move the carriage between multiple orientations.

[0060] To enable proper flow of a sample through a microfluidic tip, in some embodiments, at least one sample chamber includes a vent and a vent channel fluid-connected to the interior of the sample channel, the vent configured to provide laminar flow through the sample chamber (e.g., by preventing a vacuum). In some embodiments, the vent is located at the end of the sample chamber opposite the opening. Depending on the embodiment, the sample chamber can be any desired shape, including rectangular, square, ellipsoidal, columnar, elliptical, or other polygonal shapes. In one embodiment, the sample chamber is rectangular. In some alternative embodiments, the sample chamber is a syringe. In some such embodiments, the syringe comprises a chamber having an adapter end, the adapter end having an opening configured to fluid-connect to a microfluidic tip, and a plunger comprising a seal located within the chamber. An indenter is attached to the distal end of the plunger and configured to advance and retract the plunger. In some embodiments, the syringe includes a vent and a vent channel fluid-connected to the interior of the chamber, the vent configured to provide laminar flow through the syringe. Furthermore, in some embodiments, the syringe may optionally include a secondary syringe plunger, which is positioned within a vent channel and configured to selectively open and close the vent channel. In such embodiments, the secondary syringe is optionally coupled to the indenter such that the movement of the indenter is configured to advance and retract both the plunger and the secondary plunger. In some embodiments, the adapter end of the syringe is configured to receive a needle. In some embodiments, the syringe is configured to be detachable from the microfluidic tip, and the sample is configured to be injected directly into the injection site.

[0061] In some embodiments, each of the multiple chambers (e.g., chambers for holding microfluidic chips) is held in the opening of each of the multiple arms, and each of the multiple chambers extends through the opening of each of the multiple arms. In some embodiments, each of the multiple chambers is held along the plane of each of the multiple arms. In some embodiments, each of the multiple chambers is held in the opening of each of the multiple arms by at least one pin configured to allow out-of-plane rotation of each of the multiple chambers. In some embodiments, the out-of-plane rotation of each of the multiple chambers is configured to move each of the multiple chambers between multiple orientations. In some embodiments, each of the multiple chambers moves between rotations of 180 degrees (in-plane or out-of-plane). In some embodiments, each of the multiple chambers moves between orientations in which each of the multiple chambers is positioned along the plane of each of the multiple arms.

[0062] In some embodiments, the systems disclosed herein further comprise at least one filter configured to prevent larger sample components from entering and clogging the microfluidic chip. In some embodiments, the filter is mounted in the sample chamber to filter the sample before it passes through the microfluidic pathway of the microfluidic chip. In some embodiments, the filter is positioned upstream or before the microfluidic chip and is configured to filter the sample to prevent clogging of the microfluidic chip. In some examples, the upstream filter may include a mesh configured to cut or pulverize tissue or tissue fragments so that the sample can pass through the microfluidic chip without clogging. The cutting or pulverization of the sample is configured to generate macroscopic aggregates for the purpose of microfluidic shearing in the microfluidic chip. In some embodiments, the filter is positioned downstream or behind the microfluidic chip to allow only samples of a specific size to exit the instrument for collection.

[0063] Additional systems are also provided herein. For example, a system for processing a sample is provided, comprising a support plate, a motor coupled to the support plate and configured to rotate the support plate, and at least one carriage positioned on the support plate, the at least one carriage configured to receive a microfluidic chip and at least one sample chamber for receiving a sample for processing, and the at least one carriage configured to rotate in a plane parallel to the plane of the support plate.

[0064] Furthermore, a system for processing a sample is provided, the system comprising a support plate, a motor coupled to the support plate and configured to rotate the support plate, at least one carriage positioned on the support plate and configured to rotate in a plane parallel to the plane of the support plate, and a microfluidic chip received within at least one carriage, the microfluidic chip comprising a port, at least one microfluidic channel extending along the length of the microfluidic chip, and at least one sample chamber for receiving a sample for processing, the at least one sample chamber being fluidly connected to a first port of the microfluidic chip and configured to allow a sample to flow from at least one sample chamber along the length of the microfluidic chip.

[0065] In some embodiments, the microfluidic chip has a length of about 10 mm to 100 mm. In some embodiments, the length may be about 10 mm, about 20 mm, about 30 mm, about 40 mm, about 50 mm, about 60 mm, about 70 mm, about 80 mm, about 90 mm, or about 10 to 20 mm, about 30 to 40 mm, about 40 to 50 mm, about 50 to 60 mm, about 60 to 70 mm, about 70 to 80 mm, about 80 to 90 mm, or about 90 to 100 mm, or any value within these ranges including upper and lower limits. In some embodiments, the length of at least one microfluidic channel is less than (or equal to) the length of the microfluidic chip. In some embodiments, the width and depth of the microfluidic channel are in the range of 5 μm to 8 mm. In some embodiments, the microfluidic channels may be approximately 5–200 μm, 200–400 μm, 400–600 μm, 600–800 μm, 800–1000 μm, 1–2 mm, 2–4 mm, 4–6 mm, or 6–8 mm, or any value within these ranges, including upper and lower limits. In some embodiments, it should be understood that the microfluidic chip is removable.

[0066] Microfluidic channels can have various configurations depending on the embodiment and the tissue being processed. For example, in one embodiment, the microfluidic channel has an hourglass configuration. In some embodiments, at least one microfluidic channel has a first region including a stepped taper in which the width gradually decreases along the length of at least one microfluidic channel, a contraction region, and a second region including a stepped taper in which the width gradually increases along the length of at least one microfluidic channel. In another embodiment, at least one microfluidic channel has a series of increasing width regions and decreasing width regions. In some embodiments, at least one microfluidic channel has a diamond pattern (rhombic shape). In some embodiments, at least one microfluidic channel includes a plurality of pockets, the pockets may optionally be fin-shaped. In some embodiments, at least one microfluidic channel includes a first region including a series of branches and a second region where pairs of branched channels are recombined. In another embodiment, at least one microfluidic channel includes a plurality of wells, the plurality of wells configured to sort portions of a predetermined size of sample.

[0067] This specification provides further methods, such as methods for processing a sample. These methods include preparing a sample in a sample chamber, inserting the sample chamber into at least one of a plurality of carriages, and rotating a support plate about a first axis. In these methods, the sample chamber is fluidly connected to a microfluidic tip containing at least one microfluidic channel, one of the plurality of carriages is mounted on a support plate and configured to rotate about a first axis, at least one of the plurality of carriages is configured to rotate about a second axis, the second axis being parallel to the first axis, and the rotation of the support plate about the first axis is configured to drive the sample out of the sample chamber, through at least one microfluidic channel, in a first direction away from the sample chamber. In some embodiments, the methods further include rotating at least one of the plurality of carriages in a second direction about a second axis. In some embodiments, the method further includes rotating a support plate about a first axis, the rotation being configured to drive the sample in a second direction toward the sample chamber through at least one microfluidic channel. Furthermore, the method optionally further includes detaching the sample chamber from at least one of a plurality of carriages.

[0068] Further systems are provided, comprising a support plate including a plurality of arms, a motor, a plurality of carriages, and a case (housing). In the system, the plurality of arms extend radially from the support plate, the motor is coupled to the support plate and configured to rotate the support plate, each of the plurality of carriages is positioned on one of the plurality of arms on the support plate, each of the plurality of carriages is positioned coaxially around one of a plurality of axes, each axis extends perpendicularly from the arm on which the carriage is positioned, each of the plurality of carriages is configured to receive a microfluidic tip and at least one sample chamber for receiving a sample for processing, each of the plurality of carriages is rotatable about one of the plurality of axes, the case is configured to receive the support plate, the case is configured to protect the user from the system for processing the sample, and the case includes an opening configured to provide access to the sample chamber. In some embodiments, at least one sample chamber includes a vent and a vent channel fluid-connected to the inside of the sample channel, the vent is configured to provide laminar flow through the sample chamber. In some embodiments, at least one sample chamber includes an opening fluidly connected to a microfluidic tip. In some embodiments, at least one sample chamber includes an inlet configured to allow the sample to be processed to be inserted into or removed from at least one sample chamber. In one embodiment, the inlet is located on the opposite side of the opening, but other locations may be used as desired. In some embodiments, at least one sample chamber includes a one-way valve configured to keep the sample inside the chamber during processing. In some embodiments, the interior of the sample chamber has a surface with an inclined face, chamfer, or other shape adjacent to the inlet, which is configured to agglomerate the sample adjacent to the inlet for easy removal of the sample after processing. In one embodiment, a vent is located at the end of the sample chamber opposite the opening.In some embodiments, the inlet is configured to engage with a syringe, which is configured to take out a sample and inject the sample directly into a target site.

[0069] In some embodiments, each of the multiple chambers is held in the opening of each of the multiple arms, and each of the multiple chambers extends through the opening of each of the multiple arms. In such embodiments, each of the multiple chambers is arbitrarily held along the plane of each of the multiple arms. In some embodiments, each of the multiple chambers is arbitrarily held in the opening of each of the multiple arms using at least one pin configured to allow out-of-plane rotation of each of the multiple chambers. In some such embodiments, the out-of-plane rotation of each of the multiple chambers is configured to move each of the multiple chambers between multiple orientations. In some embodiments, each of the multiple chambers moves between rotations of 180 degrees (e.g., 0–45 degrees, 45–90 degrees, 90–135 degrees, 135–180 degrees, etc.). In some embodiments, each of the multiple chambers moves between orientations in which each of the multiple chambers is positioned along the plane of each of the multiple arms. In some embodiments, the system further comprises a filter configured to prevent larger sample components from entering and clogging the microfluidic tip. In one embodiment, the filter is attached to the sample chamber. In some embodiments, the filter is positioned upstream or before the microfluidic chip and is configured to filter the sample to prevent clogging of the microfluidic chip. In some examples, the upstream filter may include a mesh configured to cut or pulverize tissue or tissue fragments so that the sample can pass through the microfluidic chip without clogging. The cutting or pulverization of the sample is configured to create macroscopic aggregates for the purpose of microfluidic shearing in the microfluidic chip. In some embodiments, the filter is positioned downstream or behind the microfluidic chip to allow only samples of a specific size to exit the instrument for collection.

[0070] In some embodiments, a system for processing samples is provided, comprising a support plate including a plurality of arms, a motor, a plurality of carriages, a case (housing), a spin stand, a threaded rod, and a plate, wherein the plurality of arms extend radially from the support plate, the motor is connected to the support plate and configured to rotate the support plate, each of the plurality of carriages is positioned on one of the plurality of arms on the support plate, each of the plurality of carriages is positioned coaxially around one of a plurality of axes, each axis extends perpendicularly from the arm on which the carriage is positioned, each of the plurality of carriages is configured to receive a microfluidic chip and at least one sample chamber to receive a sample for processing, each of the plurality of carriages is rotatable about one of the axes, and the case includes a body and a cover, the body is configured to receive the processing system, A bar is positioned on top of the main body and configured to seal the processing system within the main body, protecting the user from the system processing the sample; the spin stand is equipped with a motor, a threaded rod is motor-mounted, the rotation of the motor rotates the threaded rod; the plate is equipped with multiple engagement structures for holding a syringe, the plate is mounted on a bearing having a female thread, the female thread is configured to engage with the male thread of the rod, the rotation of the motor is configured to raise or lower the plate vertically; the syringe comprises a chamber having an opening configured to be fluidly connected to at least one sample chamber, and a plunger positioned within the chamber, the forward or backward movement of the plunger discharges or takes in a sample for processing; the plate holds the distal end of the plunger, the plate lowers or raises the plunger vertically within the syringe chamber to discharge or take in a sample for processing.

[0071] In some embodiments, the rotary motor is mounted on the cover of the case (housing), while in other embodiments, the rotary motor is located outside the case (housing). In some embodiments, the plate is circular, while in some embodiments, the plate includes a ring attached to a bearing by a plurality of arms. In one embodiment, the plate comprises a central circular plate and a coaxial ring. In some embodiments, the engagement structure is hook-shaped and configured to allow insertion and removal of the distal end of the plunger. In some embodiments, the system includes a plurality of spaced-apart engagement structures for securing the syringe when the syringe is attached to at least one sample chamber at a first end of the microfluidic chamber, or when it is attached to at least one sample chamber at a second end of the microfluidic chamber. In some embodiments, each of the plurality of arms further includes an engagement structure configured to engage with a corresponding structure located on each of the plurality of carriages to hold each of the plurality of carriages in a first orientation. In some embodiments, the engagement structure is configured to release and engage the corresponding structure so that the carriage is configured to move between a plurality of orientations about one of a plurality of axes. In some embodiments, the engagement structure includes a magnet, and the corresponding structure includes a magnetically responsive material. Conversely, in some embodiments, the engaging structure includes a magnetically responsive material, and the corresponding structure is a magnet. In certain embodiments, an accelerating or decelerating force is configured to move the carriage between multiple orientations. In some embodiments, the carriage includes a gear assembly, which is configured to move the carriage between multiple orientations. In some embodiments, the carriage includes a centering ratchet, which is configured to move the carriage between multiple orientations.

[0072] A method is provided for processing a sample, the method comprising the steps of: providing a sample to at least one sample chamber; inserting the sample chamber into at least one of a plurality of carriages; securing a syringe to the sample chamber; and rotating a support plate around a first axis. In this method, the sample chamber includes at least one microfluidic channel fluidly connected to a microfluidic tip, one of a plurality of carriages is mounted on a support plate, the support plate is configured to rotate about a first axis, at least one of the plurality of carriages is configured to rotate about a second axis, the second axis is parallel to the first axis, the opening of a syringe is fluidly connected to the sample chamber, the distal end of the syringe plunger is detachably mounted on the plate, the plate is mounted on a motor and configured to be rotatable and move vertically, the vertical movement of the plate causes the plunger in the syringe barrel to descend or rise to discharge or take in a sample for processing into the sample chamber, and the rotation of the support plate about the first axis is configured to drive the sample out of the sample chamber through at least one microfluidic channel in a first direction. In some embodiments, this method further includes lowering the plate vertically to lower the plunger in the syringe barrel to discharge a sample for processing into the sample chamber. In such a method, there is optionally a step of raising the plate vertically so that the plunger rises within the syringe barrel to remove the sample for processing from the sample chamber. In some embodiments, the method further includes rotating at least one of a plurality of carriages in a second orientation around a second axis.

[0073] As described above, a microfluidic chip includes one or more microfluidic pathways. In some embodiments, the pathway includes compression regions (e.g., regions where the pathway walls are closer together than each other) and expansion regions (e.g., regions where the pathway walls are further apart than each other). Figure 16 shows a non-limiting embodiment of a chip having two inlets / outlets and a single expansion flow region positioned between two compression flow regions. In some embodiments, multiple expansion and compression regions are formed in the microfluidic pathways within the chip. Figure 17 shows a non-limiting embodiment in which the chip includes two inlets / outlets and three expansion regions alternately positioned between compression regions. As described above, in some embodiments, the compression / expansion regions promote the activation of specific cells in a biological sample.

[0074] In some embodiments, the carriage includes a spring-like material (e.g., a coil spring, elastomer, etc.) which pushes the carriage floor upward toward a fixed top, holding the microfluidic chip in place during the operation of the device. In some examples, the carriage floor has a wedge-shaped lateral extension on one side (a portion that extends outwards from the plane of the drawing toward the observer) which can be pushed down (e.g., in an arc shape) to move the floor downward to position and / or remove the microfluidic chip.

[0075] In yet another embodiment, the use of processed tissue samples for the treatment of a medical condition is provided. In some embodiments, the medical condition is diabetic ulcer. In some embodiments, the medical condition is improved or benefits from activated cells such as stem cells. In some embodiments, the use of activated stem cells derived from adipose tissue resulting from the processing methods and systems disclosed herein is provided for use in the manufacture of pharmaceuticals for the treatment of a disease or illness.

[0076] In certain embodiments, treatment of a subject (individual) with mechanically processed adipose-derived stem cells (ADSCs) as described herein achieves, for example, one, two, three, four, or more of the effects listed below. (i) Improvement in the severity of the disease or related symptoms (ii) Shortening of the duration of disease-related symptoms (iii) Protection from the progression of the disease or related symptoms (iv) Regression of the disease or related symptoms (v) Protection from the progression or onset of symptoms related to the disease (vi) Protection against recurrence of disease-related symptoms (vii) Reduction in hospitalizations of the target group (viii) Shortening of hospital stay (ix) Increased survival rate of subjects with the disease (x) A decrease in the number of disease-related symptoms (xi) To enhance, improve, supplement, complement, or enhance the preventive or therapeutic effect of another therapy. Administration may be via a variety of routes, including but not limited to intravenous, intra-arterial, subcutaneous, intramuscular, intrahepatic, intraperitoneal, and / or local delivery to the affected tissue.

[0077] [Biological Sample Processing System] A system for processing biological samples is disclosed. Figures 1A-1B, 2A-2B, 23A-23B, 24, 25A-25B, 26A-26B, and 27A-27F illustrate non-limiting embodiments of the system for processing biological samples. In some embodiments, each of the systems for processing biological samples may include a motor coupled to a vertically oriented rotatable chuck. A support plate is attached to or fixed to the rotatable chuck.

[0078] [Overview of Centrifugal Separators] Figure 1 shows one embodiment of a centrifuge for processing biological samples. As will be described in more detail below, the centrifuge includes a base unit with a plurality of carriages (transport units) and at least one tip assembly attached to the base unit. When rotational and / or centrifugal forces are applied to at least one tip assembly, bidirectional flow of the biological sample within the tip assembly is enabled, which can help process the biological sample for subsequent use in treatment.

[0079] Figures 1A and 1B show one embodiment of a centrifuge 100 for processing biological samples. As shown in Figures 1A and 1B, the centrifuge 100 may comprise a base 160 having a central portion 161 and a plurality of lateral arms 163. The central portion 161 is engageable with a component of a motor, which can rotate the base 160. Each of the lateral arms 163 extends radially from the central portion 161 of the base 160. As shown in Figures 1A and 1B, each of the lateral arms 163 may comprise an opening 164 and a plurality of pedestal fitting openings 166 configured to receive and hold the base portion of a carriage 130. In some embodiments, the carriage 130 may be secured to each of the lateral arms 163 using screw caps 170 and screws 132.

[0080] As shown in Figures 1A and 1B, the centrifugal separator 100 can hold a plurality of carriages 130. In some embodiments, the number of carriages 130 is the same as the number of transverse arms 163, but in some embodiments, a smaller number of carriages 130 than the number of transverse arms 163 may be used. As will be described in more detail below, each carriage 130 may comprise a clamp 110, a spring platform 140, and a base platform 150. Each base platform 150 may comprise a first arm 156a and a second arm 156b. The first arm 156a and the second arm 156b engage with the corresponding first and second fitting regions 142a and 142b of the spring platform 140. As will be described in more detail below, the carriage 130 may comprise one or more springs 134 between the base platform 150 and the spring platform 140. In some examples, the carriage 130 may include clamps 110 on which the chip assembly 120 can be secured.

[0081] Each of the carriages 130 can secure a chip assembly 120. As will be described in more detail below, each of the chip assemblies 120 may comprise a microfluidic chip 122, a sample chamber 190a fixed to a first end of the microfluidic chip 122, and a sample chamber 190b fixed to a second end of the microfluidic chip 122.

[0082] Figures 2A and 2B show another embodiment of a centrifuge 200 for processing biological samples. The centrifuge 200 is similar to the centrifuge 100 shown in Figures 1A and 1B. As shown in Figures 2A and 2B, the centrifuge 200 may comprise a base 260 having a central portion 261 and a plurality of transverse arms 263. The central portion 261 is engageable with a component of a motor, which can rotate the base 260. Each of the transverse arms 263 extends radially from the central portion 261 of the base 260. In some embodiments, as shown in Figures 2A and 2B, each of the transverse arms 263 may comprise an opening 264 and a plurality of pedestal fitting openings 266 configured to receive and hold the base portion of a carriage 230. Similar to the centrifuge 100, the carriage 230 of the centrifuge 200 may be fixed to each of the transverse arms 263 using screw caps 270 and screws 232. As will be described in more detail below, each of the lateral arms 263 of the base portion 260 may have a plurality of notches 268 that can engage with a plurality of rotor foot inserts 280.

[0083] As shown in Figures 2A and 2B, the centrifugal separator 200 can hold a plurality of carriages 230. Similar to the carriage 130 described above, the carriage 230 comprises a clamp 210, a spring platform 240, and a base platform 250. Each of the base platforms 250 may comprise a first arm 256a and a second arm 256b. The first arm 256a and the second arm 256b hold the spring platform 240 in corresponding first and second mating regions 242a and 242b. The carriage 230 may comprise one or more springs 234 between the base platform 250 and the spring platform 240. The carriage 230 may also comprise a clamp 210 that can secure a chip assembly 220 to the carriage 230. The clamp 210 may comprise a first mounting flange 212a and a second mounting flange 212b configured to engage with the shoulder portion 236 of the base platform 250. In some embodiments, the first mounting flange 212a and the second mounting flange 212b each have a first opening 214a and a second opening 214b. The first opening 214a and the second opening 214b are aligned with the first opening 236a and the second opening 236b. In some examples, the clamp 210 can be secured to the carriage 230 by inserting multiple screws into holes in the clamp 210 and the base platform 250. As shown in Figures 2A and 2B, the first screw 238a is insertable into the first opening 214a and the first opening 236a, and the second screw 238b is insertable into the second opening 214b and the second opening 236b.

[0084] Each of the carriages 230 can secure a chip assembly 220. Similar to the chip assembly 120, each of the chip assemblies 220 may comprise a microfluidic chip 222, a sample holding chamber 290a fixed to a first end of the microfluidic chip 222, and a sample holding chamber 290b fixed to a second end of the microfluidic chip 222.

[0085] [Overview of Carriage Assembly] Figures 3A and 3B show enlarged views of one embodiment of the carriage 230. Unless otherwise noted, the following description of the carriage 230 and its components is largely applicable to the carriage 130 shown in Figures 1A and 1B.

[0086] As described above, the carriage 230 may comprise a base platform 250, a spring platform 240, and clamps 210 for securing the tip assembly 220. The base platform 250 may comprise a first arm 256a and a second arm 256b that receive the spring platform 240. The base platform 250 may comprise a post portion 258. The post portion 258 extends from the base portion of the base platform 250 and extends through one of the openings 264 of the lateral arm 263 of the base portion 260. In some embodiments, the post portion 258 may comprise a channel having a distal opening 258a and an opening 252 that passes through the upper surface of the base platform 250. The distal opening 258a can receive a screw 232 into the base platform 250 through the post portion 258. In some embodiments, the outer circumferential surface of the post portion 258 may have a shape that allows it to be received by a proximal opening 272 of a screw cap 270. The screw 232 may extend through the tip (distal end) 270b of the screw cap 270, the tip opening 258a of the column 258, and the opening 252, to secure one of the carriages 230 to one of the transverse arms 263. In some embodiments, the screw 232 is secured via a screw fastener (e.g., a nut and bolt) that provides sufficient force to secure the carriage 230 to the transverse arm 263 but allows rotation of the carriage. In some embodiments, as shown in Figure 2B, the bottom surface of the base platform 250 may have a first base 251a and a second base 251b, respectively, which can engage with one of the base fitting openings 266.

[0087] The carriage 230 may also include a spring platform 240. The spring platform 240 may include a first mating region 242a and a second mating region 242b. The first mating region 242a and the second mating region 242b are configured to receive the first arm 256a and the second arm 256b, respectively. Figures 3A and 3B show the spring platform 240 and the base platform 250 as separate components, but in some embodiments, the spring platform 240 and the base platform 250 may be formed integrally.

[0088] The carriage 230 may include a clamp 210 configured to secure the tip assembly 220 to the base platform 250 of the carriage 230. As described above, the clamp 210 may include a first mounting flange 212a having a first opening 214a and a second mounting flange 212b having a second opening 214b. The first mounting flange 212a and the second mounting flange 212b (first opening 214a and second opening 214b) are configured to align with the first opening 236a and the second opening 236b on the shoulder portion 236 of the base platform 250. In some embodiments, the clamp 210 may be secured to the base platform 250 by inserting a first screw 238a into the first opening 214a and the first opening 236a and inserting a second screw 238b into the second opening 214b and the second opening 236b.

[0089] In some embodiments, the upper surface of the base platform 250 positions on it one or more springs (e.g., coil springs, leaf springs, etc.) or another deformable material (e.g., an elastomer) that returns at least substantially to its original position after a force is applied and released. As shown in Figures 3A and 3B, a plurality of springs 234 may be fixed between the spring platform 240 and the base platform 250. As shown in Figure 3A, the base platform 250 may include end spring holders (distal spring holders) 254a and end spring holders (distal spring holders) 254b that can receive the respective base portions of the springs 234. As shown in Figure 2B, the lower surface of the spring platform 240 is provided with proximal spring holders (proximal spring holders) 244a and proximal spring holders (proximal spring holders) 244b. The proximal spring holders 244a and proximal spring holders 244b can receive the respective upper portions of the springs 234. As shown in Figure 3B, when the tip assembly 220 is fixed to the carriage 230, a plurality of springs 234 are compressed between the spring platform 240 and the base platform 250. In some embodiments, the plurality of springs 234 are configured to provide an upward force to the spring platform 240. This upward force can help push the spring platform 240 upward against the tip assembly 220, thereby securing the tip assembly 220 tightly between the spring platform 240 and the clamp 210. The spring tension motion of the spring platform 240 allows the microfluidic tip 222 to be held in place by being pushed upward against the bottom surface of the clamp 210. During operation, when the central part of the device is subjected to a rotational force, and when the carriage is rotated around an axis substantially parallel to the axis of the rotational force applied to the device, the spring pressure allows the microfluidic tip 222 to be held firmly within the carriage 230.

[0090] To insert or remove the microfluidic tip 222 from the carriage 230, the user applies a downward force to the microfluidic tip 222 or the base platform 250, thereby further compressing the spring 234 and increasing the distance between the microfluidic tip 222 and the base of the clamp 210. This may allow the movement (e.g., insertion or removal) of the microfluidic tip 222 relative to the carriage 230. As will be described in more detail below, the clamp 210 may have a relief portion (cushion) or other opening to assist in the removal of the tip.

[0091] [Chip Assembly] Figures 4A to 4B show one embodiment of the chip assembly 220. As described above, unless otherwise specified, the following description of the chip assembly 220 and its components is largely applicable to the chip assembly 120 shown in Figures 1A to 1B.

[0092] The chip assembly 220 may comprise a microfluidic chip 222 fluid-coupled to a first sample holding chamber 290a and a second sample holding chamber 290b. The first sample holding chamber 290a can be fluid-coupled to at least one microfluidic chip via a first port 221a, and the second sample holding chamber 290b can be fluid-coupled to one of the microfluidic chips 222 via a second port 221b. As shown in Figures 4A-4B and 14A-14F, each of the first and second sample holding chambers 290a and 290b can be fluid-coupled to one of the microfluidic chips 222. The sample holding chamber 290a can be fluid-coupled to a first end of the microfluidic chip 222 by engaging a first port 293a of the sample holding chamber 290a with a first port 221a of the microfluidic chip 222. The sample holding chamber 290b can be fluidly connected to the second end of the microfluidic tip 222 by engaging the second port 293b of the sample holding chamber 290b with the second port 221b of the microfluidic tip 222.

[0093] In some embodiments, adapters are interposed between the first port 221a and the first sample holding chamber 290a, and between the second port 221b and the second sample holding chamber 290b. In some embodiments, at least one microfluidic chip comprises a first sample holding chamber and a second sample holding chamber located within the at least one microfluidic chip.

[0094] As will be described in more detail below, the tip assembly 220 allows samples in either or both of the sample holding chambers 290a and 290b to move bidirectionally through the sample holding chamber 290a, the microfluidic tip 222, and the sample holding chamber 290b.

[0095] [Microfluidic Chip] Figures 5A–5C, 7A–7C, 8A–8B, 9–10, 11A–11B, 12A–12B, and 13A–13B illustrate various embodiments of a microfluidic chip for use in a chip assembly 220. The disclosed microfluidic chip may comprise a fluid path having a microfluidic channel. The microfluidic channel has a plurality of expansion and contraction regions along its length. In some embodiments, the plurality of expansion and contraction regions are defined by curved walls within the microfluidic channel. In some examples, the plurality of expansion and contraction regions are defined by inclined walls within the microfluidic channel. In some embodiments, the fluid path may comprise a plurality of branching channels of decreasing size. The branching channels recombine into a plurality of branching channels of increasing size. Each branching channel may comprise a branching point. In some embodiments, the branching point may be a sharp edge. In some embodiments, the microfluidic chip may be used in a pump system. The microfluidic chip can be fluidly connected to a Luer lock and a syringe fluidly connected to the Luer lock.

[0096] Figures 5A to 5C show one embodiment of the microfluidic chip 222. Figure 5A shows a side view of the microfluidic chip 222, Figure 5B shows a top view of the microfluidic chip 222, and Figure 5C shows an isometric three-dimensional view of the microfluidic chip 222. The microfluidic chip 222 may include a body 225. The body 225 has a first port 221a at its first end and a second port 221b at its second end. The first port 221a may include an opening 223a that is fluidly connected to the first end of the fluid path of the microfluidic chip 222. The second port 221b may include an opening 223b that is fluidly connected to the second end of the fluid path of the microfluidic chip 222. The fluid path of the microfluidic chip 222 extends from the openings 223a and 223b. In some embodiments, the fluid path of the microfluidic chip 222 is symmetrical. However, in other embodiments, the fluid path of the microfluidic chip 222 may be asymmetrical or a combination of symmetrical and asymmetrical portions. The disclosure describes the microfluidic chip 222 and its associated fluid path as extending between a "first" end and a "second" end, but the orientation of the microfluidic chip 222 may be interchangeable so that the flow within the chip assembly 220 and within the microfluidic chip 222 can be bidirectional.

[0097] As shown in Figures 5A to 5C, the fluid path of the microfluidic chip 222 may comprise multiple expansion and compression portions. For example, as shown, the fluid path of the microfluidic chip 222 may include a first channel 226a, a first expansion portion 227a, a first compression portion 228a, an expansion region 229, a second compression portion 228b, a second expansion portion 227b, and a second channel 226b.

[0098] In some embodiments, the fluid path of the microfluidic chip 222 includes a first channel 226a that contracts, and the first channel 226a is closest to the opening 223a of the first port 221a. The first port 221a may have a length, width, or diameter of about 0.5 mm to about 4 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. The first channel 226a may be expanded and increased in width, height, and / or diameter so that the first channel 226a is fluidly connected to the first expansion portion 227a. As shown in Figure 5C, the first expansion portion 227a is three-dimensional (three-dimensional) teardrop-shaped. The first expansion portion 227a may have a greater width, height, and / or diameter than the first channel 226a. The first expansion portion 227a may be in the range of about 0.3 mm to about 5 mm. In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, about 4.0 mm, about 4.5 mm, or about 5.0 mm, or about 0.3 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, about 3.5 to 4.0 mm, about 4.0 to 4.5 mm, or about 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The first expansion portion 227a may be fluidly connected to the first compression portion 228a. The fluid path may contract in the first compression portion 228a, which has a smaller width, height, and / or diameter than the first expansion portion 227a. The first compression portion 228a may have a width, height, and / or diameter in the range of approximately 0.3 mm to approximately 3 mm.In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, or about 3.0 mm, or about 0.3 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, or about 2.5 to 3.0 mm, or any value within these ranges including upper and lower limits. The fluid path may be expanded and increased in width, height, and / or diameter so that the first compression portion 228a is fluidly connected to the second expansion region 229. The second expansion region 229 may be in the range of about 0.3 mm to about 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.3 mm, approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.3 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. In some embodiments, the second expansion region 229 is a spherical or elliptical expansion having a greater length, width, and / or diameter than the first compression portion 228a. The second expansion portion 229 is fluidly connected to the second compression portion 228b. The fluid path is compressed at the second compression section 228b such that the width, height, and / or diameter of the compression section 228b is smaller than the width, height, and / or diameter of the second expansion section 227b. The length, width, and / or diameter of the second compression section 228b may be in the range of about 0.3 mm to about 3 mm. In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, or about 3.0 mm, or about 0.3 to 1 mm, about 1 mm to 1.5 mm, about 1.5 to 2.0 mm, or about 2.5 to 3.0 mm, or any value within these ranges including upper and lower limits. The second compression section 228b is fluidly connected to the third expansion section 227c, and the fluid path expands to the third expansion section 227b.The third expansion portion 227b expands such that its width, height, and / or diameter are greater than the width, height, and / or diameter of the second compression portion 228b. The third expansion portion 227b may be in the range of about 0.3 mm to about 5 mm. In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, about 4.0 mm, about 4.5 mm, or about 5.0 mm, or about 0.3 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, about 3.5 to 4.0 mm, about 4.0 to 4.5 mm, or about 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The third expansion portion 227b may have a three-dimensional (three-dimensional) teardrop or other arched (bow-shaped) shape. The third expansion portion 227b may be fluidly connected to the second channel 226b and may contract to the second channel 226b. The second channel 226b may have a width, height, and / or diameter smaller than the width, height, and / or diameter of the expansion portion 227b. The second channel 226b may have a size in the range of approximately 0.5 mm to approximately 4 mm in length, width, and / or diameter. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, or approximately 4.0 mm, or approximately 0.5 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, or approximately 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. In some embodiments, the fluid path of the microfluidic chip 222 is symmetrical, but in other embodiments, the fluid path of the microfluidic chip 222 may be asymmetrical, or a combination of symmetrical and asymmetrical portions. In some embodiments, the ratio of the first channel 226a to the extension portion 227a may be in the range of 1:1 to 1:17. In some examples, the ratio of the extension portions 227b to extension portions 227b may be in the range of 1:1 to 1:17.In some embodiments, the ratio of the compressed portion 228a or compressed portion 228b to the expanded region 229 may be in the range of 1:1 to 1:17.

[0099] Figures 6A to 6D show enlarged views of the fluid pathways in the microfluidic chip 222. Figure 6A shows an enlarged view of the port 221 (i.e., the first port 221a and / or the second port 221b) and opening 223 (i.e., opening 223a and / or opening 223b) at either end of the microfluidic chip 222. In some embodiments, port 221 may be a Luer lock adapter used with a syringe, or other Luer lock connector. In some embodiments, port 221 may instead be a slip Luer, or any other type of Luer connector. The opening 223 of port 221 is fluid-connected to the inlet channel 226 (i.e., the first channel 226a and the second channel 226b). The inlet channel 226 may allow for an increase in the velocity of the fluid moving through the inlet channel 226, thereby creating laminar flow. Figure 6B shows the extension portion 227 of the first extension portion 227a and the third extension portion 227c. In some embodiments, the expansion portion 227 may have a “teardrop” shape. The expansion portion 227 can reduce the velocity and enable turbulent flow, such as mixing and vortexing the fluid flowing through the fluid path of the microfluidic tip 222. Figure 6C shows the compression zone 228 of compression portions 228a and 228b. The compression zone can change the flow to a laminar flow profile, which again helps to increase the fluid velocity. The compression zone 228 can provide high shear forces due to the interaction of the sample with the walls of the fluid path. Figure 6D shows a second expansion region 229. As described above, the expansion region 229 may have a spherical and / or elliptical shape. The expansion region 229 can reduce the velocity of the fluid by inducing turbulence and vortices. Increasing or decreasing the velocity of the fluid flow may help to break down tissues such as adipose tissue for reinjection.

[0100] The microfluidic chips disclosed above can be manufactured by a variety of methods. For example, microfluidic chips can be manufactured using additive manufacturing, subtractive manufacturing, injection molding, resin molding, urethane casting, 3D printing, or layer lamination. Microfluidic chips can be made from a variety of materials, including, for example, plastics (polycarbonate, acrylic, etc.), polyurethane, or metals (aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0101] Figures 7A to 7C show one embodiment of the microfluidic chip 322. Figure 7A shows a top view of the microfluidic chip 322. Figure 7B shows a side view of the microfluidic chip 322 in which the second expansion region 329 has a hemispherical shape. Figure 7C shows a side view of the microfluidic chip 322 in which the second expansion region 329b forms a flat, channel-shaped expansion. The microfluidic chip 322 serves the same purpose as the microfluidic chip 222 described above, except that the shapes of the expansion region and the compression region are different. The microfluidic chip 322 may include a body 325, the body 325 having at least a first port 321a at a first end of the body 325 and at least a second port 321b at a second end of the body 325. The first port 321a may include an opening 323a that is fluidly connected to the first end of the fluid path of the microfluidic chip 322. The second port 321b may include an opening 323b fluid-connected to the second end of the fluid path of the microfluidic chip 322. The fluid path of the microfluidic chip 322 may extend from openings 323a and 323b. In some embodiments, the fluid path of the microfluidic chip 322 is symmetrical. In other embodiments, the fluid path of the microfluidic chip 322 may be asymmetrical, or a combination of symmetrical and asymmetrical portions. Although this disclosure discusses the microfluidic chip 322 and its associated fluid path as extending between a “first” end and a “second” end, the orientation of the microfluidic chip 322 is interchangeable as long as the flow within the chip assembly 320 and within the microfluidic chip 322 can be bidirectional.

[0102] As shown in Figures 7A to 7C, the fluid path of the microfluidic chip 322 may include multiple expansion and compression portions. For example, as shown, the fluid path of the microfluidic chip 322 may include a first channel 326a, a first expansion portion 327a, a first compression portion 328a, a second expansion region 329, a second compression portion 328b, a third expansion portion 327b, and a second channel 326b.

[0103] In some embodiments, the fluid path of the microfluidic chip 322 includes a first channel 326a that contracts, and the first channel 326a is closest to the opening 323a of the first port 321a. The first port 321a may have a length, width, or diameter of about 0.5 mm to about 4 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. The first channel 326a may be expanded and increased in width, height, and / or diameter so that the first channel 326a is fluidly connected to the first expansion portion 327a. In some embodiments, the shape of the first expansion portion 327a may be semi-teardrop or other arched (bow-shaped). The first expansion portion 327a may have a greater width, height, and / or diameter than the first channel 326a. The first expansion portion 327a may range from about 0.5 mm to about 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.5 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The first expansion portion 327a may be fluidly connected to the first compression portion 328a. The fluid path contracts at the first compression portion 328a, and the first compression portion 328a has a smaller width, height, and / or diameter than the first expansion portion 327a. The first compression portion 328a may have a width, height, and / or diameter in the range of approximately 0.3 mm to approximately 3 mm.In some embodiments, the length, width, or diameter may be approximately 0.3 mm, 0.5 mm, 1 mm, 1.5 mm, 2.0 mm, 2.5 mm, or 3.0 mm, or approximately 0.3 to 1 mm, 1 to 1.5 mm, 1.5 to 2.0 mm, or 2.5 to 3.0 mm, or any value within these ranges, including upper and lower limits. The fluid path may be expanded and increased in width, height, and / or diameter so that the first compression portion 328a is fluidly connected to the second expansion region 329. As shown in Figure 7B, the second expansion region 329 has a hemispherical or semi-elliptical expansion shape having a greater length, width, and / or diameter than the first compression portion 328a. The second expansion region 329 may be in the range of approximately 0.3 mm to approximately 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.3 mm, approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.3 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. As shown in Figure 7C, the second expansion region 329 may be in the shape of a flat, channel-shaped expansion. The second expansion region 329 is fluidly connected to the compression portion 328b. The fluid path may be compressed in the second compression portion 328b such that the width, height, and / or diameter of the compression portion 328b is smaller than the width, height, and / or diameter of the second expansion portion 328b. The length, width, and / or diameter of the second compression portion 328b may be in the range of about 0.3 mm to about 3 mm. In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, or about 3.0 mm, or about 0.3 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, or about 2.5 to 3.0 mm, or any value within these ranges including upper and lower limits.The second compression portion 328b is fluidly connected to the third expansion portion 327b such that the fluid path expands in the third expansion portion 327b. The third expansion portion 327b may have a width, height, and / or diameter greater than the width, height, and / or diameter of the second compression portion 328b. The first expansion portion 327a may be in the range of about 0.5 mm to about 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.5 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The third expansion portion 327b may have a three-dimensional (three-dimensional) teardrop or other arched (bow-shaped) shape. The third expansion portion 327b may be fluidly connected to contract in the second channel 326b. The second channel 326b may have a width, height, and / or diameter smaller than the width, height, and / or diameter of the third extension portion 327b. The second channel 326b may have a length, width, and / or diameter in the size range of about 0.5 mm to about 4 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. In some embodiments, the microfluidic tip 322 is structured as if the microfluidic tip 222 described above were cut in half. In some examples, the microfluidic tip 322 is symmetrical. In other embodiments, the microfluidic chip 322 may be asymmetrical or a combination of symmetrical and asymmetrical portions. In some embodiments, the ratio of the first channel 326a to the extension portion 327a may range from about 1:1 to about 1:10.In some examples, the ratio of the second channel 326b to the second channel 326b may be in the range of about 1:1 to about 1:10. In some embodiments, the ratio of the compressed portion 328a or compressed portion 328b to the expanded region 329 may be in the range of about 1:1 to about 1:17.

[0104] Figures 8A and 8B show another embodiment of the microfluidic chip 422 in which the fluid path includes multiple expansion and compression portions. Figure 8A shows a top view of the microfluidic chip 422, and Figure 8B shows a side view of the microfluidic chip 422. The microfluidic chip 422 serves the same purpose as the microfluidic chips 222 and 322 described above, except that the shapes of the expansion and compression regions of the fluid path differ. The microfluidic chip 422 may include a body 425, the body 425 having a first port 421a at a first end of the microfluidic chip 422 and a second port 421b at a second end of the body 425. The first port 421a may include an opening 423a fluid-connected to the first end of the fluid path of the microfluidic chip 422. The second port 421b may include an opening 423b fluid-connected to the second end of the fluid path of the microfluidic chip 422. The fluid path of the microfluidic chip 422 may extend from opening 423a to opening 423b. In some embodiments, the fluid path of the microfluidic chip 422 is symmetrical. In other embodiments, the fluid path of the microfluidic chip 422 may be asymmetrical, or a combination of symmetrical and asymmetrical portions. Although this disclosure discusses the microfluidic chip 422 and its associated fluid path as extending between a “first” end and a “second” end, the orientation of the microfluidic chip 422 is interchangeable, as long as the flow within the chip assembly and within the microfluidic chip 422 can be bidirectional.

[0105] As shown in Figures 8A and 8B, the fluid path of the microfluidic chip 422 may include multiple expansion and compression portions. For example, as shown, the fluid path of the microfluidic chip 422 may include a first channel 426a, a first expansion portion 427a, a first compression portion 428a, a second expansion region 429, a second compression portion 428b, a third expansion portion 427b, and a second channel 426b.

[0106] In some embodiments, the fluid path of the microfluidic chip 422 comprises a first channel 426a that contracts, the first channel 426a being closest to the opening 423a of the first port 421a. The first port 421a may have a length, width, and / or diameter between about 0.5 mm and about 4 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. The first channel 426a may be expanded and increased in width, height, and / or diameter so that the first channel 426a is fluidly connected to the first expansion portion 427a. In some embodiments, the shape of the first expansion portion 427a may be semi-teardrop shaped. The first expansion portion 427a may have a greater width, height, and / or diameter than the first channel 426a. The first expansion portion 427a may be in the range of about 0.5 mm to about 5 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, about 4.0 mm, about 4.5 mm, or about 5.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, about 3.5 to 4.0 mm, about 4.0 to 4.5 mm, or about 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The first expansion portion 427a may be fluidly connected to the first compression portion 428a. The fluid path contracts in the first compression portion 428a, and the first compression portion 428a has a smaller width, height, and / or diameter than the first expansion portion 427a. The first compression portion 428a may have a width, height, and / or diameter in the range of approximately 0.3 mm to approximately 3 mm.In some embodiments, the length, width, or diameter may be approximately 0.3 mm, 0.5 mm, 1 mm, 1.5 mm, 2.0 mm, 2.5 mm, or 3.0 mm, or approximately 0.3 to 1 mm, 1 mm to 1.5 mm, 1.5 to 2.0 mm, or 2.5 to 3.0 mm, or any value within these ranges, including upper and lower limits. The fluid path may be expanded and increased in width, height, and / or diameter so that the first compression portion 428a is fluidly connected to the second expansion region 429. As shown in Figure 8A, the second expansion region 429 has a D-shape having a greater length, width, and / or diameter than the first compression portion 428a. The second expansion region 429 may be in the range of approximately 0.3 mm to approximately 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.3 mm, 0.5 mm, 1 mm, 1.5 mm, 2.0 mm, 2.5 mm, 3.0 mm, 3.5 mm, 4.0 mm, 4.5 mm, or 5.0 mm, or between approximately 0.3 and 1 mm, 1 to 1.5 mm, 1.5 to 2.0 mm, 2.5 to 3.0 mm, 3.0 to 3.5 mm, 3.5 to 4.0 mm, 4.0 to 4.5 mm, or 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. As shown in Figure 8A, the D-shape of the second extension region 429 has a right-angle end on the first side and an angled second side to provide more turbulence and vortex. The second extension region 429 may be fluidly connected to the second compression region 428b. The fluid path may be compressed in the second compression section 428b such that the width, height, and / or diameter of the second compression section 428b is smaller than the width, height, and / or diameter of the second compression section 428b. The length, width, and / or diameter of the second compression section 428b may be in the range of about 0.3 mm to about 3 mm. In some embodiments, the length, width, or diameter may be about 0.3 mm, about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, or about 3.0 mm, or about 0.3 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, or about 2.5 to 3.0 mm, or any value within these ranges including upper and lower limits.The second compression section 428b is fluidly connected to the third expansion section 427b such that the fluid path expands in the third expansion section 427b. The third expansion section 427b may have a width, height, and / or diameter greater than the width, height, and / or diameter of the second compression section 428b. The third expansion section 427b is in the range of about 0.5 mm to about 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.5 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The third expansion portion 427b may have a three-dimensional teardrop shape. The third expansion portion 427b may be fluidly connected to the second channel 426b so as to contract in the second channel 426b. The second channel 426b may have a width, height, and / or diameter smaller than the width, height, and / or diameter of the third extension portion 427b. The second channel 426b may have a length, width, and / or diameter in the size range of about 0.5 mm to about 4 mm. In some embodiments, such length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. In some embodiments, the microfluidic chip 422 is structured as if the microfluidic chip 222 described above were cut in half. In some examples, the microfluidic chip 422 is symmetrical. In other embodiments, the microfluidic chip 422 may be asymmetrical or a combination of symmetrical and asymmetrical portions. In some embodiments, the ratio of the first channel 426a to the first extension portion 427a may range from about 1:1 to about 1:10.In some examples, the ratio of the second channel 426b to the third extension portion 427b may be in the range of about 1:1 to about 1:10. In some embodiments, the ratio of the compression portion 428a or compression portion 428b to the second extension region 429 may be in the range of about 1:1 to about 1:17.

[0107] Figure 9 shows another embodiment of the microfluidic chip 522. As shown in Figure 9, the microfluidic chip 522 comprises a fluid path, which includes a microfluidic channel formed by a pair of tapered regions joined at a constriction in its central region. In some embodiments, the tapered region includes a continuous tapered region. In some examples, the tapered region includes a stepped tapered region. Figure 9 shows a top view of the microfluidic chip 522. The microfluidic chip 522 serves the same purpose as the microfluidic chips 222, 322, and 422 described above. The microfluidic chip 522 may also comprise a body 525, the body 525 having a first port 521a at a first end of the microfluidic chip 522 and a second port 521b at a second end of the body 525. The first port 521a may comprise an opening 523a fluid-connected to the first end of the fluid path of the microfluidic chip 522. The second port 521b may include an opening 523b that is fluidly connected to the second end of the fluid path of the microfluidic chip 522. The fluid path of the microfluidic chip 522 may extend from opening 523a to opening 523b. In some embodiments, the fluid path of the microfluidic chip 522 is symmetrical. In other embodiments, the fluid path of the microfluidic chip 522 may be asymmetrical, or a combination of symmetrical and asymmetrical portions. Although this disclosure discusses the microfluidic chip 522 and its associated fluid path as extending between a “first” end and a “second” end, the orientation of the microfluidic chip 522 is interchangeable as long as the flow within the chip assembly and within the microfluidic chip 522 can be bidirectional.

[0108] As shown in Figure 9, the fluid path of the microfluidic chip 522 may comprise multiple expansion and compression sections. As shown in Figure 9, the fluid path of the microfluidic chip 522 may include a first channel 526a, a first expansion section 527a, a compression section 528, a second expansion section 527b, and a second channel 526b. As shown in Figure 9, the fluid path of the microfluidic chip 522 may have an hourglass shape that provides a smooth increase in the velocity of the sample. This helps maintain laminar flow and slowly increase the shear stress on the sample as it interacts with the walls of the apparatus. In some embodiments, the first channel 526a (and the second channel 526b) provides an initial fluid velocity that enables laminar flow. The rhombic shape of the first expansion section 527a (and the second expansion section 527b) allows for deceleration of the sample, thereby generating turbulence and vortexes that mix the fluid. The compression section 528 can increase the fluid velocity and thus increase the shear force. The size of the first channel 526a and the second channel 526b may be about 0.5 mm to about 4 mm in length, width, and / or diameter. In some embodiments, such length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. The size of the first expansion section 527a and the second expansion section 527b may be larger than the width, length, and / or diameter of the first channel 526a and the second channel 526b. The first extension portion 527a and the second extension portion 527b may be in the range of approximately 0.3 mm to approximately 5 mm.In some embodiments, the length, width, or diameter may be approximately 0.3 mm, approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, approximately 3.0 mm, approximately 3.5 mm, approximately 4.0 mm, approximately 4.5 mm, or approximately 5.0 mm, or approximately 0.3 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, approximately 2.5 to 3.0 mm, approximately 3.0 to 3.5 mm, approximately 3.5 to 4.0 mm, approximately 4.0 to 4.5 mm, or approximately 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The size of the compression portion 528 may be approximately 0.3 mm to approximately 3 mm. In some embodiments, the length, width, or diameter may be approximately 0.3 mm, approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, or approximately 3.0 mm, or approximately 0.3 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, or approximately 2.5 to 3.0 mm, or any value within these ranges, including upper and lower limits. In some embodiments, the first port 521a and the second port 521b are Luer locks. In some embodiments, the first port 521a and the second port 521b may be other Luer adapters, such as slip Luers. In some embodiments, the ratio of the compressed portion 528 to the first extended portion 527a, and / or the ratio of the compressed portion 528 to the second extended portion 527b may be in the range of approximately 1:1 to approximately 1:17.

[0109] Figure 10 shows another embodiment of the microfluidic chip 622, in which the fluid path includes multiple expansion and compression portions. Figure 10 shows a top view of the microfluidic chip 622. The microfluidic chip 622 serves the same purpose as the microfluidic chips 222, 322, 422, and 522 described above, except that the shape of the expansion and compression regions of the fluid path differs. The microfluidic chip 622 may include a body 625, the body 625 having a first port 621a at a first end of the microfluidic chip 622 and a second port 621b at a second end of the body 625. The first port 621a may include an opening 623a fluid-connected to the first end of the fluid path of the microfluidic chip 622. The second port 621b may include an opening 623b fluid-connected to the second end of the fluid path of the microfluidic chip 622. The fluid path of the microfluidic chip 622 may extend from opening 623a to opening 623b. In some embodiments, the fluid path of the microfluidic chip 622 is symmetrical. In other embodiments, the fluid path of the microfluidic chip 622 may be asymmetrical, or a combination of symmetrical and asymmetrical portions. Although this disclosure discusses the microfluidic chip 622 and its associated fluid path as extending between a “first” end and a “second” end, the orientation of the microfluidic chip 622 is interchangeable as long as the flow within the chip assembly and within the microfluidic chip can be bidirectional.

[0110] As shown in Figure 10, the fluid path of the microfluidic chip 622 may comprise multiple expansion and compression sections. For example, as shown in Figure 10, the fluid path of the microfluidic chip 622 may include a first channel 626a, a first expansion section 627a, a compression section 628, a second expansion section 627b, and a second channel 626b. The first channel 626a, the first expansion section 627a, the compression section 628, the second expansion section 627b, and the second channel 626b are fluid-connected.

[0111] In the microfluidic chip 622 shown in Figure 10, the fluid path has a sharper hourglass shape to provide increased turbulence and eddy flow. The first channel 626a and the second channel 626b may provide the initial velocity of the sample to enable laminar flow. In some embodiments, the size of the first channel 626a and the second channel 626b may be in the range of about 0.5 mm to about 4 mm in length, width, and / or diameter. In some embodiments, such length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, about 3.0 mm, about 3.5 mm, or about 4.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, about 2.5 to 3.0 mm, about 3.0 to 3.5 mm, or about 3.5 to 4.0 mm, or any value within these ranges including upper and lower limits. The first expansion portion 627a and the second expansion portion 627b may have a rhomboid shape that provides fluid deceleration. This can generate turbulence and vortex for mixing the fluid sample. In some embodiments, the size of the rhomboid first expansion portion 627a and the rhomboid second expansion portion 627b is greater than the width, length, and / or diameter of the first channel 626a and the second channel 626b. The first expansion portion 627a and the second expansion portion 627b may range from about 0.3 mm to about 5 mm. In some embodiments, the length, width, or diameter may be approximately 0.3 mm, 0.5 mm, 1 mm, 1.5 mm, 2.0 mm, 2.5 mm, 3.0 mm, 3.5 mm, 4.0 mm, 4.5 mm, or 5.0 mm, or approximately 0.3 to 1 mm, 1 to 1.5 mm, 1.5 to 2.0 mm, 2.5 to 3.0 mm, 3.0 to 3.5 mm, 3.5 to 4.0 mm, 4.0 to 4.5 mm, or 4.5 to 5.0 mm, or any value within these ranges including upper and lower limits. The compression section 628 can increase the velocity of the sample to increase the shear force applied to the fluid sample. Similarly, the sharp angles on both sides of the compression section 628 can provide turbulence and eddies. In some embodiments, the compression portion 628 may have a width, length, and / or diameter in the range of about 0.3 mm to about 3 mm.In some embodiments, the length, width, or diameter may be approximately 0.3 mm, approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, or approximately 3.0 mm, or approximately 0.3 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, or approximately 2.5 to 3.0 mm, or any value within these ranges including upper and lower limits. In some embodiments, the first port 621a and the second port 621b are Luer locks. In some embodiments, the first port 621a and the second port 621b may be other Luer adapters such as slip Luers. In some embodiments, the ratio of the compressed portion 628 to the expanded portion 627a and / or the ratio of the compressed portion 628 to the expanded portion 627b may be in the range of approximately 1:1 to approximately 1:17.

[0112] Figures 11A–11B, 12A–12B, and 13A–13B illustrate multiple embodiments of a microfluidic chip having a three-dimensional channel structure. The three-dimensional channel structures in Figures 11A–11B, 12A–12B, and 13A–13B have fluid pathways that compress / contract and expand in multiple stages. This compression and expansion allows for the generation of turbulence useful for processing biological samples. For example, the device may be used to mechanically dissipate adipose tissue to reduce its size for applications such as orthopedic surgery, arthroscopic surgery, neurosurgery, gastrointestinal and related organ surgery, urological surgery, general surgery, and gynecological surgery. The disclosed microfluidic chip may be manufactured using 3D printing, additive manufacturing, subtractive manufacturing, or injection molding. The materials used to print or manufacture the device are biocompatible and sterilizable. The microfluidic chip includes an inlet and an outlet, which may be Luer locks, screws, or any other form that allows for fluid sealing. Once the seal is formed, a fluid path is created. As the sample passes through the inlet, it interacts with a compression region smaller than the diameter of the inlet, generating mechanical stress. The diameter of this region may range from about 0.5 mm to about 3.0 mm. In some embodiments, the length, width, or diameter may be about 0.5 mm, about 1 mm, about 1.5 mm, about 2.0 mm, about 2.5 mm, or about 3.0 mm, or about 0.5 to 1 mm, about 1 to 1.5 mm, about 1.5 to 2.0 mm, or about 2.5 to 3.0 mm, or any value within these ranges, including upper and lower limits. After passing through the compression region, the fluid path expands in an expansion region. In some embodiments, the expansion region may generate turbulence and vortexes. In some examples, the expansion region may provide controlled, precise laminar flow. The diameter of the expanded area can range from approximately 4 mm to approximately 25 mm.In some embodiments, the diameter of the expansion region may be approximately 4 mm, 6 mm, 8 mm, 10 mm, 12 mm, 14 mm, 16 mm, 18 mm, 20 mm, 22 mm, 24 mm, or 25 mm, or approximately 4-6 mm, 6-8 mm, 8-10 mm, 10-12 mm, 12-14 mm, 14-16 mm, 16-18 mm, 18-20 mm, 20-22 mm, 22-24 mm, or 24-25 mm, or any value within these ranges including upper and lower limits. After passing through the expansion region, the next contraction region places a large shear stress on the sample being processed. As described above, the diameter of the compression region may be in the range of approximately 0.5 mm to approximately 3.0 mm. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, approximately 1 mm, approximately 1.5 mm, approximately 2.0 mm, approximately 2.5 mm, or approximately 3.0 mm, or approximately 0.5 to 1 mm, approximately 1 to 1.5 mm, approximately 1.5 to 2.0 mm, or approximately 2.5 to 3.0 mm, or any value within these ranges, including upper and lower limits. In some embodiments, the shape of the expansion and contraction regions may be circular, elliptical, square, and rectangular, along with the overall design, but other shapes may be used.

[0113] Figures 11A and 11B show one embodiment of a microfluidic chip 722 having a single extended region. Figure 11A shows a side view of the microfluidic chip 722, and Figure 11B shows a perspective view of the microfluidic chip 722. The fluid flow through the fluid path of the microfluidic chip 722 is described in more detail above. The microfluidic chip 722 includes a first port 721a at a first end of the microfluidic chip 722 and a second port 721b at a second end of the microfluidic chip 722. The first port 721a may include an opening 723a that is fluid-connected to the first end of the fluid path of the microfluidic chip 722. The second port 721b may include an opening 723b that is fluid-connected to the second end of the fluid path of the microfluidic chip 722. The fluid path of the microfluidic chip 722 may extend from opening 723a to opening 723b. In some embodiments, the fluid path of the microfluidic chip 722 is symmetrical. As shown in Figures 11A and 11B, the fluid path of the microfluidic chip 722 may include a fluid-connected first channel 726a, an extension 727a, and a second channel 726b. As described above, in the first channel 726a and the second channel 726b, the fluid sample may be subjected to considerable shear stress. In the extension 727a, turbulence and vortex motion may occur in the fluid sample. In some embodiments, the ratio of the first channel 726a to the extension 727a and / or the ratio of the second channel 726b to the extension 727a is in the range of about 1:1 to about 1:83.

[0114] Figures 12A and 12B show one embodiment of a microfluidic chip 822 having two extended regions and a compressed region between the two extended regions. Figure 12A shows a side view of the microfluidic chip 822, and Figure 12B shows a perspective view of the microfluidic chip 822. The fluid flow through the fluid path of the microfluidic chip 822 is described in more detail above. The microfluidic chip 822 may include a first port 821a at a first end of the microfluidic chip 822 and a second port 821b at a second end of the microfluidic chip 822. The first port 821a may include an opening 823a that is fluidly connected to the first end of the fluid path of the microfluidic chip 822. The second port 821b may include an opening 823b that is fluidly connected to the second end of the fluid path of the microfluidic chip 822. The fluid path of the microfluidic chip 822 may extend from opening 823a to opening 823b. In some embodiments, the fluid path of the microfluidic chip 822 is symmetrical. As shown in Figures 12A–12B, the fluid path of the microfluidic chip 822 may include a fluid-connected first channel 826a, a first expansion portion 827a, a compression portion 828, a second expansion portion 827b, and a second channel 826b. As described above, the fluid sample in the first channel 826a, the compression portion 828, and the second channel 826b may be subjected to considerable shear stress. Turbulence and vortexing may occur in the fluid sample in the first expansion portions 827a and 827b. In some embodiments, the ratio of channels 826a, 826b to expansion portions 827a, 827b may range from about 1:1 to about 1:83. In some examples, the ratio of the compression portion 828 to expansion portions 827a, 827b may range from about 1:1 to about 1:83.

[0115] Figures 13A and 13B show one embodiment of a microfluidic chip 922 having three expansion regions and two compression regions between the three expansion regions. Figure 13A shows a side view of the microfluidic chip 922, and Figure 13B shows a perspective view of the microfluidic chip 922. The fluid flow through the fluid path of the microfluidic chip 922 is described in more detail above. The microfluidic chip 922 may include a first port 921a at a first end of the microfluidic chip 922 and a second port 921b at a second end of the microfluidic chip 922. The first port 921a may include an opening 923a that is fluid-connected to the first end of the fluid path of the microfluidic chip 922. The second port 921b may include an opening 923b that is fluid-connected to the second end of the fluid path of the microfluidic chip 922. The fluid path of the microfluidic chip 922 may extend from opening 923a to opening 923b. In some embodiments, the fluid path of the microfluidic chip 922 is symmetrical. As shown in Figures 13A to 13B, the fluid path of the microfluidic chip 922 may include a fluid-connected first channel 926a, a first expansion portion 927a, a first compression portion 928a, a second expansion portion 927b, a second compression portion 928b, a third expansion portion 927c, and a second channel 926b. As described above, the fluid sample in the first channel 926a, the first compression portion 928a, the second compression portion 928b, and the second channel 926b may be subjected to considerable shear stress. In the first expansion portion 927a, the second expansion portion 927b, and the third expansion portion 927c, turbulence and vortexes may occur in the fluid sample. In some embodiments, the ratio of channels 926a, 926b to extension portions 927a, 927b, 927c may range from about 1:1 to about 1:83. In some examples, the ratio of compression portions 928a, 928b to extension portions 927a, 927b, 927c may range from about 1:1 to about 1:83.

[0116] The examples of microfluidic chips shown in Figures 11A–11B, 12A–12B, and 13A–13B are illustrative and not limiting. Microfluidic chips may include any number of expansion and / or compression regions to achieve the intended results.

[0117] [Sample Chamber] Figures 14A to 14B show a sample chamber 290 capable of holding a sample for storage during processing, or before or after processing (e.g., during transport). The sample chamber 290 comprises a body 291a, the body 291a having a port 293 located at its first end. The port 293 of the sample chamber 290 may form a Luer lock connector for connection to a syringe for sample injection and extraction. The port 293 may also be fluidly connected to a microfluidic tip 222. As shown in Figures 4A to 4B, the port 293 of the sample chamber 290 may include an engagement surface 298 configured to engage with either the first port 221a or the second port. The port 293 of the sample chamber 290 may include an opening 294 that allows a sample to flow into or out of the sample chamber 290. The size of the opening 294 may range from about 0.5 mm to about 3 mm. In some embodiments, the length, width, or diameter may be approximately 0.5 mm, 1 mm, 1.5 mm, 2.0 mm, 2.5 mm, or 3.0 mm, or approximately 0.5–1 mm, 1–1.5 mm, approximately 1.5–2.0 mm, or 2.5–3.0 mm, or any value within these ranges, including upper and lower limits. Port 293 may be a different type of Luer connector, such as a slip Luer. As shown in Figure 14A, port 293 of the sample chamber 290 may be located near the bottom of the sample chamber 290 to facilitate the flow of fluid to the microfluidic tip 222. In some embodiments, port 293 may be located higher up in the sample chamber 290. As shown in Figure 14A, the sample chamber 290 may include a vent hole 292a at the top of the sample chamber 290, which allows for smooth fluid flow when the tip and chamber are fully assembled and prevents the formation of a vacuum within the sample chamber 290 and the fluid-connected microfluidic tip 222. The body 291a of the sample chamber 290 may include tapered surfaces 291b and tapered surfaces 291b located on either side of the port 293.In some embodiments, the tapered surface 291b may provide a smoother flow and complete discharge of the fluid sample in the sample chamber 290 during centrifugation and extraction. In some embodiments, the tapered surface 291b may have an angle greater than 25 degrees from the front of the chamber.

[0118] Figures 14C to 14D show the interior of the sample chamber 290. The sample chamber 290 includes a vent channel 292b, a vent chamber 292c, and a retaining cavity 296 fluid-connected to the vent hole 292a. The internal retaining cavity 296 can hold the sample during use. The volume of the retaining cavity 296 may range from approximately 1 mL to approximately 300 mL. The sample chamber 290 may include an arrow-shaped vent chamber 292c at the top of the sample chamber 290. The vent chamber 292c can capture any sample entering the vent channel and prevent the sample from leaving the sample chamber 290. During use, the fluid sample in the vent chamber 292c is subjected to centrifugal force. Due to the arrow shape of the vent chamber 292c, the fluid sample is pushed into the corner of the vent chamber 292c during use, away from the upper vent hole 292a. When the direction of centrifugal force is reversed, the fluid sample may be pushed back into the retaining cavity 296 through the vent channel 292b. The volume of the vent chamber 292c may range from about 0.5 mL to about 50 mL. In some embodiments, a Luer adapter may be added to the back of the chamber as an alternative extraction and filling point. In some embodiments, the sample chamber 290 may include a carriage-fitting ridge 297 which may be added to be fixed to and engage with a portion of the carriage.

[0119] [filter] Figures 15A–15B show one embodiment of a filter that may be placed between the sample chamber and the microfluidic tip in the tip assembly. As shown in Figures 16A–16B, a filter may be placed upstream or before the microfluidic tip to filter the sample to prevent clogging of the microfluidic tip. As will be described in more detail below, the filter may include a mesh configured to cut or pulverize tissue or tissue fragments so that the sample can pass through the microfluidic tip without clogging. Cutting or pulverizing the sample may generate macroscopic aggregates for the purpose of microfluidic shearing in the microfluidic tip. As shown in Figures 16A–16B, a filter may be placed downstream or behind the microfluidic tip so that only samples of a specific size can pass out of the instrument for collection.

[0120] Figures 15A and 15B show one embodiment of the filter 1000. The filter 1000 may include a first body portion 1100 and a second body portion 1200. The first body portion 1100 may include a first side having a port 1110. The port 1110 may include an engagement surface 1120 configured to engage with a first or second port of any of the disclosed microfluidic chips. The engagement surface 1120 of the port 1110 may be configured to engage with a port of the sample chamber 290 (i.e., port 293). The port 1110 may include a channel 1140, which extends from a first opening 1130 on the first side of the first body portion 1100 to a second opening 1150 on the second side of the first body portion 1100.

[0121] The second body portion 1200 may include a first side having a port 1210. The port 1210 may include an engagement surface 1220 configured to engage with either the first or second port of any of the disclosed microfluidic chips. The engagement surface 1220 of the port 1210 may be configured to engage with a port of the sample chamber 290 (i.e., port 293). The port 1210 may also include a channel 1240, which extends from a first opening 1230 on the first side of the second body portion 1200 to a second opening 1250 on the second side of the second body portion 1200. As shown in Figure 15A, the second side of the second body portion 1200 may include a mesh filter 1260. As described above, the mesh filter 1260 can filter the sample to prevent clogging of the microfluidic chip and / or to ensure that only samples of a specific size pass out of the microfluidic chip for collection. In some embodiments, the mesh size of the mesh filter 1260 is approximately 1 μm to 2000 μm, approximately 1 μm to 100 μm, approximately 100 μm to 200 μm, approximately 200 μm to 300 μm, approximately 300 μm to 400 μm, approximately 400 μm to 500 μm, approximately 500 μm to 600 μm, and approximately 600 μm to 700 μm. The range may be any of the following: μm, approximately 700 μm to approximately 800 μm, approximately 800 μm to approximately 900 μm, approximately 900 μm to approximately 1000 μm, approximately 1000 μm to approximately 1100 μm, approximately 1100 μm to approximately 1200 μm, approximately 1200 μm to approximately 1300 μm, approximately 1300 μm to approximately 1400 μm, approximately 1400 μm to approximately 1500 μm, approximately 1500 μm to approximately 1600 μm, approximately 1600 μm to approximately 1700 μm, approximately 1700 μm to approximately 1800 μm, approximately 1800 μm to approximately 1900 μm, or approximately 1900 to approximately 2000 μm, and any value within these ranges including the upper and lower limits may also be used.

[0122] Figures 16A and 16B show a diagram of a chip assembly 2020 having a first filter 1000a and a second filter 1000b at both ends of a microfluidic chip 2022. Figure 16B shows an exploded view of the chip assembly 2020. As shown in Figure 16B, in some embodiments, filter 1000a may be positioned such that port 1110a of the first body 1100a is fluidically connected to the sample chamber 2090a and port 1210a of the second body 1200a is fluidically connected to the first port 2021a of the microfluidic chip 2022. As shown in Figure 16B, filter 1000b may be positioned such that port 1110b of the first body 1100b is fluidically connected to the sample chamber 2090b and port 1210b of the second body 1200b is fluidly connected to the second port 2021b of the microfluidic chip 2022. However, Figures 16A and 16B show only one orientation of filters 1000a and 1000b on the chip assembly 2020. In some embodiments, on one or both filters 1000, the filters 1000 may be oriented such that the port 1110 of the first body 1100 is connected to the port of the sample chamber and the port 1210 of the second body 1200 is connected to the port of the microfluidic chip. In some embodiments, on one or both filters 1000, the filters 1000 may be oriented such that the port 1210 of the second body 1200 is connected to the port of the sample chamber and the port 1110 of the first body 1100 is connected to the port of one of the microfluidic chips. In some embodiments, the chip assembly 2020 may include filters 1000 on one end or both ends of the microfluidic chip 2022.

[0123] Filter 1000 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, 3D printing, injection molding, resin molding, or urethane casting. Filter 1000 can be made from various materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0124] [carriage] As described above, the carriage 230 may include a base platform 250, a spring platform 240, and a clamp 210. The assembly of the carriage 230 is described in more detail above with reference to Figures 3A and 3B.

[0125] Figures 17A to 17B show one embodiment of the base platform 250. The base platform 250 may include raised walls at one or both ends of the base portion. As shown in Figures 17A to 17B, the base platform 250 may include a first arm 256a and a second arm 256b configured to engage with a mating portion on the spring platform 240. In some embodiments, the first arm 256a may have a mating slot 257a on its side, and the second arm 256b may have a mating slot 257b on its side. In addition, to secure the tip assembly 220 on the carriage, the first arm 256a may have a retaining lip 259a on its upper part, and the second arm 256b may have a retaining lip 259b on its upper part. On the upper surface of the main body 253 of the base platform 250, the main body 253 includes end spring holders (distal spring holders) 254a and 254b for holding the base portion of the spring. The main body 253 further includes shoulder portions 236 configured to engage with the clamp 210. The main body 253 includes first openings 236a and second openings 236b configured to receive screws for securing the clamp 210 to the base platform 250. The bottom surface of the main body 253 of the base platform 250 may include a column portion 258 having a through hole extending between an end opening (distal opening) 258a and an opening 252. The bottom surface of the main body 253 also includes a plurality of pedestals (i.e., pedestals 251a and 251b). The support columns 258 and the pedestals (i.e., pedestals 251a and pedestals 251b) are configured to interact reversibly and hold the carriage 230 on one of the lateral arms 263 of the base section 260. The base platform 250 can be manufactured using a variety of methods such as additive manufacturing, subtractive manufacturing, injection molding, resin molding, or urethane casting. The base platform 250 can be made from a variety of materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0126] As shown in Figure 2B, the bottom surface of the base platform 250 may be provided with a plurality of capture elements. The capture elements serve to hold the carriage 230 in a desired position until there is a signal (or force, or lack of force) that allows the capture elements to disengage or stop interacting with the release elements, after which the carriage 230 is allowed to rotate, followed by re-engagement of the capture elements to stop the movement of the carriage 230. In some embodiments, this allows the carriage 230 to rotate through a 180-degree arc at a desired time in the tissue processing protocol. In some examples, the capture elements include magnets of a first polarity, and the release elements include magnets of the opposite polarity.

[0127] In the embodiment shown in Figure 2B, the capture element is a plurality of bases. The bases may include bases 251a and 251b. Each of bases 251a and 251b is configured to engage with a base fitting opening 266 of the transverse arm 263, thereby enabling intermittent rotation of the carriage 230 around the opening 264 of the transverse arm 263. Bases 251a and 251b can help hold the carriage 230 in a desired position until a signal or force is applied to disengage bases 251a and 251b from the plurality of base fitting openings 266. This allows for rotation of the carriage around the opening 264 and subsequent re-engagement of bases 251a and 251b with the plurality of base fitting openings 266.

[0128] Each of the multiple carriages 230 is held in the opening of each of the multiple arms using at least one pin (i.e., column 258) configured to allow out-of-plane rotation for each carriage 230. The out-of-plane rotation of each of the multiple carriages 230 allows each of the multiple carriages 230 to move between multiple orientations. For example, each of the multiple carriages 230 moves between 180-degree rotations (in-plane or out-of-plane). In some embodiments, each of the carriages 230 can move between orientations in which each of the multiple chambers is located along the plane of each of the multiple arms. Each of the multiple carriages 230 can move between 180-degree rotations (e.g., 0 to 45 degrees, 45 to 90 degrees, 90 to 135 degrees, 135 to 180 degrees, etc.).

[0129] Figures 18A and 18B show one embodiment of the spring platform 240. As described above, the spring platform 240 may include a first mating region 242a at a first end of the spring platform 240 and a second mating region 242b at a second end. The spring platform 240 has a top surface which includes a guide platform 248 positioned on the side of the spring platform 240 and a tip mating tab 246 positioned on the side of the spring platform 240 opposite to the guide platform 248. The spring platform 240 has a bottom surface which includes a proximal spring holder 244a and a proximal spring holder 244b configured to engage with the upper part of the spring 234, respectively. As shown in Figures 3A and 3B, the first mating region 242a and the second mating region 242b are configured to secure the first arm 256a and the second arm 256b of the base platform 250, respectively. The tip mating tab 246 of the spring platform 240 can be paired with the opening of the clamp 210. The spring platform 240 may also include a guide platform 248, which can be used to guide the microfluidic tip 222 onto the spring platform 240 when the microfluidic tip 222 is inserted into the carriage 230 and to assist in the compression of the spring platform 240. The spring platform 240 may also include a splash guard 245 that extends over the spring platform 240 to prevent fluid from being introduced into the spring 234 beneath the spring platform 240. The base spring holders 244a and 244b can help hold and secure the spring 234 in place during installation and use of the carriage 230. The spring platform 240 can be appropriately modified in size and shape to fit microfluidic tips 222 of any shape or size.The spring platform 240 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, 3D printing, injection molding, resin molding, or urethane casting. The spring platform 240 can be made from various materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0130] Figures 19A and 19B show one embodiment of the clamp 210. The clamp 210 can help hold the microfluidic tip 222 in place during use. The clamp 210 may include a finger groove 211 on the top of the clamp 210. This allows the user to easily access the microfluidic tip 222 and remove it from the carriage 230 if necessary. As shown in Figures 3A and 3B, the clamp 210 may include a base platform mounting area, which includes a first mounting flange 212a having a first opening 214a and a second mounting flange 212b having a second opening 214b. The base platform mounting area 214 can be fitted onto the shoulder 236 of the base platform 250 by a plurality of screws (e.g., a first screw 238a and a second screw 238b). The clamp 210 may also include a channel 215, which forms a groove into which a tip-fitting tab 246 fits, and helps to hold and position the spring platform 240. The height of the channel 215 allows vertical movement of the tip-fitting tab 246 within the channel 215, and can guide the movement of the spring platform 240 in one direction. The clamp 210 may also include guide rails 213a and 213b, which use grooves on the clamp 210 to guide and position the microfluidic tip 222 within the carriage 230. The clamp 210 may also include multiple openings 215. The openings 215 help to reduce the weight of the device. The openings 215 also allow the user to view the microfluidic tip 222 and / or adjust or remove the microfluidic tip 222 on the carriage 230. The clamp 210 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, injection molding, 3D printing, resin molding, or urethane casting. The clamp 210 can be made from various materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0131] [Other components] Figures 20A and 20B show a screw cap 270 configured to hold the base platform 250 of the carriage 230 in place. The screw cap 270 may include a proximal end 270a and a distal end 270b. The proximal end 270a includes a proximal opening 272. The proximal end 270a of the screw cap 270 has a shape that fits onto a column 258 on the bottom surface of the base platform 250. The screw cap 270 shown in Figures 20A and 20b shows a screw cap 270 with a hexagonal proximal opening 272, but the proximal opening 272 can be any shape that can accommodate the column 258 of the base platform 250. The screw cap 270 also has a through hole extending between the proximal end 270a and the distal end 270b. The through hole allows the screw cap 270 and the base platform 250 to be fastened to each other by a screw 232. The screw cap 270 may have a circular base end 270a. As described above, the base end 270a of the screw cap 270 may be configured to extend through the opening 264 of the base portion 260. The screw cap 270 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, 3D printing, injection molding, resin molding, or urethane casting. The screw cap 270 can be made from various materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0132] Figures 21A and 21B show a plurality of rotor foot inserts 280. The rotor foot inserts 280 may have a slanted portion 282 at the proximal end and a lip portion 284 at the base of the rotor foot insert 280. The rotor foot inserts 280 can assist in the smooth rotation of the carriage 230. Each of the slanted portions 282 of each rotor foot insert 280 allows the pedestals on the bottom surface of the base platform 250 (i.e., pedestals 251a and pedestals 251b) to move smoothly on the rotor base surface when the carriage 230 is rotating. Each of the rotor foot inserts 280 may also have a lip portion 284, which fits into each of the notches 268 of the lateral arm 263 of the base portion 260 to help fix each of the rotor foot inserts 280 in place during use. The rotor foot insert 280 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, 3D printing, injection molding, resin molding, or urethane casting. The rotor foot insert 280 can be made from various materials such as plastics (e.g., polycarbonate, acrylic, etc.), polyurethane, or metals (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0133] Figure 22 shows one embodiment of the base portion 260. As described above, the base portion 260 may include a central portion 261 and a plurality of transverse arms 263 extending radially away from the central portion 261 of the base portion 260. The base portion 260 in Figure 22 shows a base portion 260 having three transverse arms 263, but the base portion 260 may include any number of transverse arms 263 configured to reversibly engage with the carriage. In some embodiments, each of the transverse arms 263 is configured to be located in a plane parallel to the plane of the central portion 261. As described above, each of the transverse arms 263 may include an opening 264, at least one pedestal fitting opening 266, and a plurality of notches 268. As shown in Figure 2B, the opening 264 may be configured to receive the base of a column portion 258, and the plurality of notches 268 are configured to engage with pedestals 251a and pedestals 251b on the bottom surface of the base platform 250. Each of the notches 268 is configured to receive one of the rotor foot inserts 280. The base portion 260 can be configured to engage with various microfluidic chips. The base portion 260 may include mounting openings 262 configured to be attached to a centrifuge. In some embodiments, the mounting openings 262 are D-shaped chuck mounting openings 262 that fit into a chuck attached to the centrifuge. The mounting openings 262 have a "D" shape that helps prevent slippage during centrifugation. The base portion 260 also has a plurality of notches 268 which are shaped to help mount the rotor foot inserts. The base portion 260 also has carriage mounting holes, which allow the top of a screw cap 270 to fit into the carriage 230 and hold the carriage 230 in place. The base portion 260 also has a plurality of pedestal fitting openings 266 which fit into a pedestal on the bottom surface of the base platform 250 to keep the carriage aligned during use. The base portion 260 can be manufactured using various methods such as additive manufacturing, subtractive manufacturing, 3D printing, injection molding, resin molding, or urethane casting.The base portion 260 can be made from a variety of materials, such as plastic (e.g., polycarbonate, acrylic, etc.), polyurethane, or metal (e.g., aluminum, steel, stainless steel, surgical steel, brass, copper, etc.).

[0134] [Other embodiments] Figures 23A and 23B show one embodiment of a centrifuge. In this embodiment, each of the carriages 330 includes a clamping and locking mechanism for holding the chip assembly 220 in place during processing. As shown in Figure 23A, the carriage 330 comprises a base platform 350, the base platform 350 having a first side 355a and a second side 355b for forming an opening 353. The opening 353 can fix and hold the microfluidic chips 222 of the chip assembly 220.

[0135] The base platform 350 may include a first arm 356a at a first end of the base platform 350 and a second arm 356a at a second end of the base platform 350. The first arm 356a and the second arm 356a form clamps at either end of the carriage 330 to secure the tip assembly 220. The clamps formed by the first arm 356a and the second arm 356a can be separated and pulled away with one hand to allow the tip assembly 220 to be removed from the base platform 350. In some embodiments, the carriage 330 may be positioned on each of the transverse arms 263 of the base section 260 so that it can rotate 360 ​​degrees for reprocessing. The carriage 330 may include pedestals at both ends to help secure the carriage 330 in a direction parallel to the wing section (as described above). The carriage 330 is otherwise similar to the carriages described elsewhere in this specification.

[0136] Figure 24 shows a centrifuge having an alternative fastening for the clamp 210 described above. As shown in Figure 24, the carriage 430 utilizes a clamping and locking mechanism to hold the chip assembly 420 in place during processing. As illustrated, the base platform 450 of the carriage 430 may include a first side 455a and a second side 455b. A hinge 457 may be located on the second side 455b of the base platform 450, and an engagement portion 456 may be located on the first side 455a of the base platform 450. The hinge 457 allows a lid 452 to be movably connected to the base platform 450. The lid 452 may include a window 459 that allows viewing of the microfluidic chips of the chip assembly 420 during processing. The engagement portion 456 may include a slide lock that passes through a latch hole on the side of the lid and engages with a lock stop to cause the carriage 430 to hold the chip assembly 420. In some embodiments, the latch can be released with one hand by pulling the slide lock away from the carriage. The device is then unlocked by sliding the slide lock back into the latch hole in the lid. The lid 452 can be opened by swinging it using the hinge 457 on the second side 455b. The carriage 430 in Figure 24 can be rotated 360 degrees for reprocessing. The carriage 430 may include pedestals at both ends to help fix the carriage 430 in a direction parallel to the wing section (as described above). The carriage 430 is otherwise similar to the carriages described elsewhere in this specification.

[0137] Figures 25A and 25B show an embodiment of a centrifugal separator further including a motor 3000a, which is mounted at the bottom of a rotor base connected to a screw cap 270. The motor 3000a may be powered by a battery or connected to a power source via the base 260.

[0138] Figures 26A and 26B show one embodiment of a centrifugal separator including a motor 3000b, which is mounted on the bottom of a base 260 connected to a column 258 on the bottom surface of a base platform 250. The motor 3000b may be powered by a battery or connected to a power source via the base 260.

[0139] Figures 27A to 27D show another embodiment of the centrifuge 500 for processing biological samples. The centrifuge 500 shown in Figure 28 shows a centrifuge 500 capable of providing a multi-purpose centrifuge 500 for use in various conditions in processing protocols. The base 560 of the centrifuge 500 is capable of holding multiple syringes. As shown in Figure 27C, the centrifuge 500 comprises a base 560, which has a central section 561 with a mounting opening 562 and a number of transverse arms 563. Each of the transverse arms 563 is similar to the transverse arms 263 of the base 260 described above. Similar to the transverse arms 263 of the base 260, the transverse arms 563 of the base 560 include an opening 564 for receiving the column 258 of the base platform 250 of the carriage 230, allowing the carriage 230 to rotate around the opening 564. The lateral arm 563 also includes a plurality of base fitting openings 566, which hold the bases of the carriage 230 (e.g., bases 251a and 251b) in a desired position until a force is applied or released that allows the bases to be removed. The base 560 further includes a plurality of syringe holding arms 565a positioned between adjacent lateral arms 563. The syringe holding arms 565a may include a flared (protruding) end portion (flared distal end) 568 having an opening 565b. The end portion 568 may be angled or chamfered so that the opening 565b is not perpendicular to the syringe holding arm 565a. The flared end portion 568 may angle the opening 565b at a certain angle with respect to the plane of the syringe holding arm 565a. In some examples, the opening 565b is angled at the end portion 568 between approximately 100 and 130 degrees. In some embodiments, the terminal portion 568 may include an opening 565b at a 90-degree angle. The configuration shown in Figures 27A–27D allows for processing multiple samples to activate cells, and additional samples can be subjected to gradient separation using a syringe. This may be important because a specific subpopulation of cells from a sample can be isolated and then activated by processing it via a microfluidic tip positioned on a carriage of one of the lateral arms 263 of the base portion 260.

[0140] As shown in Figure 27D, the multiple openings 565b allow for the placement of multiple syringes 567a through the openings 565b. Each of the openings 565b is positioned with the needle hub 567e distal to the plunger 567d. In some embodiments, the syringe 567a includes a flange 567c that leans against a flared end portion 568 to hold the syringe 567a during the action of centrifugal force. In some embodiments, the openings 565b of the end portion 568 can hold the syringe 567a at an angle between 0 and 80 degrees from the horizontal position. In some examples, the openings 565b of the end portion 568 can hold the syringe 567a at an angle between 0 and 45 degrees from the vertical position. The end portion 568 of the syringe holding arm 565a is configured to fit (fit) multiple (e.g., three) syringes 567a simultaneously and to provide maximum centrifugal force on the syringes. As shown in Figures 27E to 27F, the end portion 568 of the syringe holding arm 565a allows each of the syringes 567a to slide into place. The opening 565b of the end portion 568 may include a gap 569 that allows the end portion 568 of the syringe holding arm 565a to expand to accommodate various syringes. For example, the opening 565b of the end portion 568 of the syringe holding arm 565a may be used with syringes holding volumes in the range of 1 mL to 100 mL.

[0141] In some embodiments, the end portion 568 of the syringe holding arm 565a may be machined or grooved to approximate the shape of a particular barrel 567b of the syringe 567a. In some embodiments, the end portion 568 of the syringe holding arm 565a may be able to receive and hold an insert, which is sized to hold any number of additional syringes. The insert may be configured such that the number of additional syringes is inversely proportional to the diameter of each opening for holding the syringes.

[0142] [Use of centrifuges for processing biological samples] The centrifuges disclosed herein process biological samples so that the tissue can be reinjected or reapplied to a patient to provide repair and / or regeneration. In some embodiments, the biological sample may be adipose tissue, but other types of tissue may be processed using the systems and methods disclosed herein. In some embodiments, adipose tissue, tumor tissue, cell preparations, liposuction, cultured cells, and other similar biological samples may be processed. In some examples, the biological sample comprises particles (e.g., nanoparticles, magnetic particles, reagents, or antibody-coated particles). In some embodiments, the sample comprises a cell-containing fluid.

[0143] Figure 28A shows one embodiment of a method 4000 for processing a biological sample. In the method for processing a biological sample 4000, step 4100 involves inserting the biological sample into a first sample chamber. Referring to a centrifuge 200 shown in Figures 2A-2B as a non-limiting example, in some embodiments, the biological sample may be introduced into one of the sample holding chambers 290a or 290b, which is fluid-coupled to the microfluidic chip 222 at either end of the microfluidic chip 222. As described above, the fluid path of the microfluidic chip 222 is configured to allow the biological sample to pass bidirectionally between the sample holding chamber 290a, the microfluidic chip 222, and the sample holding chamber 290b.

[0144] In method 4000 for processing a biological sample, step 4200 positions the carriage of a centrifuge in a first position. The carriage 230 of the centrifuge 200 may be fixed to a transverse arm 263 of the base 260 so as to be rotatable about an axis perpendicular or substantially perpendicular to the plane of the central portion 261 of the base 260. When the centrifuge 200 is in operation, the carriage 230 starts from a first position where the first end of the carriage 230 (i.e., the sample holding chamber 290a) is furthest from the central portion 261. The carriage 230 may rotate to a second position where the second end of the carriage 230 (i.e., the sample holding chamber 290b) is furthest from the central portion 261.

[0145] A method 4000 for processing a biological sample may include a step 4300 in which a rotational force is applied to a centrifuge. A rotational force may be applied to the centrifuge 200 so that the sample in the sample holding chamber 290a can pass from the sample holding chamber 290a through the first port 221a to the microfluidic tip 222. The sample can then pass through the fluid path of the microfluidic tip 222 from the first end of the microfluidic tip 222 to the second end of the microfluidic tip 222. The sample can then exit the microfluidic tip 222 and enter the sample holding chamber 290b through the second port 221b.

[0146] A method 4000 for processing a biological sample may include step 4400, in which the carriage is moved from a first position to a second position. The carriage 230 may then be rotated (i.e., 180 degrees) so that it moves from the first position to the second position. In some embodiments, the rotation is automated.

[0147] A method 4000 for processing a biological sample may include a step 4500 in which a rotational force is again applied to the centrifuge. At this second position, a rotational force may be applied to the centrifuge 200 so that the sample in the sample holding chamber 290b can pass from the sample holding chamber 290b through the second port 221b into the fluid path of the microfluidic tip 222. The sample can then pass through the fluid path of the microfluidic tip 222 from the first end to the second end. The sample can then exit through the first port 221a and enter the sample holding chamber 290a. The above steps of a method 4000 for processing a biological sample may be repeated until the sample is processed to the desired extent.

[0148] Method 4000 for processing biological samples may be used with a centrifuge 500 shown in Figures 27A to 27D. In the centrifuge 500, each of the syringes 567a held in the syringe holding arm 565a also contains a biological sample in the barrel 567b. When a rotational force is applied to the base 560, a centrifugal force is also applied to the biological samples in the syringes 567a, forming gradient separation of the biological samples. This may be important because it allows for the initial separation of specific subpopulations of cells from the sample before they undergo activation by processing via the microfluidic tip. Since each of the syringes 567a can be easily removed from the base 560, the biological material can be removed from the syringes 567a (or from any of the sample chambers attached to the microfluidic tip) and reattached to the tip assembly and carriage before continuing further processing of the biological material. Thus, method 4000 for processing biological samples can be adapted to various applications and uses.

[0149] In some embodiments, after processing a biological sample according to method 4000 for processing a biological sample, the processed biological sample may be injected into a patient.

[0150] [Treatment method] [overview] Figure 29 provides a schematic diagram of a non-limiting embodiment of the treatment paradigm. In this example, a subject (person) requiring tissue repair or regeneration (e.g., having a wound) undergoes a liposuction procedure (e.g., liposuction). The obtained adipose tissue is then processed according to embodiments disclosed herein and reinjected into the site and / or surrounding area of ​​the tissue requiring repair and / or regeneration.

[0151] [Administration and Medication] Furthermore, this specification provides a method for treating a subject (person) having a wound (e.g., DFU), the method comprising administering to the subject a composition comprising mechanically processed ADSCs (adipose-derived stem cells) as disclosed herein. For example, some embodiments of the compositions and methods described herein relate to the use of mechanically processed ADSCs to treat diabetic patients suffering from DFU. The use of ADSCs processed as described above to treat DFU is also provided. In certain embodiments, treatment of a subject with mechanically processed ADSCs as described herein achieves one, two, three, four, or more of the effects listed below, for example. (i) Reduction or improvement in the severity of the disease or related symptoms (ii) Shortening of the duration of disease-related symptoms (iii) Protection from the progression of the disease or related symptoms (iv) Regression of the disease or related symptoms (v) Protection from the progression or onset of symptoms related to the disease (vi) Protection against recurrence of disease-related symptoms (vii) Reduction in hospitalizations of the target group (viii) Shortening of hospital stay (ix) Increased survival rate of subjects with the disease (x) A decrease in the number of disease-related symptoms (xi) To enhance, improve, supplement, complement, or enhance the preventive or therapeutic effect of another therapy. Each of these comparisons is a contrast with different treatment methods for the disease, including the implementation of invasive procedures for treating deep and complex wounds such as DFU, for example.

[0152] Administration can be carried out by various routes including, but not limited to, intravenous, intra-arterial, subcutaneous, intramuscular, intrahepatic, intraperitoneal, and / or local delivery to the affected tissue. The dosage of mechanically processed ADSCs can be easily determined for a given subject based on its body weight, the type and condition of the disease, and the desired treatment aggressiveness. Depending on the embodiment, the number of cells per gram is about 10 5 ~ about 10 12 in the range (for example, 10 5 ~ 10 7 、10 7 ~ 10 10 、10 10 ~ 10 12 、and overlapping ranges thereof). In one embodiment, a dose escalation regimen is used. In some embodiments, for example, the number of cells per gram is about 1×10 6 ~ about 1×10 8 of ADSCs are administered. Depending on the embodiment, various types of wounds can be treated. In some embodiments, DFU is treated. Further embodiments provided herein include the treatment or prevention of non-limiting examples of wounds including venous stasis ulcers, arterial ulcers, and pressure ulcers (i.e., bedsores).

[0153] The dosage of mechanically processed ADSCs can be easily determined for a given subject based on its body weight, the type and condition of the disease, and the desired treatment aggressiveness. Depending on the embodiment, the number of cells per gram is about 10 5 ~ about 10 12 in the range (for example, 10 5 ~ 10 7 、10 7 ~ 10 10 、10 10 ~ 10 12 、and overlapping ranges thereof). In one embodiment, a dose escalation regimen is used. In some embodiments, for example, the number of cells per gram is about 1×106 ~Approx. 1×10 8 Mechanically processed ADSCs within this range are administered. Depending on the embodiment, various types of wounds can be treated.

[0154] [Examples] [Example 1: Determining the optimal CD-LOC processing parameters] The primary objective of this portion of the study examined is to determine the optimal fully automated lab-on-a-chip (CD-LOC) processing parameters for generating the maximum proportion of CD34-DPP4+ / CD55+ and multi-system differentiated stress-resistant cells (Muse cells) at an acceptable level of overall cell viability. This will be achieved by a block randomized experiment collecting five adipose tissue samples (10 ml each) from each of 15 diabetic patients. Body mass index (BMI) and anatomical location of adipose tissue harvesting will be recorded at the time of procedure. Adult diabetic patients are defined as those with a hemoglobin A1c (HbA1c) greater than 6.5. In addition, patients receiving any form of immunosuppressive therapy or suffering from an active systemic infection will be excluded from the study.

[0155] Based on the current design of the CD-LOC platform, the shear force is adjusted by adjusting the RPM of the processing unit. Adipose tissue samples are collected, stored at room temperature, and processed within 24 hours. Each sample is thoroughly washed with sterile PBS. Patient-specific tissue samples are randomized and processed at 0 (baseline), 25, 50, 75, and 100 kilodynes / cm². 2 The samples are subjected to five shear forces. A portion of each sample is flash-frozen in liquid nitrogen and stored at -80 degrees Celsius for subsequent immunohistochemistry.

[0156] To isolate stem cells from fat, all tissue samples are digested at 37°C for 30 minutes using 0.1% collagenase. This process is used solely for single-cell isolation and subsequent analysis and is not an element of device processing for clinical application. Each resulting pellet is filtered through a 100 μm strainer and subjected to RBC lysis before final single-cell analysis. Automated cell count and viability are determined using a dual-fluorescence cell counter (Luna-STEM, Logos Biosystems, Annandale, Virginia). Phenotypic marker analysis is also performed by staining tissue samples for CD45, CD34, and CD31 and analyzed by flow cytometry. Thus, five processing parameters are included to allow for a depiction of the log-linear nature of the inductive relationship.

[0157] To test cell activity / efficacy, cells were seeded in triplicate in a 96-well plate in standard control medium, and the activity of water-soluble tetrazolium salt was evaluated according to the manufacturer's protocol (Dojindo Molecular Technologies, Rockville, MD). To test population doubling, cells were seeded in a 1 cm³ 6-well plate containing control medium. 2 2x10 4 The cells are seeded in triplicate at a density of one cell and placed under standard culture conditions. When the first set of cells approaches 70-80% confluence (cell density), the number of cells is counted and evaluated via the following formula (Equation 1).

[0158]

number

[0159] Phenotypic marker analysis is performed by staining cells with the following combinations of markers. MSC:CD13-APCVio770 / CD45-VioBlue / CD34-PerCP-Vio700 / CD31-FITC / CD73-PE / CD146-APC Muse cells: CD13-APC-Vio770 / CD45-VioBlue / CD34-PerCP-Vio700 / CD31-FITC / SSEA-3-PE DPP4+ / CD55+:CD45-VioBlue / CD34-PerCP-Vio700 / CD31-FITC / CD55-APC, DPP4-APC Apoptosis detection (Annexin-V-FITC kit) All samples undergo transcriptional analysis, are treated with TRIzol reagent (Thermo Fisher Scientific) for RNA extraction, reverse transcribed, and then subjected to qRT-PCR analysis (Applied Biosystems Real-Time PCR 7300 system, TaqMan gene expression assay: angiogenesis, cell proliferation, wound healing, aging, tumorigenesis, cell survival). The main targets include DPP4, CD55, HIF-1α, IL-6, TNF-α, PPARy, HGF, VEGF, CXCL2, SCUBE3, DLL1, NR4A2, ADAMTS9, AK5, SOX2, RPA1, SGK1, HGF, IGF-1, SDF-I, PDGF-B, NGF-b, SCF, bFGF, POU5F1, and REX1.

[0160] The samples will also undergo secretome analysis. Each sample will be seeded (2 × 10) in serum-free medium (StemPRO MSC medium, ThermoFisher Scientific) in a T75 culture flask. 6 Cells are cultured and placed under standard culture conditions until confluence is reached. The culture medium (CM) is collected, centrifuged, and stored for subsequent assays. The conditioned medium is thawed and filtered using a 0.22 μm filter. An enzyme immunosorbent assay (ELISA) kit is purchased (R&D Systems, Minneapolis, Minnesota, or Signosis, Santa Clara, California), and the medium is assayed for cytokines (VEGF, HGF, IGF-1, SDF-1, PDGF-BB, NGF-β, SCF, bFGF, TNF-α) according to the manufacturer's protocol. Absorbance is measured at 450 nm by spectrophotometric method using an infinite microplate reader.

[0161] The paracrine activity of cells is tested using cell migration assays and scratch assays. A Boyden chamber (Neuroprobe, Gaithersburg, MD) is used for the cell migration assay. The Boyden chamber is equipped with an 8 μm pore size polycarbonate filter (Nucleopore, Whatman Incorporated, Clifton, New Jersey) coated with a 5 μg / mL gelatin solution, and keratinocytes, which are fibroblasts of ADSCs, are placed in migration medium at a rate of 1 × 10⁶ 5 Cells are introduced at their concentration. ADSC-conditioned medium is added to the lower compartment of the Boyden chamber, and the plate is placed under standard culture conditions. After 16 hours, the filter is removed from the Boyden chamber, fixed with 4% paraformaldehyde / PBS, stained with 0.5% crystal violet, and cell migration is counted under a microscope.

[0162] In the scratch essay, a monolayer of keratinocytes and fibroblasts is plated in a 6-well plate containing control medium and placed under standard culture conditions. Upon confluence, the center of each well is scratched using a P-200 tip to create a uniform cell-free zone. Cell debris is removed in a PBS washing step, and the wells are treated with CM from the control and experimental groups, as well as serum starvation medium serving as a negative control. Cultures are photographed at multiple consecutive time points to assess migration.

[0163] Preliminary data was obtained and additional data was prepared using the methods and experiments detailed above. The preliminary data was obtained using the "microfat" treatment (75 kilodynes / cm³). 2This process shows a quarter-reduction in the number of nucleated cells per mg / tissue compared to untreated tissue. Furthermore, no significant decrease in the viability of these cells was observed. In addition, preliminary data indicate that automated processing of "microfat" under increasing shear stress leads to a logarithmically correlated stem cell phenotype compared to manual processing, and a linear relationship. Finally, based on pilot data, cell viability is optimally 75 kilodynes / cm². 2 The shear force is 90%, and 100 kilodynes / cm². 2 The shear force reduces it by 50%. These results indicate that the shear forces involved in microfat processing lead to a significant upregulation of the regenerative phenotype. Further processing under various parameters can also make it possible to turn adipose tissue into nanofat.

[0164] [Statistical method] A linear mixed-effects regression is performed, with the unit of analysis being tissue samples from patients. The outcome variable is the proportion of CD34+ cells. The fixed effect includes shear force. The random effect is tissue samples from the same patient. Based on this mixed-effects model, the average proportion of CD34+ cells at each processing rate is calculated, taking into account intra-patient and inter-patient heterogeneity.

[0165] [Analysis of sample size and statistical power] Based on pilot data, the optimal cell viability is 75 kilodynes / cm³. 2 The shear force is 90%, and 100 kilodynes / cm². 2 It decreases by 50% at a shear force of 75 kilodynes / cm². Therefore, as the shear force increases, the average percentage of CD34+ cells increases linearly, up to 75 kilodynes / cm². 2It is hypothesized that the maximum level is reached at the shear force, resulting in a cell viability exceeding 75%. Using 15 diabetic patients and 5 tissue samples from each patient, 82% power is achieved to test this hypothesis based on a two-sided paired t-test at a 5% significance level. Secondly, it is expected that subpopulations of MSCs, Muse, and diabetic wound healing cells will reflect CD34 activity. Furthermore, it is expected that non-tumorogenic transcriptional patterns similar to those observed in previous adipose-derived Muse cell populations will be observed.

[0166] [Example 2: Characterization of micro-fragmented adipose tissue] Experiments were conducted to characterize adipose tissue obtained as a result of processing using the methods and systems disclosed herein. The adipose tissue was processed according to the non-limiting approach disclosed in Example 1. In some embodiments, since the final treatment paradigm is an autologous transplantation approach (see, for example, Figure 29), adipose tissue was collected from both healthy and diabetic donors to determine whether diabetic adipose tissue behaves similarly to that of healthy subjects.

[0167] The cellular profiles of the obtained microfragmented adipose tissue were evaluated by determining the changes in the CD26+ / CD55+ cellular component of the treated tissue. The CD26+ / CD55-positive phenotype indicates an adipose-derived stem cell subtype, and the resulting cells are thought to enhance the ability of the cells to contribute to tissue repair and / or regeneration. Figure 30A shows that treatment of adipose tissue using the systems and methods disclosed herein results in an increase in the percentage of CD26+CD55+ cells in the treated adipose tissue (labeled "Syntr") compared to untreated tissue. Furthermore, this is true whether the treated tissue comes from a healthy subject or a diabetic subject. Thus, regardless of whether the donor adipose tissue is from a healthy subject or a diabetic subject, treating the adipose tissue according to the methods disclosed herein and using the systems disclosed herein results in a higher percentage of the resulting population in the adipose tissue having a regenerative phenotype. Figure 30B also shows a similar phenomenon with respect to the detection of CD34+ cells (e.g., stem cells).

[0168] Cell density and cell viability within the treated adipose tissue were also evaluated. These data are shown in Figures 31A and 31B. As seen in Figure 31A, treatment of adipose tissue significantly reduces the number of cells per milliliter of adipose tissue (considering both healthy and diabetic adipose tissue samples). Figure 31B shows that the viability of cells in the treated adipose tissue (e.g., cells that survived the treatment procedure) is maintained at a level not statistically significantly different from that of untreated tissue. Considered together with the data from Figure 30, treating adipose tissue as disclosed herein reduces the cell density within the treated tissue, but the remaining cells are not only as viable as those in the untreated tissue, but the resulting cell population is rich in cells with stem cell-related phenotypes, suggesting that the post-treated adipose tissue has an improved ability to induce, promote, or facilitate tissue repair and / or regeneration.

[0169] Taking this concept of enriched cell populations to promote tissue repair and / or regeneration further, and examining the types of cells present in the treated tissue, we see that the relative proportion of cells classified as various types of stem cells increases after treatment. For example, Figure 32A shows a significant increase in the percentage of endothelial progenitor cells (EPCs) in the treated tissue, whether using healthy or diabetic adipose tissue. Similarly, Figure 32B shows a significant increase in the relative proportion of mesenchymal stem cells (MSCs) after treatment. Finally, Figure 32C shows an increase in the percentage of regenerative Muse cells after adipose tissue treatment.

[0170] Coupled with the relative enrichment of regenerating cell phenotypes, similar data reflecting this overall trend of cell number reduction are shown in Figures 33A–33C. Furthermore, these figures compare different adipose tissue processing approaches. Macroadipose tissue (MF) represents unprocessed adipose tissue. Nanoadipose tissue (NF30) is adipose tissue processed manually by passing between two syringes 30 times. Each of the remaining data points represents adipose tissue processed 10, 20, or 30 times using the systems disclosed herein. As seen in Figure 33A, processing of adipose tissue does not essentially change the overall viability of the cell types obtained that survive the processing procedure. Figure 33B shows that mechanical processing of adipose tissue results in an overall decrease in cell number. In particular, the cell numbers obtained remain stable even when the adipose tissue processing methods disclosed herein are used 10, 20, or 30 times. Figure 33C presents additional data for each histogram related to specific subtypes of cell populations within the adipose tissue. From left to right, MF, NF30, Syntr10, Syntr20, and Syntr30 are repeated in the same order, displaying multiple groups. The histogram on the far left of Figure 33C shows the relative percentage of CD34+ cells in the tissue sample. As seen in the figure, each of the treatment approaches performed results in a relative increase in the percentage of CD34+ cells in the treated tissue. In particular, with increasing the number of treatment runs, when adipose treatment is performed according to the embodiments disclosed herein and using the systems disclosed herein, the percentage of CD34+ stem cells in the treated tissue increases in a treatment run-dependent manner. Generally, similar results are seen not only for the percentage of EPCs shown in the third histogram, but also for Muse cells shown in the fourth histogram. Mesenchymal stem cells shown in the second histogram do not show this treatment run-dependent increase. However, it is worth noting that at least a similar level of enrichment occurs even when not exceeding NF30 syringe-based treatment. Similar results are observed for the CD26+ / CD55+ positive cell subtype.Taken together, these data indicate that adipose tissue treatment enriches tissues treated with cells exhibiting a regenerative phenotype and no decrease in viability, suggesting that adipose tissue treated in the manner disclosed herein and using the systems disclosed herein induces or enhances tissue repair and / or regeneration. In some embodiments, tissue repair is enhanced compared to adipose tissue treated in other ways.

[0171] [Example 3: The role of treated adipose tissue in promoting wound healing] This non-limiting example was conducted to compare the effects of standard, untreated diabetic interstitial vascular fraction (SVF) and microfragmented diabetic adipose tissue (SA) with higher levels of cellular activity in promoting wound healing in diabetic mice. Two adipose tissue samples (10 ml each) were collected from each of 20 diabetic patients. The collection methods, parameters, and criteria were those used in Non-Limiting Example 1 described above. Forty 6-week-old male db / db mice were obtained (The Jackson Laboratory, Sacramento, California) and maintained under controlled environmental conditions in an animal resource facility (constant laminar flow, temperature 20–23°C, humidity 40–60%, 12-hour light-dark cycle). Block randomized, paired-controlled, blinded experiments were conducted with two treatment groups.

[0172] Mice were intraperitoneally anesthetized with 12 mg / kg xylazine (Vedco, St. Joseph, Missouri) and 80 mg / kg ketamine (Ketathesia, Butler Scein Animal Health Supply, North Dublin, Ohio). A 6 mm full-thickness wound was created through the cutaneous-muscular layer on the back of each mouse and splinted with silicone. Mice were observed for 48–72 hours prior to the procedure. The bilateral wounds of each mouse were randomized, and one wound was injected with micro-fragmented activated diabetic adipose tissue or standard diabetic SVF resulting from the use of the systems and methods disclosed herein (see, e.g., Example 1), while the other wound (paired control group) was injected with placebo saline. In the experimental group, a total of 1 × 10⁶ per 125 μl of saline was injected. 5 Cells were subcutaneously injected into four sites around the wound margin, while the control group received only saline. The wounds were covered with occlusive dressings, and digital photographs were taken on days 0, 5, 9, 13, 17, 21, and 24. The wound area was measured using the digital photographs. At the earliest point of wound closure, the wounds were excised, fixed, and frozen sections were prepared. Slides were stained with hematoxylin & eosin (H&E), trichrome, and vimentin, and imaged using a fluorescence microscope (Evos FL, Thermo Fisher Scientific) with anti-CD31 antibody for basic histological and vascular density analysis. Research staff who performed the experiments / measurements were blinded from the treatment randomization scheme.

[0173] Data from this in vivo study are provided in Figures 34A–34C. Figure 34A shows histological results from excised wounds after closure. The leftmost panel of the figure represents wounds treated with saline, the middle column of the figure represents wounds treated with adipose tissue treated according to embodiments disclosed herein, and the rightmost panel of the figure relates to untreated SVF-treated negative control wounds. The top row shows hematoxylin-eosin staining, the second row shows Masson's trichrome staining, the third row shows immunohistochemistry for detecting vimentin, and the fourth row shows immunohistochemistry for detecting CD31. In the middle column, arrows indicate the presence of adipose tissue derived from treated human adipose tissue. The presence of adipose-derived stem cells is generally shown within circles, and rectangles generally indicate areas of increased skin regeneration. The improved staining of vimentin using SA adipose tissue indicates epithelial-to-mesenchymal transition, evidence of wound healing. Furthermore, increased expression of CD31 (also known as PECAM-1) has been shown to have a positive effect on inflammatory responses and angiogenesis (in particular, anti-PECAM-1 antibodies are known to block normal intercellular contact and affect cell migration, demonstrating the role of PECAM-1 in angiogenesis and wound healing).

[0174] Figure 34B shows photographic evidence of the treatment of induced wounds and healing timelines. As can be seen from these data, wounds treated with saline took the longest to heal, taking 19 days to close in the example shown in Figure 34B. Similarly, wounds treated with SVF took more than 3 weeks to heal. In contrast, wounds treated with adipose tissue processed as disclosed herein healed in less than 2 weeks. Photographic data are summarized in Figure 34C. These data showed that the time to wound closure was statistically significantly reduced when the adipose tissue process disclosed herein was applied to wounds compared to the administration of saline or SVF (by ANOVA or Student's t-test, p<0.0003 saline vs. SVF, p<0.00003 saline vs. SA, p<0.001 SA vs. SVF. Error bars represent SEM). These data support that the methods, processes, and systems disclosed herein and used for processing adipose tissue result in process tissue with enhanced wound-healing properties.

[0175] While embodiments of the present invention have been disclosed in connection with specific preferred embodiments and examples, those skilled in the art will understand that the present invention extends beyond the specifically disclosed embodiments to other alternative embodiments and / or uses of the present invention, as well as obvious modifications and equivalents thereof. In addition, while many variations of the present invention have been shown and described in detail, other modifications that fall within the scope of the present invention will be readily apparent to those skilled in the art based on this disclosure. Various combinations or subcombinations of specific features and aspects of the embodiments can be created and are intended to still be included in one or more of the present invention. Furthermore, any specific features, aspects, methods, characteristics, properties, qualities, attributes, elements, etc., disclosed herein relating to an embodiment can be used in all other embodiments described herein. Therefore, it should be understood that various features and aspects of the disclosed embodiments can be combined or substituted for each other to form various modes of the disclosed invention. For all embodiments described herein, the steps of the method do not need to be performed sequentially. Therefore, it is intended that the scope of the present invention disclosed herein should not be limited by the specific embodiments disclosed above.

[0176] [Statistical method] A linear mixed-effects regression analysis is performed, with mouse wounds as the unit of analysis. The outcome variable is the percentage of wound healing area. Fixed effects include two categorical variables for treatment groups (activated diabetic SVF and standard diabetic SVF) and an indicator of whether the wound is treated or placebo. Random effects include two wounds per mouse and two samples per patient. Based on this linear mixed-effects model, the mean increase in the percentage of healing area between treated wounds and placebo wounds is compared between the two treatment groups, taking into account heterogeneity between patients and mice.

[0177] [Analysis of sample size and statistical power] Using a total of 40 mice with bilateral wounds treated with either activated diabetic SVF or microfragmented diabetic adipose tissue, obtained from fat samples (2 samples per patient) taken from 20 diabetic patients, a two-sided paired t-test at a significance level of 5% was used to test the hypothesis that, 14 days after treatment, diabetic mice treated with microfragmented diabetic adipose tissue had a 95% mean increase in wound healing area compared to 65% of diabetic mice treated with standard diabetic SVF. This achieved 81% power to test this hypothesis.

[0178] The present invention is not limited to the embodiments described above. Of course, various changes and modifications can be made without departing from the scope and spirit of the invention.

[0179] Those skilled in the art will readily recall additional advantages and modifications. Therefore, in its broader embodiments, the present invention is not limited to the specific details and representative embodiments shown and described herein. Accordingly, various modifications can be made without departing from the spirit or scope of the general inventive concept defined by the appended claims and their equivalents.

Claims

1. A system for processing biological samples, It comprises a support plate and at least one carriage assembly, The support plate has a central portion configured to reversibly interact with a motor for providing rotational force, and at least one lateral arm. Each of the at least one lateral arm includes a first opening and at least one fixed opening, The at least one carriage assembly is configured to interact reversibly with one of the at least one transverse arms, Each of the at least one carriage assemblies comprises a base, a spring platform, at least one spring, and a fixing element. The base portion comprises an upper surface including at least one circular groove, a bottom surface including a column portion configured to extend through the first opening in one of the at least one lateral arms, a first arm, and a second arm. The spring platform comprises at least one circular groove located on the bottom surface of the spring platform, a first engaging portion, and a second engaging portion. The spring platform is positioned adjacent to the upper surface of the base portion, The at least one spring is held between the base portion and the spring platform. The at least one spring is fixed at its first end in the at least one circular groove of the base portion, and at its upper end in the at least one circular groove of the spring platform. The at least one spring is configured to provide an upward force to the spring platform. The fixing element is configured to fix the chip assembly on the at least one carriage assembly, A system in which each of the at least one carriage assemblies is rotatable by approximately 180 degrees.

2. The system according to claim 1, wherein the spring platform is configured to move between a first position and a second position in response to the insertion or removal of the chip assembly.

3. The aforementioned spring platform further includes tabs, The fixing element further comprises a channel configured to accept the tab of the spring platform, The system according to claim 2, wherein the tab is configured to move within the channel of the fixed element when the spring platform moves between the first position and the second position.

4. The system according to claim 1, wherein the fixing element includes at least one guide rail configured to guide and position the chip assembly.

5. The first engaging portion of the spring platform is configured to hold the first arm of the base portion, The system according to any one of claims 1 to 4, wherein the second engaging portion of the spring platform is configured to hold the second arm of the base portion.

6. The system according to any one of claims 1 to 5, wherein the spring platform further comprises a platform configured to guide the chip assembly into the at least one carriage assembly.

7. The spring platform is provided with a splash guard on the bottom surface of the spring platform. The system according to any one of claims 1 to 6, wherein the splash guard is configured to prevent fluid from being introduced into the at least one spring.

8. The chip assembly comprises a microfluidic chip, a first sample chamber, and a second sample chamber. The microfluidic chip has a fluid path extending from a first end to a second end of the microfluidic chip. The first sample chamber is fluidly connected to the first end of the microfluidic chip. The system according to claim 1, wherein the second sample chamber is fluidly connected to the second end of the microfluidic chip.

9. The system according to claim 8, wherein the fluid path has at least one expansion region and at least one compression region.

10. The at least one of the extended regions increases its radius along a first axis, a second axis, and a third axis, each perpendicular to the central axis of the fluid path. The at least one compression region has a diameter smaller than the diameter of the at least one expansion region. The system according to claim 9, wherein the diameter of the at least one compression region does not change.

11. The system according to claim 10, wherein the first axis, the second axis, and the third axis are perpendicular to each other.

12. The system according to claim 9, wherein the at least one extension region is increased in dimensions by three or more dimensions.

13. The system according to any one of claims 1 to 12, wherein the fluid path includes a plurality of teardrop-shaped extended regions.

14. The system according to any one of claims 1 to 13, wherein the fluid path includes a spherical or elliptical extended region.

15. The system according to any one of claims 1 to 12, wherein the fluid path includes a plurality of semi-teardrop shaped extended regions.

16. The system according to claim 15, wherein the fluid path includes a hemispherical or semi-elliptical extended region.

17. The system according to claim 15, wherein the fluid path includes a D-shaped extension region.

18. The system according to any one of claims 1 to 17, wherein the fluid path includes an hourglass-shaped portion.

19. The system according to any one of claims 1 to 12, wherein the fluid path comprises at least two expansion regions and at least one compression region disposed between the at least two expansion regions.

20. The fluid path comprises at least three expansion regions and at least two compression regions. The system according to any one of claims 1 to 12, wherein each of the at least two compression regions is located between adjacent expansion regions.

21. The system according to any one of claims 8 to 20, wherein a filter is disposed between the microfluidic chip and the first sample chamber, and between the microfluidic chip and the second sample chamber.

22. The system according to any one of claims 1 to 21, wherein the microfluidic chip is provided with Luer locks at a first end and a second end of the microfluidic chip.

23. A system for processing biological samples, It comprises a support plate, at least one carriage assembly, and at least one chip assembly, The support plate has a central portion configured to reversibly interact with a motor for providing rotational force, and at least one lateral arm. Each of the at least one transverse arm includes an interaction region, The at least one carriage assembly is configured to interact reversibly with one of the at least one transverse arms, Each of the at least one carriage assembly comprises a base portion, a spring platform positioned on the base portion, at least one spring held between the base portion and the spring platform, and a fixing element. The at least one spring is configured to provide an upward force to the spring platform. The at least one chip assembly comprises a microfluidic chip, a first sample chamber, and a second sample chamber. The microfluidic chip has a fluid path extending from a first end to a second end of the microfluidic chip. The fluid path has at least one expansion region and at least one compression region. The at least one of the expanded regions has an increased diameter along a first axis, a second axis, and a third axis, each perpendicular to the central axis of the fluid path. The at least one compression region has a diameter smaller than the diameter of the at least one expansion region. The aforementioned at least one compression region is configured such that its diameter does not change. The first sample chamber is fluidly connected to the first end of the microfluidic chip. The second sample chamber is fluidly connected to the second end of the microfluidic chip. A system in which each of the at least one carriage assemblies is rotatable by approximately 180 degrees.

24. The system according to claim 23, wherein the first axis, the second axis, and the third axis are perpendicular to each other.

25. The system according to claim 23, wherein the at least one extension region is increased in three or more dimensions.

26. The system according to any one of claims 23 to 25, wherein the fluid path includes a plurality of teardrop-shaped extended regions.

27. The system according to any one of claims 23 to 26, wherein the fluid path includes a spherical or elliptical extended region.

28. The system according to any one of claims 23 to 28, wherein the fluid path includes a plurality of semi-teardrop shaped extended regions.

29. The system according to any one of claims 23 to 28, wherein the fluid path includes a hemispherical or semi-elliptical extended region.

30. The system according to claim 29, wherein the fluid path includes a D-shaped extension region.

31. The system according to any one of claims 23 to 25, wherein the fluid path includes an hourglass-shaped portion.

32. The system according to any one of claims 23 to 31, wherein the fluid path comprises at least two expansion regions and at least one compression region disposed between the at least two expansion regions.

33. The fluid path comprises at least three expansion regions and at least two compression regions. The system according to any one of claims 23 to 31, wherein each of the at least two compression regions is located between adjacent expansion regions.

34. The system according to any one of claims 23 to 33, wherein a filter is disposed between the microfluidic chip and the first sample chamber, and between the microfluidic chip and the second sample chamber.

35. The system according to any one of claims 23 to 34, wherein the microfluidic chip is provided with Luer locks at a first end and a second end of the microfluidic chip.

36. A system for processing biological samples, It comprises a support plate and at least one carriage assembly, The support plate has a central portion configured to reversibly interact with a motor for providing rotational force, a plurality of lateral arms, and a plurality of holding arms. Each of the aforementioned plurality of lateral arms includes an interaction region, Each of the plurality of holding arms is positioned between each of the plurality of lateral arms, Each of the plurality of holding arms is configured to hold a syringe, The at least one carriage assembly is configured to interact reversibly with one of the at least one transverse arms, Each of the at least one carriage assemblies is configured to accept at least one chip assembly, A system in which each of the at least one carriage assemblies is rotatable by approximately 180 degrees.

37. A method for treating damaged tissue, In subjects with damaged or diseased tissue, a population of activated adipose stem cells is administered at approximately 1 × 10⁶ times per gram. ４ ~1 x 10 １０ This includes administering in amounts within a range of individual values. A method wherein the activated adipose stem cells result in one or more of the following: upregulation of a regenerating phenotype (e.g., CD34, CD13, CD73, or CD146), reduced inflammation, rapid tissue proliferation, rapid tissue remodeling, increased angiogenesis, or a combination thereof.

38. The method according to claim 37, wherein the administration is performed by subcutaneous injection.

39. The method according to claim 37, wherein the administration is performed by intravenous injection.

40. The method according to any one of claims 37 to 39, wherein the damaged tissue is a result of an ulcer.

41. The method according to claim 40, wherein the ulcer is selected from the group consisting of diabetic foot ulcers, pressure ulcers (i.e., bedsores), venous stasis ulcers, and arterial ulcers.

42. The method according to any one of claims 37 to 42, wherein the damaged tissue is a result of the wound.

43. The method according to any one of claims 37 to 42, wherein the wound is a burn-related wound, an abrasion (e.g., a road rash), a laceration (e.g., a knife wound), a stab wound, or a blunt force trauma (e.g., a bullet wound or a wound from another weapon).

44. The method according to any one of claims 37 to 43, wherein the administration results in the time to which the damaged tissue heals being shortened by at least 10% compared to damaged tissue that has not been exposed to the activated adipose stem cells.

45. The method according to any one of claims 37 to 44, wherein the administration results in the time to which the damaged tissue heals being reduced by at least 50% compared to damaged tissue that has not been exposed to the activated adipose stem cells.

46. A method for treating damaged tissue, A method comprising administering to a subject having damaged or diseased tissue a population of activated adipose stem cells in an amount sufficient to increase angiogenesis at the site of injury.

47. The use of activated adipose stem cells for the treatment of damaged tissue, The aforementioned population of activated adipose stem cells is approximately 1 × 10⁶ per gram of the target tissue with damage. ４ ~1 x 10 １０ It is provided in quantities within a range of individual units. The use wherein exposure of the damaged tissue to the activated adipose stem cells shortens the time it takes for the damaged tissue to heal by at least 10% compared to damaged tissue that has not been exposed to the activated adipose stem cells.

48. The use according to claim 47, wherein the activated adipose stem cells result in one or more of the following: upregulation of a regenerating phenotype (e.g., CD34, CD13, CD73, or CD146), reduced inflammation, rapid tissue proliferation, rapid tissue remodeling, increased angiogenesis, or a combination thereof.

49. The use according to claim 47, wherein exposure of the damaged tissue to the activated adipose stem cells reduces the time it takes for the damaged tissue to heal by at least 50% compared to damaged tissue that has not been exposed to the activated adipose stem cells.

50. A method for activating adipose stem cells used in tissue repair, The steps include: extracting a sample of adipose tissue from a patient in an amount sufficient to produce a sufficient amount of mechanically processed adipose-derived stem cells; The steps include inserting the adipose tissue sample into a first sample chamber located at one end of a microfluidic chip, A step of rotating a microfluidic chip using a motor-driven rotary chuck so that the adipose tissue sample is subjected to shear force, wherein while the microfluidic chip is rotating, the adipose tissue sample reciprocates along a plurality of microfluidic channels from a first sample chamber to a second sample chamber located at the opposite end of the microfluidic chip; The steps include extracting mechanically processed adipose-derived stem cells from the microfluidic chip, A method comprising the step of administering the mechanically processed adipose-derived stem cells to the site of injury in a patient.

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