Magnetically driven micro robot and manufacturing method thereof

Abstract

The present invention provides a magnetically driven microrobot and a manufacturing method thereof, which are used for carrying and delivering cells, and the method comprises the following steps: photocuring a photocurable material composition to form a main body of the magnetically driven microrobot. The photocurable material composition comprises: a degradable component, a structural component, a magnetic component, and a photocuring accelerating composition comprising a photoinitiator component and a photosensitizer component.

Description

[0001] Cross-references Related references

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 055,963 filed on July 24, 2020 and priority to U.S. Patent Application No. 17 / 128,345 filed on December 21, 2020, the entire contents of which are incorporated by reference into this application. Technical Field

[0003] The invention relates to a magnetically driven micro robot and a method for manufacturing the magnetically driven micro robot. Background Art

[0004] Microrobots are often used for biomedical applications in vivo and in vitro environments because they are small and can be controlled relatively easily. Some existing biomedical microrobots are non-degradable, while other biomedical microrobots are degradable. Non-degradable microrobots are relatively difficult to remove from their environment and are therefore not particularly suitable for in vivo applications. On the other hand, degradable microrobots may have relatively weak structures, rendering them unsuitable for carrying cells, etc. Summary of the invention

[0005] In a first aspect, the present invention provides a method for manufacturing a magnetically drivable microrobot. The microrobot is used to carry and deliver cells, drugs, medicaments, etc. The method comprises the following steps: photocuring a photocurable material composition to form a main body of a magnetically drivable microrobot. The photocurable material composition comprises: a degradable component, a structural component, a magnetic component, and a photocuring promoting composition including a photoinitiator component and a photosensitizer component. The degradable component can be used as a material matrix. The structural component can be used to provide strength or structural integrity to the main body. The magnetic component makes the main body magnetically drivable. The photoinitiator component and the photosensitizer component can be used to crosslink the various components of the photocurable material composition.

[0006] The photocurable material composition may include one or more additional components. Alternatively, the photocurable material composition substantially includes: a degradable component, a structural component, a magnetic component, and a photocuring accelerating composition having a photoinitiator component and a photosensitizer component.

[0007] The degradable component may include poly(ethylene glycol) diacrylate (PEGDA) or similar poly(ethylene glycol) (PEG) derivatives. In one example, the degradable component includes 74 vol% or about 74 vol% PEDGA (relative to the photocurable material composition).

[0008] The structural component may include pentaerythritol triacrylate (PETA). The structural component may include at least 20 vol%, at least 24 vol%, or at least 25 vol% PETA (relative to the photocurable material composition).

[0009] Optionally, the ratio of vol% of PEGDA to vol% of PETA is between about 2:1 to about 4:1, about 3:1 or 3:1.

[0010] The magnetic component may include Fe3O4 particles. The Fe3O4 particles may include Fe3O4 particle nanoparticles. The Fe3O4 particles may not exceed 2 vol% (relative to the photocurable material composition).

[0011] The photoinitiator component may include parabens.The photosensitizer component may include 2-isopropyl-9H-thioxanthen-9-one.

[0012] In one embodiment, the photocurable material composition substantially includes: PEGDA, PETA, Fe3O4 particles, a photoinitiator, and a photosensitizer. In one example, the photocurable material composition includes: about 74 vol% PEGDA, about 24 vol% PETA, about 2 vol% Fe3O4 particles, and trace amounts of photoinitiator and photosensitizer.

[0013] Optionally, photocuring (eg, laser writing) is selectively performed using a lithography technique such as 3D laser lithography or multiphoton lithography.

[0014] Optionally, the method further comprises the step of coating or applying a contrast agent on at least a portion of the body. The contrast agent may include a material that makes the microrobot easily imaged by an imaging device.

[0015] Optionally, the method further comprises the steps of coating or otherwise applying a photoacoustic imaging contrast agent on at least a portion of the body. The photoacoustic imaging contrast agent may include gold. In embodiments where the photoacoustic imaging contrast agent forms a coating, the coating may include one or more layers. In one example, the coating may have a thickness of nanometers (e.g., 10 nm).

[0016] Optionally, the method further comprises the following steps: forming a photocurable material composition by the following steps: mixing a degradable component and a structural component based on a first ratio to form a first mixture, and mixing the first mixture with a magnetic component based on a second ratio to form a second mixture. The method may also comprise the following steps: determining or optimizing the composition of the photocurable material composition, in particular the first ratio and the second ratio, before forming the photocurable material composition. The determination or optimization (thereby determining the first ratio and the second ratio) depends on the desired characteristics or application of the microrobot.

[0017] Optionally, the method further comprises the step of mixing the photocurable material composition before photocuring. The mixing may be performed using a mixer or a shaking machine.

[0018] Optionally, the method further comprises the step of developing the formed main body before light curing. The developing may include drying the main body, cooling the main body, blowing air towards the main body, etc.

[0019] The formed body may include a porous body having a three-dimensional structure having burr features. The three-dimensional structure may be of any shape, for example, a generally spherical shape.

[0020] Optionally, the photocuring of the photocurable material composition forms the corresponding bodies of multiple magnetically drivable micro-robots. These bodies can be formed sequentially (one by one) or approximately simultaneously. Adjacent bodies can be overlapped by direct material connection or without direct material connection, such as mechanical bonding. In one example, mechanical bonding can include two or more interconnected metal rings or rings without direct material connection.

[0021] Optionally, the method further comprises the following steps: attaching or loading cells, drugs, agents, etc. to the main body and / or the coating.

[0022] In a second aspect, the present invention provides one or more magnetically drivable microrobots manufactured at least using the method of the first aspect. The one or more microrobots are suitable for biomedical applications.

[0023] In a third aspect, the present invention provides a method for manipulating a magnetically drivable microrobot such as the second aspect. The method comprises the following steps: imaging a magnetically drivable microrobot in an environment using a photoacoustic imaging device, and based on the imaging, applying a magnetic field to the magnetically drivable microrobot to move the magnetically drivable microrobot in the environment. The environment may be an in vivo environment, such as a blood vessel. The photoacoustic imaging device may include a probe, an optical fiber, and the like.

[0024] Optionally, when the magnetically drivable microrobot moves due to the magnetic field, imaging is performed in real time (eg dynamically or continuously) to image the moving magnetically drivable microrobot and thus track it. The magnetic field may be a gradient magnetic field.

[0025] In a fourth aspect, the present invention provides a system for manipulating a magnetically drivable microrobot such as the second aspect. The system includes: a photoacoustic imaging device for imaging a magnetically drivable microrobot in an environment; and a magnetic field generator; the magnetic field generator is formed by one or more coils, and the magnetic field generator is used to provide a magnetic field to move the magnetically drivable microrobot in the environment. The environment can be an in vivo environment, such as a blood vessel. The photoacoustic imaging device can include a probe, an optical fiber, etc.

[0026] Optionally, the system further comprises a controller operatively connected to the magnetic field generator to control the operation of the magnetic field generator based on feedback from the photoacoustic imaging device and / or user input. The magnetic field generator may be a gradient magnetic field generator.

[0027] Optionally, the photoacoustic imaging device is configured to image the magnetically drivable microrobot in real time (e.g. dynamically or continuously) as the magnetically drivable microrobot moves due to the magnetic field, so as to image the moving magnetically drivable microrobot and thus track it.

[0028] By considering the detailed description and the accompanying drawings, other features and aspects of the present invention will become apparent.Any feature described herein with respect to one aspect or embodiment may be combined with any other feature described herein with respect to any other aspect or embodiment when appropriate and applicable. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Embodiments of the present invention are described by way of example with reference to the accompanying drawings, in which:

[0030] Figure 1 is a schematic diagram illustrating a method for manufacturing a magnetically drivable microrobot in one embodiment of the present invention;

[0031] Figure 2 is an example of an embodiment of the present invention for use in operation Figure 1 Schematic diagram of a system of a magnetically driven microrobot manufactured by the method;

[0032] Figure 3 is a graph showing degradation test results of microrobots fabricated from different ratios of PEGDA (vol%) to PETA (vol%);

[0033] Figure 4is a graph showing mechanical testing results of microrobots fabricated from different ratios of PEGDA (vol%) to PETA (vol%);

[0034] Figure 5A is a scanning electron microscope image of a microrobot fabricated using 90 vol% PEGDA:10 vol% PETA, showing the microrobot structural deformation and collapse;

[0035] Figure 5B is a scanning electron microscope image of a microrobot fabricated using 85 vol% PEGDA:15 vol% PETA, showing microrobot mesh deformation and burr fracture;

[0036] Figure 5C is a scanning electron microscope image of a microrobot fabricated using 80 vol% PEGDA:20 vol% PETA, showing burr fracture;

[0037] Figure 5D is a scanning electron microscope image of a microrobot fabricated using 75 vol% PEGDA:25 vol% PETA, showing that the microrobot is structurally intact;

[0038] Figure 5E is a scanning electron microscope image of a microrobot fabricated using 70 vol% PEGDA:30 vol% PETA, showing that the microrobot is structurally intact;

[0039] Fig. 5F is a scanning electron microscope image of a microrobot made using 65 vol% PEGDA:35 vol% PETA, showing that the microrobot is structurally intact;

[0040] Figure 6 is a graph showing the magnetic driving capability of microrobots fabricated using different percentages of Fe3O4 nanoparticles;

[0041] Fig. 7A is a scanning electron microscopy image of a microrobot fabricated using 2.5 vol% Fe3O4 nanoparticles;

[0042] Figure 7B yes Fig. 7A Magnified scanning electron microscopy image of the microrobot in the dashed rectangle;

[0043] Figure 7C yes Figure 7B A magnified scanning electron micrograph of a portion of the microrobot in the dashed rectangle shows that the burr is gone;

[0044] Figure 8is a graph showing the fluorescence signals of microrobots fabricated using different ratios of PEGDA to PETA in NaOH;

[0045] Fig. 9 is a graph showing the fluorescence signals of microrobots fabricated using different ratios of PEGDA to PETA in PBS;

[0046] Fig. 10A This is the initial image of the microrobot in NaOH;

[0047] Fig. 10B yes Fig. 10A Image of the microrobot after 2 hours;

[0048] Fig. 10C yes Fig. 10A Image of the microrobot after 3 hours;

[0049] Fig. 10D yes Fig. 10A Image of the microrobot after 7 hours;

[0050] Fig.11A This is the image of the microrobot that was originally in PBS;

[0051] Fig. 11B yes Fig.11A Image of the microrobot after 20 days;

[0052] Fig. 12A is a confocal scanning image showing a tissue section of a tissue implanted with a microrobot made using 75 vol% PEGDA and 25 vol% PETA at week 2;

[0053] Fig. 12B Is implanted Fig. 12A Fluorescent images of tissue sections of tissues containing microrobots;

[0054] Fig. 12C is a confocal scanning image showing a tissue section of a tissue implanted with a microrobot manufactured using 100 vol% PETA at week 2;

[0055] Fig.12D Is implanted Fig. 12C Fluorescent images of tissue sections of tissues containing microrobots;

[0056] Fig.13A is a confocal scanning image showing a tissue section of a tissue implanted with a microrobot made of 75 vol% PEGDA and 25 vol% PETA at week 4;

[0057] Fig. 13B Is implanted Fig.13AFluorescent images of tissue sections of tissues containing microrobots;

[0058] Fig. 13C is a confocal scanning image showing a tissue section of a tissue implanted with a microrobot manufactured using 100 vol% PETA at 4 weeks;

[0059] Fig.13D Is implanted Fig. 13C Fluorescent images of tissue sections of tissues containing microrobots;

[0060] Fig.14 is a graph showing the fluorescence intensity of microrobots fabricated with different ratios of PEGDA to PETA in vivo;

[0061] Fig.15A is a graph showing cell viability on day 1 in different cell culture media (MIHA, MSC, MNCC97L) with different concentrations of microrobot degradation products;

[0062] Fig. 15B is a graph showing cell viability on day 3 in different cell culture media (MIHA, MSC, MNCC97L) with different concentrations of microrobot degradation products;

[0063] Fig. 15C is a graph showing cell viability on day 5 of different cell culture media (MIHA, MSC, MNCC97L) with different concentrations of microrobot degradation products;

[0064] Fig.16A is a graph showing the comparison of photoacoustic magnitudes of different numbers of 10 nm gold-coated and non-gold-coated microrobots;

[0065] Fig. 16B is a graph showing the comparison of photoacoustic magnitudes of 10 nm gold-coated and non-gold-coated microrobots at different tissue thicknesses;

[0066] Fig.17A This is an ultrasound image of the left lobe pre-injected with 200 microrobots;

[0067] Fig. 17B yes Fig.17A Photoacoustic image of the left lobe in the figure, arrows indicate 200 microrobots pre-injected;

[0068] Fig. 17C yes Fig.17A Merged ultrasound-photoacoustic image of the left lobe in;

[0069] Fig.18A is an ultrasound image of the left lobe without the microrobot;

[0070] Fig.18B yes Fig.18A Photoacoustic image of the left lobe in the figure, arrows indicate background signal;

[0071] Fig. 18C yes Fig.18A Merged ultrasound-photoacoustic image of the left lobe in;

[0072] Fig.19A Photoacoustic images of in vivo navigation of a cell-laden microrobot at position a in the inferior vena cava of a nude mouse;

[0073] Fig.19B yes Fig.19A Photoacoustic image of in vivo navigation at position b;

[0074] Fig.19C yes Fig.19A Photoacoustic image of in vivo navigation at position c in FIG; and

[0075] Fig. 20 It is shown FIG. 19A to FIG. 19C A plot of the position error of the cell-laden microrobot at different locations in the inferior vena cava. DETAILED DESCRIPTION

[0076] Figure 1 A method 100 of making a magnetically drivable microrobot 10 for carrying and delivering cells in one embodiment of the present invention is shown. The microrobot can carry cells, etc. In one exemplary application, during a path in vascular tissue, the microrobot can be guided inside the tissue using real-time photoacoustic (PA) imaging and ultrasound (US) imaging techniques, thereby enabling optical adsorption to be detected with high resolution.

[0077] The microrobot 10 is manufactured using an optimized photocurable composite material, which includes: a degradable component configured to provide the degradability of the microrobot, a mechanical or structural component configured to provide mechanical strength or support for the microrobot, and a magnetic component configured to provide magnetic drive capability (which can make the microrobot respond to magnetic manipulation). The composition of the microrobot material can be optimized based on the application, especially taking into account factors such as degradability, mechanical strength (for carrying cells, drugs, etc.) and magnetic drive capability. The composition also includes a photocuring accelerating composition, which includes a photoinitiator and a photosensitizer used for the components in the cross-linked composition. The degradable component can include a derivative of polyethylene glycol (PEG), such as poly (ethylene glycol) diacrylate (PEGDA), which is used for drug delivery and tissue engineering-based applications. The structural component can include a derivative of acrylate, such as pentaerythritol triacrylate (PETA). The magnetic component can include magnetic or ferromagnetic particles, such as Fe3O4 particles, especially Fe3O4 nanoparticles. The photoinitiator and photosensitizer may include p-hydroxybenzoate and 2-isopropyl-9H-thioxanthen-9-one, respectively.

[0078] The method 100 for manufacturing a microrobot begins at step 102. In 102A of step 102, a degradable component used as a material matrix is ​​first combined with a structural component in a first ratio to form a first mixture, and mechanical strength is provided to the microrobot. Then, in 102B of step 102, a magnetic component is mixed with the first mixture of the degradable component and the structural component in a second ratio to obtain a second mixture, thereby providing a material composition with magnetic drive capability. The mixing in 102A and 102B of step 102 can be performed using a mixer or a shaker. The composition of the photocurable material composition, in particular the first ratio and the second ratio, can be determined or optimized before step 102 to achieve optimized degradability and mechanical strength according to the desired application. Specifically, the first ratio can be determined based on the structural integrity that bears the cell tension. The second ratio can be determined based on the drive capability requirements and manufacturing constraints.

[0079] After step 102, the optimized material composition is photocured using photolithography (such as 3D laser lithography or multiphoton lithography) to define a plurality of micro-robots in step 104. In the illustrated embodiment, the plurality of micro-robots 10 are defined as individual micro-robots without material connections.

[0080] Subsequently, in step 106, the photocured composition is developed to form a plurality of microrobots 10 for loading cells, etc. In the illustrated embodiment, the formed microrobots 10 include a porous body having a three-dimensional structure, which is generally spherical and has a plurality of burr members. The burr members extend approximately orthogonally relative to the outer surface of the porous body.

[0081] In one embodiment, the method 100 may include the following steps: coating or applying a contrast agent, such as gold, on at least a portion of the formed body. The contrast agent may be a photoacoustic imaging contrast agent that facilitates photoacoustic imaging or subject tracking based on photoacoustic imaging. The coating may have a thickness in the nanometer range (e.g., 10 nm).

[0082] After step 106, in step 108, cells are loaded onto the microrobot 10 so that the microrobot loaded with cells can be applied to an environment to carry and / or deliver the cells. The cells are loaded or attached to a coating of a main body or a photoacoustic imaging contrast agent. The cells can be loaded or attached between adjacent burr members of the same microrobot or between different burr members of adjacent microrobots.

[0083] Figure 2 A system 200 for manipulating a magnetically drivable microrobot 20 carrying cells in a living mouse is shown. The magnetically drivable microrobot 20 can be a microrobot manufactured based on the method 100 in one embodiment of the present invention. In this embodiment, a cluster of microrobots carrying cells is delivered to a blood vessel of a mouse. The microrobot 20 is designed with a light absorbing material to ensure a high light absorption coefficient and high contrast for hemoglobin molecules in the visible spectrum.

[0084] System 200 includes a photoacoustic imaging device with a probe 202 and an optical fiber 204, which is used to image the microrobot 20 in a mouse blood vessel (e.g., the inferior vena cava). System 200 also includes a sharp needle 206 and a catheter 208 covered with the sharp needle 206, which is used to pierce and deliver the microrobot 20 loaded with cells into the blood vessel. System 200 also includes a magnetic field generator 210, which is formed by one or more coils in this embodiment, and the magnetic field generator is used to provide a magnetic field to interact with the microrobot 20 in the mouse and thereby move the microrobot. The magnetic field generator 210 can be a gradient magnetic field generator. The photoacoustic imaging device can be used to image the microrobot 20 in real time (e.g., dynamically or continuously) when the microrobot 20 moves or is otherwise manipulated due to the magnetic field. Similarly, the movement of the magnetically driven microrobot 20 can be tracked by imaging. The system 200 may further include a controller (not shown) operably connected to the magnetic field generator 210 to control the operation of the magnetic field generator 210 based on feedback from the photoacoustic imaging device and / or user input.

[0085] Fabrication of magnetically actuated microrobots

[0086] In one experiment, a photocurable material composition including PEGDA (Sigma, 437441), PETA (Sigma, 246794), a magnetic nanoparticle solution (100 nm size, 260 mg / mL suspended in γ(gamma)-butyrolactone (GBL), custom made by chemicell GmbH), a photoinitiator (paraben, EasepiEDB, Curease Chemical, China), and a photosensitizer (2-isopropyl-9H-thioxanthen-9-one, Easepi ITX, Curease Chemical, China) was used to fabricate microrobots. The prepolymer solution (photocurable material composition) was mixed using a vortex shaker before forming the microrobot using photolithography. The composite material was then drop cast onto a clean glass substrate and loaded into Nanoscribe, a commercial two-photon direct writing system (GmbH, Germany) for structures written using a 63x oil immersion objective (1.4 numerical aperture from Zeiss). The substrates were then developed in toluene (Sigma, 179965) and isopropyl alcohol (IPA, Sigma, 67-63-0) and dried in a fume hood with air flow.

[0087] The cells are then loaded onto the formed microrobot. The formed microrobot is sterilized using UV irradiation and treated in a plasma cleaner for 30 seconds. Human iPSC10 MSC-GPx3 is trypsinized and resuspended at a concentration of 105 cells / mL. Then, this cell solution is drop-cast on the microrobot and incubated for 15 minutes at 37°C in a humidified incubator with 5% CO2 for cell attachment. Then, an additional 2ml of culture medium is added. After an overnight culture, the cells are stably attached to the microrobot. The microrobot is dehydrated in a critical point dryer (LEICA EM CPD300), and the morphology of the microrobot with cells is observed under SEM (FESEM, FEI Nova 450), as discussed in more detail below.

[0088] In addition, a microrobot cluster was prepared for a photoacoustic tomography (PAT) experiment. In this example, the microrobots were manufactured one by one. In order to avoid the scattering of the microrobots, the manufactured microrobots had a small overlap with the adjacent microrobots through the burr member (e.g., mechanical engagement, with or without direct connection), and the array of connected microrobots formed a square. In this example, a microrobot cluster of ten microrobots was established on four production lines: the first three production lines each contained three microrobots, while the last production line contained only one microrobot.

[0089] Optimization of photocurable material composition

[0090] In order to determine or optimize the composition of a photocurable material for a particular application (particularly with reference to Figure 1 The first ratio and the second ratio described in method 100) take into account the degradability and mechanical strength requirements of the microrobot.

[0091] Figures 3 to 5F Degradation and mechanical testing results of microrobots made from different ratios of PEGDA (vol%) and PETA (vol%) are shown. The ratios tested include: 90:10, 75:25, 50:50, 25:75, and 0:100. During the fabrication of the microrobots under test, the different ratios of materials were doped with the same amount of thiol polyethylene glycol rhodamine (Rhodamine B-PEG-Thiol, RB-PEG-SH).

[0092] For the degradation test, the fabricated microrobots were immersed in a PBS environment. At different times, images of the microrobots were captured (Zeiss fluorescence microscope), and the fluorescence intensity of the microrobots was analyzed using ImageJ software. Figure 3, it can be seen that the fluorescence intensity of the microrobot with a high PEGDA ratio decays faster, indicating that the microrobot with a high PEGDA ratio is beneficial to its degradability.

[0093] For mechanical testing, the fabricated microrobots were tested in a Hysitron TI950 dual-head nanoindentation system. Figure 4 The mechanical strength of the microrobot determined in this way is shown, where the inset shows the mechanical test of the microrobot in 75vol%PEGDA:25vol%PETA. As can be seen from Table 4, the mechanical strength of the microrobot increases with the increase in the proportion of PETA, and the material with a higher PETA composition requires less laser power in the microrobot fabrication. These indicate that a higher PETA composition can be beneficial to the structural integrity of the microrobot.

[0094] To determine the minimum PETA composition required for microrobot fabrication, materials with different ratios of PEGDA and PETA were tested based on structural integrity. It was found that a minimum composition of 10 vol% PETA was required to form properly constructed microrobots.

[0095] Then, the microrobots with different ratios of PEGDA and PETA were further evaluated in terms of mechanical strength based on loading mesenchymal stem cells (MSCs). Figure 5A , Figure 5B , Figure 5C Figure 5D and Fig. 5F It is shown that microrobots with less than 25 vol% PETA may collapse relatively easily after loading cells, while microrobots with 25 vol% or higher PETA can remain stable and intact when loaded with cells. In this example, in order to provide the best degradability and structural integrity, it may be advantageous to use 75 vol% PEGDA and 25 vol% PETA in the composition of the microrobot material.

[0096] Figures 6 to 7C Determining the reference based on magnetic drive capability and manufacturing constraints is illustrated Figure 1 The second ratio described in method 100. A microrobot made using a 75:25 ratio of PEGDA (vol%) to PETA (vol%), plus different ratios (1 vol%, 1.5 vol%, and 2 vol%) of Fe3O4 nanoparticles was tested. The results show that as the ratio of nanoparticles increases from 1 vol% to 2 vol%, the microrobot can achieve a higher driving speed. In this example, the ratio of the magnetic component is limited to 2 vol% because a higher ratio in the composition may reduce or block laser penetration, potentially making the microrobot defective.

[0097] Fig. 7A , Figure 7B and Figure 7C A microrobot made with 2.5 vol% magnetic content is shown. Since too much magnetic component in the composite material blocks laser writing during photocuring, defects are found in the scanning electron microscope (SEM) image. Therefore, in one example, 2 vol% magnetic nanoparticles are incorporated into a microrobot material of 75 vol% PEGDA and 25 vol% PETA for magnetic actuation of microrobots. In this example, in view of the above experimental results, a 74 vol% PEGDA, 24 vol% PETA and 2 vol% Fe3O4 nanoparticle composition solution is used.

[0098] Degradability and biocompatibility of magnetically actuated microrobots

[0099] The degradability and biocompatibility of the designed microrobots were evaluated. Figure 8 and Fig. 9 The fluorescence signals of the microrobots made of degradable 75vol% PEGDA:25vol% PETA hydrogel and the microrobots made of hard-to-degrade 100vol% PETA hydrogel in NaOH solution and PBS environment are respectively illustrated. The results show that the fluorescence intensity of the degradable 75vol% PEGDA:25vol% PETA microrobots decreased by 92% within 24 hours in NaOH solution, while the hard-to-degrade 100vol% PETA microrobots only decreased by 13% ( Figure 8 ). In the PBS environment, the fluorescence intensity of the degradable microrobots decreased by 40% after two weeks of culture, while the fluorescence intensity of the non-degradable microrobots remained almost unchanged ( Fig. 9 ). These results indicate that the fabricated microrobots can be degraded by hydrolysis, and the degradation in alkaline environment is much faster than that in PBS.

[0100] Fig. 10A , Fig. 10B Fig. 10C and Fig. 10D shows the degradation of the microrobot in NaOH solution, Fig.11A and Fig. 11B The degradation of the microrobot in a PBS environment is shown. PEGDA-based hydrogels will undergo a bulk degradation mode. In bulk degradation, the physical dimensions of the polymer network will not change significantly until the polymer network is almost completely degraded, but the fraction of polymer remaining in the hydrogel will decrease over time.

[0101] The local tissue environment, enzymatic oxidation, and macrophage activity may affect hydrogel degradation. In vivo testing of the degradability of microrobots was performed in the subcutaneous (SC) tissue of nude mice. Degradable 75vol% PEGDA:25vol% PETA microrobots and difficult-to-degrade 100vol% PETA microrobots were subcutaneously implanted into the left and right sides of each mouse. All microrobots were made using a prepolymer solution containing 1mg / mL RB-PEG-SH and were rinsed and dried before implantation. The mice were killed at the 2nd and 4th weeks. The size of the implanted area was approximately 1×1cm 2 The skin fragments were fixed with formaldehyde solution. The fixed skin tissue was then embedded in a cryogenic matrix freezing medium and cut vertically to a thickness of 50 μm using a cryostat.

[0102] Fig. 12A , Fig. 12B , Fig. 12C , Fig.12D , Fig.13A , Fig. 13B , Fig. 13C and Fig.13D A tissue section of a tissue implanted with a microrobot is shown. The confocal scanning image shows that the microrobot is not washed away. The fluorescence signal of Rhodamine B is only present inside or near the microrobot. There is no leakage of Rhodamine B in a large area. The release process of Rhodamine B is related to the degradation of the microrobot. The fluorescence signal of Rhodamine B on the microrobot has been characterized. Fig.14 The fluorescence intensity of the degradable 75 vol% PEGDA:25 vol% PETA microrobot decreased by about 65% 2 weeks after implantation and decreased by 82% 4 weeks after implantation. The fluorescence intensity of the difficult-to-degrade 100 vol% PETA microrobot decreased by 19% and 30%, respectively. These experimental results confirm the in vivo degradability of the microrobot.

[0103] Considering that the degradation products of the microrobots would stay in the body for a certain period of time, a survival test was then performed to examine the biocompatibility of the fully degraded products. The MTT assay was performed on days 1, 3, and 5 using cells cultured in medium supplemented with different concentrations of the degradation products. Fig.15A , Fig. 15B and Fig. 15C It is shown that even when 800 ppm of degradation products are added to the culture medium (equivalent to the degradation of 10k microrobots in 1 uL solution), the cell survival rate is not substantially impaired. Even at ultra-high concentrations of degradation products, the survival rate of all cell types is above 80%.

[0104] In vivo imaging of magnetically actuable microrobots

[0105] To facilitate in vivo experiments using microrobots, microrobots can be imaged in real time at depths of a few millimeters to a few centimeters. In one example, a microrobot is designed and PA tomography (PAT) is tailored for in vivo microrobot imaging. The microrobot is designed with light absorbing materials to ensure high light absorption coefficients and high contrast for hemoglobin molecules in the visible spectrum. By tuning the optical excitation wavelength, the contrast of the microrobot in the blood is optimized, and the blood and microrobots are quantified simultaneously. To visualize the tissue environment, co-registered PA and US images are acquired simultaneously.

[0106] In this example, in order to enhance the PA signal of the microrobot and improve the contrast between the microrobot and the blood, 1064nm was selected for in vivo imaging. The microrobot was coated with a 10nm thick gold layer (photoacoustic imaging contrast agent) to further enhance absorption while ensuring good biocompatibility. In order to deliver more therapeutic cells to the diseased site, a cluster of microrobots (from a few to hundreds of microrobots) was used in a single treatment. A large number of microrobots increased the PA signal. Fig.16A The PA magnitude is positively correlated with the number of microrobots, both at a 1064nm laser wavelength. Compared to uncoated microrobots under the same excitation, the gold-coated microrobots produce five times the signal intensity.

[0107] To demonstrate microrobot imaging in deep tissue, two microrobot clusters were imaged in chicken tissue, e.g. Fig. 16B As shown. One cluster consists of 10 gold-coated microrobots, while the other cluster consists of 10 uncoated microrobots. Compared with the uncoated microrobot cluster, the coated microrobots show stronger PA signals at various tissue thicknesses. The gold-coated microrobot cluster can be clearly imaged at a depth of 2 cm, while the uncoated cluster is almost undetectable at a depth of 2 cm. This experiment shows that engineered microrobots have good contrast in deep PA imaging.

[0108] The imaging of the microrobots was further tested in the mouse liver. 200 microrobots were injected into the left lateral lobe (LLL) via the portal vein, and the microrobots were then collected and fixed in paraformaldehyde (PFA) and imaged using US and PAT. Fig.17A , Fig. 17B , Fig. 17C , Fig.18A , Fig.18B and Fig. 18CThe imaging results are shown. The US image shows the morphology of LLL, and the PA image shows the absorption contrast of the microrobot to the liver tissue. The combined US and PA images show the distribution position of the microrobot in the LLL. The signal magnitude of the LLL with 200 injected microrobots is 2.8 times that of the control.

[0109] Magnetically actuable microrobots navigate in vivo in a mouse model

[0110] In vivo navigation of cell-laden microrobots guided by PA imaging was performed at a depth of 6 mm in the inferior vena cava of nude mice (male, 6 to 8 weeks). Figure 2 The microrobot cluster consisted of five gold-plated microrobots. The cell-carrying microrobot cluster was delivered into the inferior vena cava of nude mice using a needle-covered catheter. Guidance was provided by real-time PA and US imaging (20 frames per second per imaging mode), and the microrobots inside the catheter were activated by using the gradient magnetic field in the vein. A proportional-integral-derivative (PID) controller was used for motion control, and position feedback was obtained by visual processing of the PA images. During microrobot navigation, two target positions were set and labeled b and c (respectively Fig.19B and Fig.19C The microrobot cluster with cells starts from the starting position a ( Fig.19A ) to the target position b, and then move from b to the target position c, with a total displacement of 5.1 mm. Fig. 20 As shown, the position error of the microrobot indicates that the microrobot with cells can accurately navigate in the vascular environment to transport therapeutic cells.

[0111] The above-mentioned embodiments of the present invention provide a method for manufacturing a magnetically driven and / or degradable microrobot that can be loaded with cells, which promotes wireless and minimally invasive methods for accurately treating diseases. In one example, the microrobot can be used for accurate cell delivery in the living body of a human or other animal (for example, in vascular tissue). The microrobot can be navigated and tracked by in vivo photoacoustic imaging for targeted therapy. The microrobot will degrade after performing a task in the body (for example, delivering cells or drugs), and the side effects are relatively few. The above-mentioned embodiments of the present invention also provide an imaging method for guiding the microrobot to move in vivo or in vitro. Photoacoustic imaging can enable real-time navigation and provide high-resolution images, especially at millimeters to centimeters of tissue, potentially promoting the surgical and / or therapeutic applications of the microrobot.

[0112] Those skilled in the art will appreciate that, without departing from the scope of the present invention, various changes and / or modifications may be made to the illustrated embodiments. Therefore, the described embodiments of the present invention are to be considered in all aspects as illustrative and non-restrictive.

[0113] The illustrated method for making a microrobot can be used to make microrobots with different structures, shapes, forms, sizes, etc. Moreover, the illustrated microrobot can be manipulated in different environments using unillustrated methods or systems. For example, the method for making a microrobot may be different from the illustrated method. Different photocuring techniques (including but not limited to photolithography) can be used to form a microrobot. The method can be used to form one or more microrobots. The photocurable material composition may include: one or more degradable components; one or more structural components; one or more magnetic components; and one or more photocuring promoting compositions (including one or more photoinitiator components and one or more photosensitizer components). The degradable component does not have to be PEGDA. The structural component does not have to be PETA. The magnetic component does not have to be Fe3O4 particles, but can be other ferromagnetic or magnetic materials. The contrast agent can be coated or applied to at least a part of the body. The photocurable material composition can be photocured using other techniques other than photolithography. Multiple microrobots can be formed one by one or approximately simultaneously (e.g., photocuring and developing). Adjacent microrobot bodies can be overlapped using direct material connection or without direct material connection, such as mechanical bonding. For example, the microrobot may be different from the illustrated microrobot. The microrobot may have a solid body instead of a porous body. The microrobot may have a non-spherical shape. The burr members on the body of the microrobot may have different orientations, forms, sizes, lengths, etc. In some embodiments, the body of the microrobot may not have any burr members on the body. In some embodiments, the body of the microrobot may include a surface depression. The microrobot can be used to carry and deliver cells, drugs, etc. The illustrated methods and systems for manipulating microrobots can be modified for different control applications. The illustrated methods and systems can be modified for use in other in vivo or in vitro environments. The system for manipulating a microrobot may include additional or alternative devices (instead of sharp needles and catheters) for delivering the microrobot to or into an environment.

Claims

1. A method for manufacturing a magnetically drivable microrobot cluster, the method comprising the following steps: photocuring the photocurable material composition to form a main body of a magnetically drivable microrobot; The photocurable material composition is photocured to form a plurality of main bodies of magnetically drivable micro-robots; the plurality of micro-robot main bodies of the micro-robot cluster meet the following requirements: (1) The main body comprises a porous body having a three-dimensional structure, wherein the three-dimensional structure has a burr component; (2) Adjacent bodies overlap; the overlapping adjacent bodies are mechanically joined by burr members; (3) coating or applying a photoacoustic imaging contrast agent on at least a portion of the body to form a coating; Wherein, the photocurable material composition comprises: Degradable components; Structural components; a magnetic component; and A photocuring accelerating composition comprising a photoinitiator component and a photosensitizer component.

2. The method according to claim 1, wherein: The degradable component includes a polyethylene glycol derivative.

3. The method according to claim 2, wherein: The polyethylene glycol derivative is polyethylene glycol diacrylate.

4. The method according to claim 1, wherein: The structural component includes pentaerythritol triacrylate.

5. The method according to claim 1, wherein: The magnetic component includes Fe3O4 particles.

6. The method according to claim 5, wherein: The Fe3O4 particles include Fe3O4 nanoparticles.

7. The method according to claim 1, wherein: The photocuring is selectively performed using photolithography.

8. The method according to claim 7, wherein: The photocuring is selectively performed using 3D laser lithography or multiphoton lithography.

9. The method according to claim 1, wherein: The contrast agent comprises gold.

10. The method according to claim 1, further comprising the steps of: The photocurable material composition is formed by the following steps: mixing the degradable component and the structural component based on a first ratio to form a first mixture, and The first mixture is mixed with the magnetic component based on a second ratio to form a second mixture.

11. The method according to claim 10, further comprising the steps of: The composition of the photocurable material composition is determined before forming the photocurable material composition.

12. The method according to claim 11, wherein: The step of determining the composition of the photocurable material composition includes determining the first ratio and the second ratio.

13. The method according to claim 1, further comprising the steps of: The photocurable material composition is mixed before the photocuring.

14. The method according to claim 1, further comprising the steps of: The formed body is developed prior to the photocuring.

15. The method according to claim 1, wherein: The degradable component includes polyethylene glycol diacrylate, and the structural component includes pentaerythritol triacrylate; and wherein the ratio of vol% of polyethylene glycol diacrylate to vol% of pentaerythritol triacrylate is 3:

1.

16. The method according to claim 1, wherein: The overlapping adjacent bodies are mechanically joined without direct material connection.

17. The method according to claim 1, further comprising the steps of: Cells are attached or loaded to the body.

18. The method according to claim 1, further comprising the steps of: Cells are attached or loaded to the coating.

19. A magnetically driven micro-robot cluster, comprising: a plurality of bodies manufactured by photocuring a photocurable material composition, each body comprising a porous body having a three-dimensional structure having a burr member; Adjacent subjects are overlapping; Overlapping adjacent bodies are mechanically joined by burr members; The photocurable material composition comprises: Degradable components; Structural components; a magnetic component; and A photocuring accelerating composition comprising a photoinitiator component and a photosensitizer component; The magnetically drivable microrobot further comprises a contrast agent material or coating disposed on at least a portion of the body.

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