Introducing the IVM Intravital Microscope
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In this second session of the IVM webinar series, we take a deeper look at the IVM all-in-one intravital microscope. The IVM system is a carefully engineered state-of-the-art intravital fluorescence microscopy platform optimized to perform real-time imaging of dynamic phenomena in in vivo tissues at a cellular level. This technique can serve as a next generation core technology to elucidate the pathophysiology of various human diseases and assist in the discovery of new cures. We will review the different modules of the system allowing for intravital confocal and/or two-photon imaging, along with the features and benefits of each set-up. Key Features All-in-one intravital microscopy system with a flexible design for modification and updates Optimized for in vivo observation of dynamic processes in mouse models of human disease Four-color simultaneous confocal/two-photon imaging Sub-µm image resolution and ultrafast video-rate imaging (max. 100 fps - 512x512 pixels) Integrated automatic high-precision motion artifact compensation Key Applications In Vivo 4D cell imaging, tracking and monitoring In Vivo visualization of dynamic molecular & cellular mechanisms In Vivo efficacy monitoring of novel drug compound In Vivo monitoring of material delivery target tissues In Vivo real-time imaging of microcirculation
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Introducing the IVM Intravital Microscope
- 1. Introducing the IVM Intravital Microscope All-in-One Intravital Confocal/ Two-Photon Microscopy System
- 2. WWW.SCINTICA.COM 2 Topics of Discussion • Intravital Microscopy • IVM Systems Overview • Key Research Applications
- 4. WWW.SCINTICA.COM Optical Imaging Resolution vs Penetration Depth Nguyen et al. EMJ Radiology, 2020
- 5. WWW.SCINTICA.COM In Vivo Imaging Optimal Imaging Window Adapted from Ruggiero et al. Dalton Transactions, 2016 Optimal Imaging Window Tissue Penetration (mm) UV IR VIS + NIR H2O H2O Relative Absorbance 100 10 1 0.1 0.01 Hb HbO2 Hb
- 6. WWW.SCINTICA.COM Fluorescence Imaging Fluorophores • Genetically modified fluorescent proteins: • Organic dyes: FITC/TRITC, Alexa (conjugated antibodies) • Inorganic dyes: Quantum Dots • Endogenous species: elastin, collagen, NADH/FAD
- 8. WWW.SCINTICA.COM Confocal/ Two-Photon Microscope Mostany et al. Advanced Fluorescence Microscopy, 2014 Upright 100 um 500 um Upright Confocal Two-Photon
- 9. WWW.SCINTICA.COM Confocal/ Two-Photon Microscope Mostany et al. Advanced Fluorescence Microscopy, 2014 Single Photon Excitation Two-Photon Excitation
- 10. WWW.SCINTICA.COM 10 Conclusions Optical imaging has excellent resolution but poor penetration depth Propagation of light through biological tissue is affected by hemoglobin and water The optimal window for in vivo optical imaging lies between 600 and 1000 nm Fluorescence is the commonly used optical technique for in vivo imaging Up to 4 fluorophores can be imaged simultaneously when chosen carefully to avoid spectral overlap
- 11. WWW.SCINTICA.COM Intravital Microscopy | Why? Macroscopic-scale Imaging & Diagnosis (Organ/Tissue-level :StructuralInformation) Microscopic-scale Imaging & Analysis (3D Cell-level: Molecular/FunctionalInformation) Intravital microscopy (IVM) Islet cell (MIP-GFP) Vessel (CD31) PET / CT / MRI X-Ray Ultrasound Cancercell (B16F10-GFP) Vessel (CD31) Sinusoid (LYVE-1) Blood circulation (FITC-Dextran) Neutrophil (Ly6G) disease stage, development, and treatment response In situ Spatial information Real-time Quantitative Longitudinal Reduces animals
- 12. WWW.SCINTICA.COM Intravital Microscopy | Where?
- 13. WWW.SCINTICA.COM Intravital Microscopy | How? Coste et al. Cytometry, 2019
- 14. WWW.SCINTICA.COM Intravital Microscopy | Window Chambers Dorsal Skinfold Chamber - Cancer Xenograft imaging Cranial Imaging Window - Long-term repeated brain imaging Abdominal Imaging Window - Long-term abdominal organ imaging
- 15. WWW.SCINTICA.COM Intravital Microscopy | What? For cancer and drug development, intravital microscopy enables a direct imaging analysis of the tumor development and drug delivery to target tissue as well as efficacy, and mode of action (MOA) of new therapeutic candidates at a microscopic, cellular level in various preclinical model of human disease. Cancer Metastasis - CTC Cancer cell dissemination to circulation Drug Delivery - Nanoparticle Anti-cancer nanoparticle delivery Circulating Tumor cell (CTC) Cancer ll Stromal cell Drug carrier Cancer cell Vessel Bone Marrow Transplanted BM cell, HSPC cell H2B Sinus
- 16. WWW.SCINTICA.COM Skin Inflammatory response KARS Granulocyte Intravital Microscopy | What? Intravital microscopy enables dynamic 3D imaging of various cellular-level dynamics such as cell trafficking, cell-cell interaction, and cell-microenvironment interaction inside the living body in vivo, providing a new insight in the processes of human disease development. Cancer Xenograft - T cell Triple Negative Human Breast Cancer T cell Cancer cell Vessel Lung Microcirculation in Sepsis Model Neutrophil Blood flow
- 17. WWW.SCINTICA.COM Intravital Microscopy | Quantification Intra-/extravasation Cell motion tracking Cellular interactions Intracellular changes Cell area/count Vascular analysis Adapted from Evans et al. Experimental Neurology, 2019
- 18. WWW.SCINTICA.COM Intravital Microscopy | Motion Artifacts Uncorrected Motion Artifacts Corrected for Motion Artifacts
- 19. WWW.SCINTICA.COM 19 Conclusions Successful intravital imaging depends on: • Before imaging System chosen Animal model optimization Window chamber quality • During imaging Animal positioning Animal maintenance Imaging parameters • After imaging Pre-processing corrections Post-processing quantification
- 21. WWW.SCINTICA.COM 21 IVM Systems Overview
- 22. WWW.SCINTICA.COM IVIM Technology | Who? Intravital Imaging of Various Organs in Human DiseaseAnimal Model Real-time Intravital Imaging Endomicroscopy https://scholar.google.com/citations?user=RRDHF9oAAAAJ&hl=en
- 23. WWW.SCINTICA.COM IVIM’s All-in-One System Conventional Approach IVIM Technology | Why?
- 24. WWW.SCINTICA.COM IVIM Technology | When? 2017.06 • Founded • KAIST Munji Campus 2017.08 • Series A Investment (2.7mil.USD/30억원) 2017.09 • Exclusive Licensing Technology from KAIST 2018.07 2018.02 • Establishment Seoul Marketing Office 2018. 09 • First Release All-in-One IVM IVM-C/IVM-CMmodel 2018.11 • First Installation IVM-C model SNU Bundang Hosp. R&D center 2020.12 | Cumulative Sales • IVM System | 2 Mil USD - SNU Med. School / IBS / Curacle, etc • IVM Imaging Service | 300k USD - Academia (University, Hospital) Industry (Bio-tech, Pharmaceutical Company) - Total 21 projects, completed/on-going 2019.09 2019.02 2019.10 2020.02-08 2020.07 http://imnews.imbc.com/replay/2018/nwdesk/article/4846789_22663.html • Establishment R&D Center • New Model Release Multi-Photon IVM IVM-M/ IVM-MSmodel • New Installation IVM-CM model SNU Medical School Yongon Campus • Series B Investment (7.3mil.USD/80억원) • New Installation IVM-C/M/MS model YMC, AMC, IBS, KAIST Knotus, Curacle, etc • USAsite Installation IVM-MS model Harvard Med. School Boston, MA, USA
- 25. WWW.SCINTICA.COM IVIM’s All-in-One Intravital Microscopy (IVM) System IVIM Technology | What? All-in-onesinglebox package for easy installation,operationandmaintenance Co-optimizedH/W and S/W for superbintravitalimagingperformance • Integrateddevices for live animalmaintenance • No limitationin imagingvarious internalorgans • Ultrafast imagingspeed • Live tissue motioncompensation
- 26. WWW.SCINTICA.COM IVIM Technology | Product Lines
- 27. WWW.SCINTICA.COM Confocal Microscopy (IVM-C, -CM) Wide-area Intravital Microscopy
- 28. WWW.SCINTICA.COM Two-Photon Microscopy (IVM-M, -CM, -MS) Sacomere (SHG) Vessel (CD31) Neuromuscular Junction (Thy1-YFP) Skeletal Muscle
- 29. WWW.SCINTICA.COM IVIM Technology | Product Lines Comparison Specifications IVM-C IVM-M IVM-CM IVM-MS Confocal laser Yes No Yes No Tunable two-photon laser No Yes Yes No Fixed two-photo laser No No No Yes (920nm) Fluorescence Detector Confocal Two-Photon Both Two-Photon Scan head Polygonal mirror, Galvano scanner Imaging head Max. 6 objectives FOV 100x100 um^2 / 10x10 mm^2 Motion correction X, Y, Z, and T motion 3D stage range 50,000 x 50,000 x 75,000 um Imaging speed 30 fps @ 512 x 512 pixels (Max. 100 fps), 15 fps @ 1,024 x 1,024 pixels (Max. 50 fps) Fully integrated software Yes
- 30. WWW.SCINTICA.COM IVIM Technology | Filter Sets and Dyes
- 31. WWW.SCINTICA.COM IVIM Technology | Components
- 32. WWW.SCINTICA.COM IVIM Technology | Components
- 33. WWW.SCINTICA.COM Ultrafast Scanning with Uniform Illumination
- 34. WWW.SCINTICA.COM Total Darkness with Black-Out Curtain 34 Sliding black-out curtain protects system from potentially polluting ambient light during imaging
- 35. WWW.SCINTICA.COM Platform for Live Animal Maintenance Body Temperature Monitoring & Feedback Heater Control: Rectal Probe & Body Plate Heater Imaging Tissue Temperature Monitoring & Heater Control: 2 Indicators & Cover Glass Heater
- 36. WWW.SCINTICA.COM Platform for Live Animal Maintenance Gas anesthesia ready with connector ports for gas and exhaust tubing
- 37. WWW.SCINTICA.COM Platform for Live Animal Maintenance Movable stage that connects to animal platform
- 38. WWW.SCINTICA.COM Customizable Objective Lens for Optimal Access Full 3-axis (X, Y, Z) Rotation of OBJ lens: +70° to -70° X-Axis Rotation (roll) Y-Axis Rotation (pitch) Z-Axis Rotation (yaw) Axial Translation of OBJ (Fine Focusing, Fast Z-stack)
- 39. WWW.SCINTICA.COM Customizable Objective Lens for Optimal Access Full 3-axis (X, Y, Z) Rotation of OBJ lens: +70° to -70°
- 40. WWW.SCINTICA.COM Software | 3D rendering Confocal Microscopy (IVM-C), Z-stack Adiponectin-Cre x mTmG SF44 Adiponectin-Cre x mTmG SF44 3D Rendering, Z-stack In Vivo,Inguinaladiposetissue
- 41. WWW.SCINTICA.COM Software | Animal Motion Compensation Real-time Intravital Microscopy (Video-rate imaging) Heart
- 42. WWW.SCINTICA.COM Software | Animal Motion Compensation
- 43. WWW.SCINTICA.COM Software | Animal Motion Compensation Real-time Intravital Microscopy (Video-rate imaging) No Motion Compensation Motion Compensation Liver
- 44. WWW.SCINTICA.COM Software | Animal Motion Compensation Real-time Intravital Microscopy (Video-rate imaging) No Motion Compensation Motion Compensation Lung
- 45. WWW.SCINTICA.COM IVIM Technology | IVM Product & Imaging Service All-in-One IntraVital Microscope (IVM) Sample IntraVital Imaging R&D Service BM cell (DsRed) Histone (H2B-GFP) BM Sinus (CD31) KRS protein Neutrophil (LysM-GFP) Neutrophil (Ly6G) Blood Circulation (FITC-Dextran)
- 46. WWW.SCINTICA.COM 46 Conclusions First all-in-one confocal and/or two-photon microscopes optimized for in vivo imaging Very small footprint with easy installation and integration into any lab space Equipped with a superb live animal maintenance platform and ultrafast scanner Optimized to image a great variety of internal organs Capable of real-time motion compensation
- 47. WWW.SCINTICA.COM 47 Key Research Applications
- 48. Oncology
- 49. WWW.SCINTICA.COM Longitudinal Cancer Xenograft Imaging Intravital Imaging of dorsal skinfold chamber for cancer xenograft imaging Objective Lens Heating Pad Rectal Probe Chamber holder Dorsal Skinfold Chamber Dorsal Skinfold Chamber - Cancer Xenograft Imaging (H460, Human non-small cell lung carcinoma) Day 0 Day 4 SC. Injection of H460-GFP (Human Lung Cancer; NSCLC) H460-GFP (0.1 mil / 20 μL) Intravenous Injection of Tetramethyl-rhodamine dextran TAMRA dextran H460-GFP Vessel (TMR-dextran)
- 50. WWW.SCINTICA.COM Longitudinal Cancer Xenograft Imaging Intravital Imaging of dorsal skinfold chamber for cancer xenograft imaging No Treatment Anti-Angiogenic Treatment Day7 Day10 Day13 Day10 Day13 Day7 LLC-GFP Vessel (CD31) 50 μm Objective Lens Heating Pad Rectal Probe Chamber holder Dorsal Skinfold Chamber Dorsal Skinfold Chamber - Monitoring of anti-angiogenic treatment effect in vessel morphology and dilation
- 51. WWW.SCINTICA.COM Longitudinal Cancer Xenograft Imaging Intravital Imaging of dorsal skinfold chamber for cancer xenograft imaging Dorsal Skinfold Chamber - Monitoring of nanoparticle delivery to triple-negative breast cancer, MDA-MB-231 50 μm 2 hour 6 hour 24 hour MDA-MB-231-GFP Nanoparticle Vessel (CD31) Objective Lens Heating Pad Rectal Probe Chamber holder Dorsal Skinfold Chamber
- 52. WWW.SCINTICA.COM Longitudinal Cancer Xenograft Imaging Intravital Imaging of dorsal skinfold chamber for cancer xenograft imaging Dorsal Skinfold Chamber - Monitoring of nanoparticle delivery to triple-negative breast cancer, MDA-MB-231 50 μm 2 hour 6 hour 24 hour MDA-MB-231-GFP Nanoparticle Vessel (CD31) Objective Lens Heating Pad Rectal Probe Chamber holder Dorsal Skinfold Chamber
- 53. Disease & Drug Development
- 54. WWW.SCINTICA.COM Real-time Abdominal Imaging Abdominal Window Chamber Monitoring of cellular behavior in kidney, pancreas, and spleen Cover glass Kidney Spleen Abdominal Imaging chamber Pancreas Islet cell (MIP-GFP) Vessel (Evans Blue) MIP-GFP mouse - endogenously expresses GFP in pancreatic beta-cells under the control of mouse insulin 1 promotor Monocyte/DC (CX3CR1- GFP) Vessel (TAMRA-dextran) CX3CR1-GFP mouse - endogenously expresses GFP in monocytes, macrophage, brain microglia and DCs under control of endogenous Cx3cr1 locus Pancreas Spleen 100 μm Kidney Islet cell (MIP-GFP) Vessel (Evans Blue)
- 55. WWW.SCINTICA.COM Real-time Gastric Imaging Intravital Imaging of colon cancer implanted in caecum 250 µm Cancer (CT26:H2B-GFP) Vessel (CD31) Cancer (CT26:H2B-GFP) Vessel (CD31) Vessel (CD31)
- 56. WWW.SCINTICA.COM Intravital Imaging of Liver Disease Biomed. Opt. Express 11(8):4835 (2020) Hepatic Lipid Droplet (SF44) Liver Sinusoid (CD31) Portal Vein Periportal Sinusoid
- 57. WWW.SCINTICA.COM Intravital Imaging of Liver Disease Normal diet MCD, 2 days MCD, 7 days MCD, 14 days Hepatic Lipid Droplet (SF44) Liver Sinusoid (CD31) Intravital Imaging of Non-alcoholic Fatty Liver Disease (NAFLD) Biomed. Opt. Express 11(8):4835 (2020)
- 58. Immunology
- 59. WWW.SCINTICA.COM Heating pad Objective Lens Motorized XYZ translational stage Heating pad sensor Rectal probe: body temp. monitoring Cover glass holder Dynamic Immune Cell Imaging In Vivo Blood Vessel Imaging - Endothelial cell labeled in vivo by intravenous injection of anti- CD31 antibody conjugated with far-red fluorophore LysM-GFP mouse - endogenously expresses green fluorescence protein (GFP) in the neutrophil and macrophage by genetically knocking eGFP gene into the lysozyme M (LysM) locus Intravital Imaging of ear skin Neutrophil/Macrophage (LysM-GFP) Vessel (CD31) J. Cell Biology, 216(7):2201 (2017)
- 60. WWW.SCINTICA.COM Dynamic Immune Cell Imaging KARS protein - KARS protein labelled with far-red fluorophore Alexa647 was intradermally injected by using microinjector 3 hurs after KARS injection 6 hours after KARS injection J. Cell Biology, 216(7):2201 (2017) KARS protein Neutrophil/Macrophage (LysM-GFP) LysM-GFP mouse - endogenously expresses green fluorescence protein (GFP) in the neutrophil and macrophage by genetically knocking eGFP gene into the lysozyme M (LysM) locus
- 61. WWW.SCINTICA.COM Real-time Lymphatics Imaging Heating pad Popliteal lymph node Cover glass Warm water circulator Rectal probe: body temp. monitoring Temperature Sensor Objective Lens Cover glass holder Motorized XYZ translational stage Tail vein catheter Intravital Imaging of popliteal lymph node Parenchyma HEV Lumen Intravital imaging of extravasation of T cells & B cells in the high endothelial venule (HEV) of Lymph Node • T cell & B cell obtained from actin-DsRed & actin-GFP mice then adoptively transferred to wildtype C57BL/6 mouse • FRC labeled by anti-ER-TR7 antibody conjugated with Alexa Fluor 647 • HEV Lumen labeled by IV injection of 2MD FITC-Dextran
- 63. WWW.SCINTICA.COM Longitudinal Transplanted Cell Imaging Day 1 Day 3 Day 4 1 mm Heating pad Cover glass Cranium Stereotaxic Instrument: Mouth Adapter, Ear Bar Objective Lens Motorized XYZ translational stage Coronal suture Central vein Sagittal suture Cranium Transplanted cell Vessel (CD31) Longitudinal repetitive wide-area intravital imaging of cranial bone marrow after bone marrow transplantation of c-kit+ BM cell (DsRed) Intravital Imaging of cranial bone marrow PLoS ONE, 12(11):e0187660 (2017)
- 64. WWW.SCINTICA.COM Longitudinal Transplanted Cell Imaging H2B-GFP / β-actin-DsRed mouse - expresses green fluorescence protein (GFP) in the nucleus and DsRed in cytoplasm Longitudinal repetitive wide-area intravital imaging of cranial bone marrow after bone marrow transplantation of c-kit+ BM cell (DsRed) Day 1 Intravital Imaging of cranial bone marrow Day 3 Day 4 1 mm Transplanted cell Vessel (CD31) PLoS ONE, 12(11):e0187660 (2017)
- 65. Infection
- 66. WWW.SCINTICA.COM Real-time Thoracic Imaging Intravital Imaging of lung Stabilized lung imaging - Suction-assisted lung stabilization imaging window chamber Illuminating fiber Ventilator Lung Imaging Window Chamber Heating Pad Motorized XYZ translational stage Pulse Oximetry Objective Lens Homeothermic Controller Temperature Sensor Chamber Titling Mount Protected Ag Mirror Cover glass Mouse Intubation Imaging chamber holder Tilting mount Lung Lung Imaging Window chamber Suction tube Suction hole
- 68. Globally linking scientists with precision tools for research through expertise in science, engineering and support INFO@SCINTICA.COM WWW.SCINTICA.COM
Hinweis der Redaktion
- IVIM Technology’s All-in-One intravital confocal/two-photon microscopy system (IVM-C/M/CM/MS) is extensively optimized and carefully engineered to provide superb performance in the intravital imaging of live animal models in vivo
- Considerations for Optical Imaging: Better resolution at the cellular level than traditional in vivo methods (ex: MRI, CT) Other imaging techniques provide better information at the level of the organ (larger scale) General rule of thumb (any form of imaging): maximum depth ~ 10-100 x spatial resolution
- This slide shows the tissue penetration of the different colors in the spectrum ranging from UV to IR light, and shows that Hb absorbs light maximally in the blue-green-yellow wavelengths, while water absorbs maximally in IR wavelengths, which means the optimal imaging window for in vivo is for light in the orange/red wavelengths of the visible spectrum and NIR wavelengths, meaning between 600 and 1000 nm. This is important to know when selecting fluorophores in fluorescence imaging since fluorophores have to be excited by distinct wavelengths and will emit at a distinct wavelength as well.
- Use of fluorophores makes fluorescence imaging Specific Sensitive Dynamic processes Multiplexing Fluorescent proteins can be incorporated into model or introduced externally Endogenous species = autofluorescence!
- To excite your chosen fluorophore specifically, you need to use the right wavelength that has the right amount of enegry to bring the electron from ground state to excited state. Calculates for you how much the excitation wavelength for one fluorophore potentially also excites a different fluorophore if they are too close to each other in the spectrum
- So you have chosen your fluorophores that you know can be distinguished from one another, but how will you actually achieve separating and imaging them? That is where fluorescence microscopes come into play and they exist of specific components to make that happen: a dichroic mirror and filter sets. 2 specialized fluorescence microscopes most commonly used in intravital imaging are the confocal and the two-photon microscope. They distinguish themselves from basic fluorescence microscopes such as the one in the previous slide by focusing a laser beam into the sample and scanning there point by point rather than illuminating the whole sample at once, thus increasing spatial resolution drastically. In order to collect the emitted light from the illuminated scanned area, the confocal microscope makes use of a pinhole before the detector. This pinhole rejects all photons emitted from outside the focus, as well as those that come from the focus but are scattered on their way to the detector. This way, only unscattered photons coming from the focal plane are able to pass through the pinhole (e.g., photon #5) and contribute to the signal. This inefficiency demands a high laser power for imaging though, which could create unwanted photodamage (such as photobleaching and phototoxicity) and limits imaging depth to a max of 100 μm. In contrast, a 2-photon microscope does not require a pinhole since all emitted photons contributing to the signal come only from the focal point of the excitation spot (e.g., photon #5), regardless of how much they scatter on their path to the detector. Greater tissue penetration up to 500 um is reached with 2-photon microscopy and fewer excitation events are required to achieve the same signal due to the improved collection efficiency, limiting potential photodamage. Confocal: fluorescence from out of focus planes Two-photon: fluorescence from focal spot only
- To understand why the signal in two-photon microscopy comes only from the focal point of the excitation spot, we need to have a look at the principles of two-photon microscopy, which differ from single-photon. Single photon excitation requires the absorption of a high energy photon to excite an electron of a fluorophore to an excited state. When this electron relaxes to its ground state, it emits a photon of light of a different wavelength than the excitation photon. In two-photon microscopy, reaching the excited state of an electron is achieved by the simultaneous absorption of two photons. The likelihood for this simultaneous absorption is basically only reachable in the focal point of the laser beam, which is why there is no out of focus fluorescence from scattered photons, eliminating the need for a pinhole. Since 2 low energy photons with a long wavelength are used to create the energy needed to excite the fluorophore, the penetration depth will be higher.
- IntraVital Microscopy (IVM) enables dynamic 3D imaging of various cellular-level dynamics such as cell trafficking, cell-cell interaction and cell-microenvironment interaction inside the living body in vivo, providing a new insight in the processes of human disease development In situ Spatial information Real time Longitudinal Quantitative Reduce animals
- Endoscopy!
- coverslip thickness of #1.5 is usually required
- applications stem cell biology oncology drug development
- applications immunology infection inflammation
- Motion Analysis (Speed, Acceleration, Track length) Cells (Volume, Vesicles per Cell, Distance to Membrane, Vesicles per Nucleus, Distance to Cell Center) Spots (Number (count), Position (x, y, z), Distance between Spots, Distance to Surface object) Surfaces (Area, Volume, Intensity, Elipcity, Sphericity) Vessels (Area, Density, Number of branches, Number of terminal points, Length, Volume) Neuro (Dendrites (length, branch angle, density), Filaments (branch points, terminal points), Spines (length, volume, diameter) Imaris/Image J/Fiji/In house software
- Lung
- gaining more and more experience in microscopy, animal models, window chambers and applying this in various appications
- Conventionally, users have to prepare the each required functions by themselves. It was very difficult and non-optimal solution with low performance. IVIM is the world’s first and only company providing new All-in-One system providing the best intravital imaging performance. Conventional: technically challenging for non-expert users, not optimized for in vivo imaging, hard to standardize, limited reproducibility Limitations: Imaging depth Motion artifacts Anesthetics and temperature control IVIM: IVIM Technology’s All-in-One intravital confocal/two-photon microscopy system) is extensively optimized and carefully engineered to provide superb performance in the intravital imaging of live animal models in vivo
- Korea Advanced Institute of Science and Technology CEO/CTO/Co-Founder Pilhan Kim Education 2000, B.S. Electrical Engineering, Seoul National University 2005, M.S./ Ph.D. Electrical Engineering, Seoul National University Research Experience 2005 – 2010, Research Harvard Medical School, Boston, USA Fellow Massachusetts General Hospital (MGH), Boston, USA Wellman Center for Photomedicine Work Experience 2010 – pres., Associate Prof. KAIST (Korea Advanced Institute of Science & Technology) Graduate School of Medical Science & Engineering 2017 – pres., CEO/CTO IVIM Technology
- All-in-one single box package enables easy installation, operation and maintenance. It has co-optimized hardware and software for superb intravital imaging performance with ultrafast imaging speed and no limitation in imaging various internal organs. IVM: fully integrates key functionalities for in vivo imaging, consists of an animal stage that can accommodate a wide variety of imaging windows, user friendly design sub-micrometer imaging resolution Ultra high speed imaging Up to 4 simultaneous color channels Integrated motion artifact compensation
- IVM-MS: fully integrated 920nm 2-photon laser
- IVM-MS: fully integrated 920nm 2-photon laser
- IVIM’s unique selling point 1
- Graphics processing unit, a specialized processor originally designed to accelerate graphics rendering. GPUs can process many pieces of data simultaneously, making them useful for machine learning, video editing, and gaming applications.
- LIver
- Lung
- Nonalcoholic fatty liver disease (NAFLD) is a rapidly increasing chronic liver disorder worldwide. effective treatment strategy for NAFLD has not yet been established, which has been hampered by the limited understanding of the pathophysiological drivers for NAFLD. IVM observation of hepatic microenvironments over extended periods of time In this work, novel fluorescent lipid droplet labeling dye, Seoul-Fluor 44 (SF44) to visualize individual lipid droplets and anti-CD31 antibody conjugated with Alexa Fluor 647 to visualize microvasculature simultaneously. longitudinally visualized and quantitatively analyzed the gradual accumulation of hepatic LDs and their changes with the liver sinusoid simultaneously during the progression of hepatic steatosis in a cellular resolution up to 21 days in vivo. Spatial distribution of hepatic lipid droplet in MCD diet-induced NASH mouse model.
- Longitudinal intravital imaging of hepatic lipid droplet accumulation in MCD-diet induced NASH mouse model. (a) Representative maximum intensity projection (MIP) images of hepatic LD (yellow, SF44) and sinusoid (cyan, CD31) in the liver of mice fed normal diet or MCD diet. Scale bars, 20 µm.
- LysM-GFP model which possess the enhanced green fluorescent protein (EGFP) inserted in the Lysozyme M (LysM) promoter region (expressed primarily by neutrophils), when used in conjunction with in vivo fluorescence imaging (FLI) provide a means of quantifying neutrophil emigration noninvasively and longitudinally into wounded skin.
- lysyl-tRNA synthetase (KRS), was previously shown to be secreted from cancer cells to induce inflammatory responses In this study, we present the results of our investigation into how KRS is secreted to extracellular space and whether any posttranslational modifications are involved in this process. Time-lapse intravital microscopy showing the recruitment of macrophages/monocytes (green) to the injected naked KRS stained with Alexa Fluor 647 as described in the Intravital imaging section of Materials and methods. This study suggests a novel functional connection between caspase-mediated signaling and a key enzyme for protein synthesis, KRS, which is also active in immune stimulation when secreted into the extracellular space.
- Bone marrow transplantation (BMT) has been widely performed in patients with blood disorders and cancers. However, the cellular-level behaviors of the transplanted bone marrow cells over wide-areas of the host bone marrow after the BMT are not fully understood yet. longitudinal wide-area cellular-level observation of the calvarial bone marrow after the BMT in vivo. Using a H2B-GFP/β-actin-DsRed double-transgenic mouse model as a donor, a subcellular-level nuclear-cytoplasmic visualization of the transplanted bone marrow cells was achieved, which enabled a direct in vivo dynamic monitoring of the distribution and proliferation of the transplanted bone marrow cells.
- Proliferation events observed during the 5 hour time-lapse imaging (S1 and S2 Movies). (B, C) Representative time-lapse image sequences showing (B) proliferation (S3 and S4 Movies) or (C) migration of the transplanted BM cell (S5 Movie). From day 1 to day 4 after the BMT, the transplanted BM cells greatly increased in number and distributed widely where they formed clusters which also increased over time