Multiphoton Imaging Brochure
Transcription
Multiphoton Imaging Brochure
Multiphoton Imaging Multiphoton Microscopes: Rotating and Standard Imaging Platforms The Bergamo II Series is Thorlabs’ platform for multiphoton microscopy. Used to acquire high-resolution, multi-channel images in vivo, ex vivo, and in vitro, our versatile, sensitive microscopes are specifically engineered for exceptional performance under a wide variety of experimental conditions. Overview of the Bergamo II Series Multiphoton Microscopes Thorlabs' multiphoton imaging platforms are designed around various experimental needs. Customize and future-proof your investment with a system specifically tailored to your application. Choose between two body options: Flexibility with the Rotating Body Configuration Best For Rotating Body In Vivo Deep Brain Imaging Performed with Rotating Body (Courtesy of Dr. Hajime Hirase and Katsuya Ozawa, RIKEN Brain Science Institute, Wako, Japan) Applications Advantages u Live Imaging u The Industry’s Only Microscope that Rotates Around the Focal Spot of the Objective – Awake and u Motion in X, Y, Z, and θ to Image Around Orientation-Sensitive Experiments u -5° to +95° or -50° to +50° of Rotation, 2" Travel in X and Y, 5" Microscope Base Travel in Z, and Behaving Animals – Large Animals – Small Animals 1" Objective Travel u 24+ Cubic Inches (393 cm3) of Total Objective Viewing Volume u Maintain Beam Alignment Throughout the Full Range of Motion via Articulating Periscopes Versatility with the Standard Body Configuration Standard Body Best For In Vitro Ex Vivo In Vivo Mouse Retina Imaged with DIC Mouse Retina Imaged with Dodt Contrast Applications Advantages u Imaging u All-in-One Rig for a Wide Array of Applications – Cells – Tissues –Live Small Animals u DIC or Dodt Contrast; Camera or Laser Scanned u High-Sensitivity Forward Fluorescence Detection u Removable Transmitted Light Path to Configure the Microscope on the Fly for In Vivo or In Vitro u Large Working Space Around the Objective to Accommodate Various Samples and Equipment Rotating Bergamo II Microscopes Overview of Options and Benefits Widefield Viewing uS cientific Camera Scan Paths u Galvo-Galvo and/or Galvo-Resonant uE xcitation Path Optimized for 400 - 1100 nm or 680 - 1600 nm Wavelength Range Rotational Configuration u - 5° to +95° u Thorlabs’ Epi-Illuminator with High-Power LED u -50° to +50° Dichroic Changers u Manual or Computer Controlled Piezo Objective Focus Control u High-Speed Z Stacks PMT Detection Module u Up to Four Channels u Microscope Controller u Advanced 5-Axis Controller with Touchscreen With or Without Shutters Beam Conditioning Modules u Variable Beam Expander u Pockels Cell u Variable Attenuator Epi-Detection Module u Three Collection Angle Options: 8°, 10°, and 14° u Easy-to-Exchange Filters u Emission Detection Optimized at 400 - 800 nm Features Details u -5° to +95° or -50° to +50° (Factory Configured) Rotational Configuration – Image Around Orientation-Sensitive Experiments – Optical Alignments are Maintained Throughout the Full Range of Motion u Galvo-Resonant – Image at 2 fps (4096 x 4096 Pixels), 15 fps (1024 x 1024 Pixels), 30 fps (512 x 512 Pixels), and Up to 400 fps (512 x 32 Pixels) u Galvo-Galvo Scan Paths – Custom Scan Geometries: Squares, Rectangles, Circles, Ellipses, Lines, and Polylines – Capture Weak Signals with Long Dwell Time Integration – Achieve Consistent Dwell Times Across Field of View u Super Broadband Optics Optimized for 400 - 1100 nm or 680 - 1600 nm Excitation Wavelengths – Designed for the Latest Widely Tunable Ti:Sapphire and OPO Systems u Up to Four Channels PMT Modules – Cooled or Non-Cooled PMTs – Configure Up to Four Channels with Simultaneous Detection u Choice of Signal Collection Angle to Accommodate Different Sample Depths Epi-Detection Modules – 8° (Two PMTs) – 10° (With or Without Shutters and Up to Four PMTs) – 14° (With or Without Shutters and Up to Two PMTs) (Angles Quoted When Using an Objective with a 20 mm Entrance Pupil) u Thorlabs or Third-Party Scientific Cameras Widefield Viewing – Visualize Samples with Fluorescence or Reflected Light – Compatible with C-Mount-Threaded Scientific Cameras u Single- or Multi-Filter-Cube Epi-Illuminator – Locate Areas of Interest Without Unnecessary Laser Excitation – Thorlabs’ LEDs Provide Bright Illumination – Industry-Standard Liquid Light Guides Illuminate Across the Visible Spectrum u Piezo Objective Focus Control Objective Focus Control – Fast Z Stacks – Nanometer-Level Axial Resolution u Choice of Sample Holders Sample Holders – Rigid Stand with Platform, Slide Holder, or Recording Chamber – XY Platform with Micromanipulators u Advanced 5-Axis Controller with Touchscreen Microscope Controller – Control X, Y, Z (Base), Z (Objective), and Rotation Angle – Speed Control, Saved Positions, and the Convenience of a Touchscreen Interface u Variable Beam Expander – 1X - 3X Beam Diameter Modulation into the Objective Back Aperture Using Software Control u Pockels Cell Beam Conditioning Modules – Edge and Fly-Back Blanking to Minimize Sample Photobleaching – High-Speed Masking for ROIs – Customize Laser Power at Each Slice Using Software Control u Variable Attenuator – Improves Pockels Cell Performance – One-Click Shutter – Manual and Computer Control of Laser Power in Systems Without a Pockels Cell u ThorImageLS™ Software Acquisition Packages – – – – Designed for Seamless Integration with Thorlabs’ Imaging Systems Powerful and Intuitive Interface Direct Support from Software Engineers Open Source for Custom Features and Participation with the Life Science Community u ScanImage and LabVIEW – Large User Base Standard (Z and XYZ Motion) Bergamo II Microscopes Overview of Options and Benefits Widefield Viewing uS cientific Camera u Thorlabs’ Epi-Illuminator with High-Power LED Scan Paths u Galvo-Galvo and/or Galvo-Resonant u Binoculars or Trinoculars uE xcitation Path Optimized for 400 - 1100 nm or u Hold up to Six Filter Cubes 680 - 1600 nm Wavelength Range Beam Conditioning Modules Piezo Objective Focus Control u High-Speed Z Stacks u Variable Beam Expander u Pockels Cell u Variable Attenuator Detection Modules u Forward and Epi-Detection Modules u Easy-to- Exchange Filters PMT Detection Module u Up to Four Channels u Microscope Controllers u Advanced 5-Axis Controller with Touchscreen u 3-Axis Controller With or Without Shutters Contrast Imaging u Dodt Contrast u Differential Interference Contrast (DIC) Sample Holders u Rigid Stand with Platform u Rigid Stand with Slide Holder or Recording Chamber u XY Platform with Micromanipulators Features Details u Galvo-Resonant – Image at 2 fps (4096 x 4096 Pixels), 15 fps (1024 x 1024 Pixels), 30 fps (512 x 512 Pixels), and Up to 400 fps (512 x 32 Pixels) u Galvo-Galvo Scan Paths – Custom Scan Geometries: Squares, Rectangles, Circles, Ellipses, Lines, and Polylines – Capture Weak Signals with Long Dwell Time Integration – Achieve Consistent Dwell Times Across Field of View u Super Broadband Optics Optimized for 400 - 1100 nm or 680 - 1600 nm Excitation Wavelengths – Designed for the Latest Widely Tunable Ti:Sapphire and OPO Systems u Up to Four Channels PMT Modules – Cooled or Non-Cooled PMTs – Configure Up to Four Channels with Simultaneous Detection u Choice of Signal Collection Angle to Accommodate Different Sample Depths Epi-Detection Modules – 8° (Two PMTs) – 10° (With or Without Shutters and Up to Four PMTs) – 14° (With or Without Shutters and Up to Two PMTs) (Angles Quoted When Using an Objective with a 20 mm Entrance Pupil) u Thorlabs or Third-Party Scientific Cameras Widefield Viewing – Visualize Samples with Fluorescence or Reflected Light – Compatible with C-Mount-Threaded Scientific Cameras u Single- or Multi-Filter-Cube Epi-Illuminator – Locate Areas of Interest Without Unnecessary Laser Excitation – Thorlabs’ LEDs Provide Bright Illumination – Industry-Standard Liquid Light Guides Illuminate Across the Visible Spectrum u Piezo Objective Focus Control Objective Focus Control – Fast Z Stacks – Nanometer-Level Axial Resolution u Choice of Sample Holders Sample Holders – Rigid Stand with Platform, Slide Holder, or Recording Chamber – XY Platform with Micromanipulators u Advanced 5-Axis Controller with Touchscreen Microscope Controller – Control X, Y, Z (Base), Z (Objective), and Rotation Angle – Speed Control, Saved Positions, and the Convenience of a Touchscreen Interface u 3-Axis Controller – Control X, Y, Z Axes with Speed Adjustment u Dodt Contrast and DIC Modules Contrast Imaging – Removable Modules to In Vivo or In Vitro Configurations – View Features in Tissue Slices with Dodt Contrast or Laser-Scanned Dodt Contrast – View Features in Thinner, Transparent Samples with DIC Imaging or Laser-Scanned DIC Imaging u Variable Beam Expander – 1X - 3X Beam Diameter Modulation into the Objective Back Aperture Using Software Control u Pockels Cell Beam Conditioning Modules – Edge and Fly-Back Blanking to Minimize Sample Photobleaching – High-Speed Masking for ROIs – Customize Laser Power at Each Slice Using Software Control u Variable Attenuator – Improves Pockels Cell Performance – One-Click Shutter – Manual and Computer Control of Laser Power in Systems Without a Pockels Cell u ThorImageLS™ Software Acquisition Packages – – – – Designed for Seamless Integration with Thorlabs’ Imaging Systems Powerful and Intuitive Interface Direct Support from Software Engineers Open Source for Custom Features and Participation with the Life Science Community u ScanImage and LabVIEW – Large User Base Innovations in Detail Image with Speed and Accuracy with Galvo-Resonant and/or Galvo-Galvo Scan Paths Rotating Body Standard Body Bergamo II multiphoton microscopes can be configured with a galvo-resonant scan path, a galvo-galvo scan path, or both, depending upon the needs of the experiment. Equip both types of scan paths to get the best of both worlds. Scan at up to 400 fps at 512 x 32 pixels using our galvo-resonant scanners. Achieve uniform images with edge blanking and protect samples with high-speed masking in tandem with our Pockels cells. Alternatively, use our galvo-galvo scanners to accurately photoactivate or photobleach specific areas of the sample with preset (line, square, rectangle, circle, and ellipse) or custom-drawn geometries. Galvo-Resonant Scanner FPS 512 x 32 Frames per Second 400 300 200 100 Primary Galvo-Resonant Scan Path Secondary Galvo-Galvo Scan Path Single-Photon Fiber Laser in Secondary Scan Path 512 x 512 0 Pockels Cell Edge Blanking Pockels Cell Masking Square/Rectangle Circle/Ellipse Line Polyline Galvo-Galvo Scan Modes Superior Photodetection Systems Rotating Body Standard Body Bergamo II systems can be configured with up to four PMT detection channels and are fully compatible with industry-standard fluorescence emission filters. Our unique design makes it simple and quick to exchange filters without having to worry about maintaining optical alignment. Capture weak fluorescence signals or protect highly photosensitive samples using high-sensitivity, high-quantum-efficiency GaAsP PMTs. Full software integration of the protection circuit ensures maximum PMT longevity. Sensitivity Quantum Efficiency 1000 100 10 Sensitivity (mA/W) Quantum Efficiency (%) 100 1 10 1 0.1 0.1 100 200 300 400 500 600 700 800 900 100 200 300 400 Wavelength (nm) Super Broadband Scan Optics 500 600 700 800 900 Wavelength (nm) Rotating Body Standard Body Bergamo II microscopes feature proprietary scan optics that are optimized for 400 - 1100 nm or 680 - 1600 nm excitation wavelengths, allowing single or multiphoton excitation of a wide variety of fluorophores. Designed with the latest widely tunable Ti:Sapphire lasers and OPO systems in mind, the Bergamo II platform future-proofs your investment. Image more by taking full advantage of the optical designs used in low-magnification, high-numerical aperture objectives that have become the standard in multiphoton objectives. Capture large sample areas with the Bergamo II’s maximum field number (FN) of 20. Customizable Software Options In addition to hardware versatility, Thorlabs offers a selection of software acquisition packages. Choose either the ThorImageLS™ software package, which is designed in parallel with the Bergamo II Multiphoton Microscope, or the popular ScanImage software from Vidrio Technologies. Regardless of the selection, Thorlabs will install and test the performance of your system with the chosen software platform. Rotating Body Standard Body ThorImageLS™ Custom Open-Source ThorImageLS™ ScanImage Innovations in Detail Imaging Optimization using the Variable Beam Expander Rotating Body Standard Body Utilize the Variable Beam Expander to adapt to and optimize the experimental conditions. Match the beam diameter to the back aperture of the objective to utilize the entire NA, overfill to achieve more even illumination, or underfill for higher power through thick samples. The software-controlled variable beam expander adjusts the input beam diameter from 1X to 3X. Beam Diameter Matches Back Aperture, which Uses the Entire NA Back Aperture is Overfilled for More Even Illumination Back Aperture is Underfilled, Leading to Higher Power in Thick Samples Large-Collection-Angle Optics Rotating Body Standard Body A high photon collection efficiency in the PMTs is made possible by a wide collection angle while simultaneously minimizing the distance between the collecting lens and the sample. This system design allows Bergamo II Series Multiphoton Microscopes to achieve high sensitivity and a high signal-to-noise ratio for deep physiological imaging. To PMTs Collecting Lens Objective Sample Plane Advanced Multi-Axis Controller with Touchscreen Rotating Body Specifically designed for the rotating Bergamo II Microscope, this controller allows individual control of up to five directions of movement. In addition to the standard X, Y, and Z axes, the controller also provides fine movement along the optical axis of the objective. The objective can be factory configured to rotate from -5° to 95° or -50° to 50°. The integrated touchscreen allows two quick memory settings, while the ThorImageLS™ image acquisition software suite can save up to eight positions, allowing convenience and enabling experimental repeatability. z z z Rotation Angle, θ y + x x x y y Objective Axis Superior Engineering for a Flexible, Rotating Platform Base Axis Rotating Body The Bergamo II Multiphoton Microscope maintains alignment during rotation around the sample through multi-jointed articulating periscopes. Our periscopes' engineering and design offer enhanced flexibility to allow the entire scanning system to rotate around the sample while still maintaining laser alignment into the scan head. 2" 2" 6" 24+ Cubic Inches of Accessible Volume Multi-Jointed Articulating Periscopes Maintain Alignment into Galvo-Resonant and Galvo-Galvo Scanners Innovations in Detail Adaptability for Specialized Objectives Rotating Body Standard Body Bergamo II microscopes directly accept infinity-corrected objectives that have M32 threads, while also having highly stable mechanical adapters for M34, M25, and RMS threads. Together, these options encompass the vast majority of low-magnification, RMS M25 M32 high-NA objectives. Our scan optics are designed with these specialized objectives in mind to ensure that the microscope is always able to handle light collection over a large field of view. M34 Highly Sensitive Scientific-Grade Cameras Rotating Body Standard Body Thorlabs' scientific cameras are designed to locate point-of-reference markers in specimens being studied with Bergamo II microscopes. Their low noise and high quantum efficiency make it easy to find fluorescence labels and provide the high sensitivity needed to obtain contrast in thick, unlabeled specimens using DIC or Dodt contrast. These cameras operate independently from the multiphoton laser scanning apparatus, protecting delicate samples by minimizing their exposure to laser light. By combining epi-fluorescence images with images obtained by DIC, Dodt contrast, and laser scanning, co-registered, multimodal pictures that yield deeper insights into your specimen are created. 1.4 Megapixel Scientific-Grade Camera Thorlabs' Customer Advantages Our Approach Advantages u Our Systems Are Engineered at a Single Location, Maximizing Efficiency and Ensuring Cost Fully Designed and Manufactured In House Savings u In-House Expertise in Every Component and System Modular System Construction Professional Installation u Future-Proof Your Investment and Upgrade as Needs Evolve Without Sacrificing Existing Capabilities u Thorlabs' Technicians Meticulously Assemble, Test, and Verify Each System to Meet Our High Standards of Quality and Performance u Receive Support for Your Experimental Needs from Our Life Science Application Engineers Unmatched Professional Support u Receive Direct Support from Engineers Who Designed the Bergamo II Series Multiphoton Microscopes u Receive Direct Support from Software Engineers Who Wrote ThorImageLS™ u Receive Support via Video Teleconferencing (Receive a Camera and Microphone from Quick Support Thorlabs if Needed) u Let Thorlabs Do the Work! With Your Permission, Thorlabs Can Remotely Access Your Computer to Address a Software-Related Issue Specifications Configuration Laser Scanning Scan Path Wavelength Range u 400 - 1100 nm or 680 - 1600 nm ✓ ✓ Field of View u 20 mm Diagonal Square (Max) at the Intermediate Image Plane ✓ ✓ Scanners u Single or Dual Scan Paths: Galvo-Resonant, Galvo-Galvo, or Both ✓ ✓ Galvo-Resonant Scan Speed u 30 fps at 512 x 512 Pixels; 400 fps at 512 x 32 Pixels ✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ ✓ Compatible u M34 x 1.0, M32 x 0.75, M25 x 0.75, and RMS Objective Threading ✓ ✓ Epi-Detection u Up to 4 GaAsP PMTs, Cooled or Non-Cooled ✓ ✓ Forward Detection u 2 GaAsP PMTs, Non-Cooled Collection Optics u 8°,10°, or 14° Collection Angle (Angles Quoted When Using an Objective with a 20 mm Entrance Pupil) Galvo-Galvo Scan Speed u 3 fps at 512 x 512 Pixels; 48 fps at 512 x 32 Pixels; 70 fps at 32 x 32 Pixels Galvo-Galvo Scan Modes u Imaging: Line, Polyline, Square, or Rectangle Scan Zoom u 1X - 16X (Continuously Variable) Scan Resolution Detection Rotating Standard Body Body u Pixel Dwell Time: 0.4 - 20 µs u Non-Imaging: Circle, Ellipse, Polygon, or Point u Bi-Directional: 2048 x 2048 Pixels (Max) u Unidirectional: 4096 x 4096 Pixels (Max) ✓ ✓ ✓ ✓ ✓ u User-Changeable Emission Filters and Dichroic Cubes Widefield Viewing Transmitted Light Imaging u Manual or Motorized Switching Between Scanning and Widefield Modes u Thorlabs’ High-Power LED or Liquid Light Guide for Illumination u C-Mount Threads for Scientific Cameras u Widefield and (Laser-Scanned) Dodt Contrast or Differential Interference Contrast (DIC) u Visible and/or NIR LED Illumination ✓ u Air or Oil Immersion Condenser X and Y u 2" (50.8 mm) Total Travel; 0.5 µm Encoder Resolution ✓ ✓ Objective Axis u 1" (25.4 mm) Total Travel; 0.1 µm Encoder Resolution ✓ ✓ Base Axis (Z) u 5" (127 mm) Total Travel; 1 µm Encoder Resolution ✓ Translation Rotation u -5° to + 95° or -50° to +50° Around Objective Focus u 0.1° Encoder Resolution ✓ Selected Publications Using Thorlabs' Imaging Systems Kato HK, Gillet SN, Peters AJ, Isaacson JS, and Komiyama T. "Parvalbumin-expressing interneurons linearly control olfactory bulb output." Neuron. 2013 Dec 4; 80 (5): 1218-31. Lu W, Tang Y, Zhang Z, Zhang X, Yao Y, Fu C, Wang X, and Ma G. "Inhibiting the mobilization of Ly6C(high) monocytes after acute myocardial infarction enhances the efficiency of mesenchymal stromal cell transplantation and curbs myocardial remodeling." Am J Transl Res. 2015 Mar 15; 7 (3): 587-97. Ehmke T, Nitzsche TH, Knebl A, and Heisterkamp A. "Molecular orientation sensitive second harmonic microscopy by radially and azimuthally polarized light." Biomed Opt Express. 2014 Jun 12; 5 (7): 2231-46. Boyd AM, Kato HK, Komiyama T, and Isaacson JS. "Broadcasting of cortical activity to the olfactory bulb." Cell Rep. 2015 Feb 24; 10 (7): 1032-9. Liu J, Wu N, Ma L, Liu M, Liu G, Zhang Y, and Lin X. "Oleanolic acid suppresses aerobic glycolysis in cancer cells by switching pyruvate kinase type M isoforms." PLoS One. 2014 Mar 13; 9 (3): e91606. Cai F, Yu J, Qian J, Wang Y, Chen Z, Huang J, Ye Z, and He, S. "Use of tunable second-harmonic signal from KNbO3 nanoneedles to find optimal wavelength for deep-tissue imaging." Laser & Photon Rev. 2014 8: 865-874. Peters AJ, Chen SX, Komiyama T. "Emergence of reproducible spatiotemporal activity during motor learning." Nature. 2014 Jun 12; 510 (7504): 263-7. Cossell L, Iacaruso MF, Muir DR, Houlton R, Sader EN, Ko H, Hofer SB, and Mrsic-Flogel TD. "Functional organization of excitatory synaptic strength in primary visual cortex." Nature. 2015 Feb 19; 518 (7539): 399-403. Palmer LM, Shai AS, Reeve JE, Anderson HL, Paulsen O, and Larkum ME. "NMDA spikes enhance action potential generation during sensory input." Nat Neurosci. 2014 Mar; 17 (3): 383-90. Chen SX, Kim AN, Peters AJ, and Komiyama T. "Subtype-specific plasticity of inhibitory circuits in motor cortex during motor learning." Nat Neurosci. 2015 Aug; 18 (8): 1109-15. Jia Y, Zhang S, Miao L, Wang J, Jin Z, Gu B, Duan Z, Zhao Z, Ma S, Zhang W, and Li Z. "Activation of platelet proteaseactivated receptor-1 induces epithelial-mesenchymal transition and chemotaxis of colon cancer cell line SW620." Oncol Rep. 2015 Jun; 33 (6): 2681-8. Takata N, Nagai T, Ozawa K, Oe Y, Mikoshiba K, and Hirase H. "Cerebral blood flow modulation by Basal forebrain or whisker stimulation can occur independently of large cytosolic Ca2+ signaling in astrocytes." PLoS One. 2013 Jun 13; 8 (6): e66525. Partridge JG, Lewin AE, Yasko JR, and Vicini S. "Contrasting actions of group I metabotropic glutamate receptors in distinct mouse striatal neurones." J Physiol. 2014 Jul 1; 592 (Pt 13): 2721-33. Show Us Your Work! We would like to showcase your hard-earned results! Let us know that you published an article with images obtained using a Thorlabs system and gain increased visibility within the Thorlabs and scientific community. By sharing your successes, you can increase the overall awareness of the capabilities of your system while also learning more about what others have done. Transgenic Zebrafish (Courtesy of Katie Kindt, Porter Neuroscience Research Center, National Institutes of Health, Bethesda, MD) Sales Contact Our dedicated Thorlabs sales team is ready to help you achieve your goals. Whether you are seeking more information or are ready to make a purchase, please contact us for immediate assistance. (703) 651-1700 u ImagingSales@thorlabs.com Confocal Microscopy Systems Thorlabs’ Confocal Imaging System can be used to upgrade any research-grade microscope into a confocal workstation. Its high-speed galvo-resonant scan head provides a video frame rate of 30 fps at 512 x 512 pixels and a maximum frame rate of 400 fps at 512 x 32 pixels. Compatible with upright and inverted microscopes, it supports a large field number (FN) of 25. Confocal imaging has several advantages over widefield imaging, including the elimination of out-of-focus light and improved spatial resolution. Combined with multi-channel acquisition, it creates detailed 3D volumetric images of biological samples. 400 µm Live, GFP-labeled hippocampal mouse neuron. Courtesy of the 2015 Cold Spring Harbor summer course: “Imaging structure and function in the nervous system.” Confocal Rig with Fast XY Scanning Stage and Dodt Contrast Worldwide Locations Thorlabs Ultrafast Optoelectronics Thorlabs Ltd. Thorlabs Sweden AB Thorlabs Imaging Systems Thorlabs Vytran Division Thorlabs SAS France Thorlabs China Ltd. Thorlabs Quantum Electronics (TQE) Thorlabs Canada Thorlabs GmbH Thorlabs Japan Thorlabs Scientific Imaging (TSI) Thorlabs Ltda, Brazil Thorlabs, Inc. 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