Opterra. Multipoint Scanning Confocal Microscope. Innovation with Integrity. Cell-Friendly, High-Speed, High-Resolution Imaging

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Caitlin O’Brien’
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1 Opterra Multipoint Scanning Confocal Microscope Cell-Friendly, High-Speed, High-Resolution Imaging Innovation with Integrity Fluorescence Microscopy

Opterra Multipoint Scanning Confocal Microscope Superior Integration, Versatility, and Cell-Friendly Performance The Opterra Multipoint Scanning Confocal Microscope is designed specifically for

The system utilizes a CCD camera as a detector, and can be configured through selectable aperture sizes so the optimal balance of speed and resolution for each experiment can be set to meet your

With its short acquisition times and cell-protecting minimization of photobleaching and phototoxicity, Opterra is ideal for advanced live sample studies, including protein localization and

2 Opterra Multipoint Scanning Confocal Microscope Superior Integration, Versatility, and Cell-Friendly Performance The Opterra Multipoint Scanning Confocal Microscope is designed specifically for live-cell microscopy. It uniquely combines the resolution of traditional confocal systems with the speed typically associated with wide-field imaging. The system utilizes a CCD camera as a detector, and can be configured through selectable aperture sizes so the optimal balance of speed and resolution for each experiment can be set to meet your specific application requirements. Opterra s unconventional optical design makes spectral imaging of dynamic processes possible. With its short acquisition times and cell-protecting minimization of photobleaching and phototoxicity, Opterra is ideal for advanced live sample studies, including protein localization and trafficking, intracellular ion imaging, microtubule and vesicle dynamics, and nuclear structure and dynamics. Uncompromised Performance High-speed, multidimensional imaging of live cells and small organisms Photoactivation module designed specifically for cellular kinetic studies Intuitive and powerful application software

Resolution and Speed Matched to Applications Efficient and Effective Fluorescence Confocal One aspect of Opterra s unique scanner design is a motorized aperture plate that contains pinholes of three

This enables the scientist to match aperture size to the objective and optimize the system for specific applications.

3 Resolution and Speed Matched to Applications Efficient and Effective Fluorescence Confocal One aspect of Opterra s unique scanner design is a motorized aperture plate that contains pinholes of three different sizes and slits of four different widths. Pinholes provide maximum resolution, while slits allow for higher speed acquisitions. This enables the scientist to match aperture size to the objective and optimize the system for specific applications. Plus, aperture selection is controlled by software, so hardware changes are not required for setting up different experiment protocols. Opterra s use of one-dimensional pinhole arrays offers significant advantages over systems based on two-dimensional arrays, such as spinning disks. Opterra s array produces one half or less of the crosstalk of a two-dimensional array. The result is that Opterra produces sharper images with higher resolution at greater depths versus the spinning disk. C. elegans embryo labeled with GFP and mcherry. Mitosis in C. elegans embryo labeled with GFP tubulin. The Best Multi-Pinhole Confocal Solution in the Market Opterra Spinning Disk High-speed operation Reduction in bleaching and phototoxicity compared to point scanning confocals Availability of slits for acquisitions of up to 1,000 frames per second 2D crosstalk between pinholes Improved axial resolution and improved imaging depth with linear pinhole configuration Ability to select different pinhole sizes to match objectives utilized and obtain highest resolution possible with no hardware changes Restriction to only two pinhole sizes with the purchase of additional hardware and requiring a hardware change to switch between pinhole sizes Spectral Imaging at 4 frames per second with 15 spectral channels per frame with a 512x512 pixel image Simultaneous photoactivation and imaging Some Systems Simultaneous photoactivation and imaging with no emission signal loss

Easy, Versatile, High-Performance Data Acquisition Opterra tightly integrates scanner, CCD camera, and motion control devices to provide high-speed, timed volumetric,

Piezo Z-focus can be combined with stage movement for efficient collection of 3D stage montages, as well as timed acquisition at multiple stage locations in individual

Opterra is powered by Prairie View, the same software that drives Bruker s multiphoton microscope systems.

Time lapse, Z-series, stage montage and multidimensional acquisitions can be defined in a matter of seconds.

4 Easy, Versatile, High-Performance Data Acquisition Opterra tightly integrates scanner, CCD camera, and motion control devices to provide high-speed, timed volumetric, 4D imaging. Piezo Z-focus can be combined with stage movement for efficient collection of 3D stage montages, as well as timed acquisition at multiple stage locations in individual sample chambers or multi-well plates. Stage montage of zebrafish. Opterra is powered by Prairie View, the same software that drives Bruker s multiphoton microscope systems. Prairie View provides scientists with a user-friendly interface that allows even the most complex protocols to be set up with ease. Time lapse, Z-series, stage montage and multidimensional acquisitions can be defined in a matter of seconds. Prairie View also enables users to integrate external devices into their protocols by providing a rich environment for creating triggers and analog signals, as well as recording analog signals from other devices. Applications Protein localization and trafficking Vesicle trafficking Mitosis Organelle trafficking Microtubule dynamics Cell migration Intracellular ion concentration FRAP Developmental biology Photoconversion Cell membrane wound healing Photoactivation GFP labeled neurons in zebrafish, max projection of 100 µm Z-series. Max projection of C. elegans labeled with GFP.

Flexible and Feature-Rich Software Multifield Imaging Simplified Prairie View s Atlas Imaging module makes setting up stage montages and other types of multiple stage location paradigms simple and

Easy navigation in X, Y and Z allows the user to quickly find and record individual locations of interest, or define a grid of overlapping locations which can be used to construct a high-resolution

From the quick-scan preview, users can define a montage of overlapping locations for final acquisition.

5 Flexible and Feature-Rich Software Multifield Imaging Simplified Prairie View s Atlas Imaging module makes setting up stage montages and other types of multiple stage location paradigms simple and intuitive. Easy navigation in X, Y and Z allows the user to quickly find and record individual locations of interest, or define a grid of overlapping locations which can be used to construct a high-resolution image of up to an entire specimen. The preview window shows a coarse tiling of all positions scanned, and the live window shows the currently selected field of view. From the quick-scan preview, users can define a montage of overlapping locations for final acquisition. Tiles in areas not containing sample can be turned off, speeding acquisition and saving storage space. Intuitive navigation in the X, Y and Z dimensions is performed by clicking and dragging in the live or preview windows. On the left is a coarse preview scan of many fields of view covering a zebrafish. On the right is a real-time image of the current field of view, highlighted in the green box on the preview window. Mitosis in frog oocytes cell membrane: mtag BFP, tubulin (spindles and membrane): egfp, histone (nuclei): mcherry. Zebrafish heart.

Image + Activate = Discover Simultaneous Confocal Imaging and Photoactivation Opterra s unique optical design allows photostimulation to occur simultaneously with imaging with no loss of image

It can be interfaced with UV and visible lasers, as well as with pulsed IR lasers.

Prairie View software includes a rich set of tools for controlling photostimulation protocols to take full advantage of Opterra s capabilities.

Protocols can be run in a precisely timed synchronous mode, or in an interactive asynchronous mode. Intuitive interfaces make both basic and complex protocols simple to set up and run.

6 Image + Activate = Discover Simultaneous Confocal Imaging and Photoactivation Opterra s unique optical design allows photostimulation to occur simultaneously with imaging with no loss of image emission signal. The optional photoactivation module uses the same powerful scanning technology found in Bruker s multiphoton systems. It can be interfaced with UV and visible lasers, as well as with pulsed IR lasers. Opterra provides a comprehensive solution for FRAP, ablation, photo-damage, photoconversion, optogenetics, and uncaging. Photoconversion of Dendra in glioma cell using masked area. Prairie View software includes a rich set of tools for controlling photostimulation protocols to take full advantage of Opterra s capabilities. Activation patterns include points, lines, rectangles, arbitrary shapes and high speed spiral activation. Protocols can be run in a precisely timed synchronous mode, or in an interactive asynchronous mode. Intuitive interfaces make both basic and complex protocols simple to set up and run. Point photostimulation applications are run from Prairie View s Mark Points interface, which is ideal for applications such as uncaging and optogenetics. Mark Points also includes a robust tool for photobleaching and ablation experiments that require individual points or lines. For larger areas, the Spiral Activation feature enables high-speed stimulation of circular areas. Scientists also can define custom-shaped regions with the Photoactivation Mask interface. Multiple laser lines can be employed within each protocol so that more than one type of photosensitive molecule can be used in a single sample. Photobleaching of GFP-labeled tubulin in HeLa cells using masked area. Photobleaching of GFP-labeled tubulin in HeLa cells using lines. Uncaging fluorescein with Spiral Activation, sequential frames at 50 fps.

Dynamic Spectral Scanning for Live Cell Imaging Opterra s Spectral Scanning feature performs spectral imaging at

By incorporating an Amici prism into the Opterra image scanner, full spectral data sets of 15 channels can be

Prairie View software provides real time processing of spectral data sets to provide a live image in which up to

Spectral Scanning also can be used with time lapse, Z-series, and stage control.

3D stage montages, and acquisitions from multiple stage locations.

replayed in the Prairie View software or loaded into third party programs.

Activation. Setup shown for photomanipulation of arbitrary areas.

7 Dynamic Spectral Scanning for Live Cell Imaging Opterra s Spectral Scanning feature performs spectral imaging at speeds appropriate for live cell and small organism imaging. By incorporating an Amici prism into the Opterra image scanner, full spectral data sets of 15 channels can be acquired at speeds of up to four spectral data sets per second for a 512x512 pixel image. Prairie View software provides real time processing of spectral data sets to provide a live image in which up to four channels of data can be visualized. Spectral Scanning also can be used with time lapse, Z-series, and stage control. This provides a full range of multidimensional data acquisition methods, including time-based volumetric imaging, 3D stage montages, and acquisitions from multiple stage locations. Spectral data sets are saved as individual OME-TIFF images, each representing one channel of data, and they can be replayed in the Prairie View software or loaded into third party programs. Before Conversion After Conversion Green Signal Red Signal Photoconversion of EoS in fibroblasts with Spiral Activation. Setup shown for photomanipulation of arbitrary areas. Red masks show the areas that will be scanned by the photomanipulation scanner. Prairie View Image Window showing location of targets for point photoactivation. Each point represents a diffraction-limited spot that will be illuminated by a user-selected laser at a defined power and time. Setup shown for running spiral activation on three areas. Circles show the areas that will be scanned, with blue lines showing the trajectory of the galvanometers between spirals.

8 Bruker Nano Surfaces Division is continually improving its products and reserves the right to change specifications without notice Bruker Corporation. All rights reserved. Opterra and Prairie View are trademarks of Bruker Corporation. All other trademarks are the property of their respective companies. B2000, Rev. B0 Opterra Specifications Scanhead Scanning Method Apertures Scan Speeds Filters and Dichroics Triggering Image Collection CCD Camera Optical Inputs Visible Laser Platform and Automation Microscope Specimen Stage Z Focus Combination galvanometer and piezoelectric crystal scanning Fully motorized aperture plate with 7 software-selectable apertures; 3 pinhole settings (30, 45, and 60 µm); 4 slit settings (22, 35, 50, and 70 µm) Scanner or camera master timed imaging; Up to 50 fps in pinhole mode; Up to 1000 fps in slit imaging mode (with appropriate camera) Motorized 6-postion emission filter wheel; Custom dichroics and polychroics for multiple excitation/emission wavelengths Frame trigger out and frame trigger in; 4 auxillary device triggers Camera port with adapter for c-mount cameras; Driver support for Photometrics and Q-Imaging cameras; Evolve Delta EMCCD or Rolera EM-C2 EMCCD camera standard; Software support for gain settings, integration time, binning, and ROIs; Software support for DV2 and QV2 emission splitting systems Helios launch with up to 5 diode laser lines; Lasers analog modulated with digital blanking; Launch designed to minimize thermal drift over extended time periods; Fiber optic input interface to Opterra scanner Nikon TI-E inverted and Nikon FN-1 upright; Software control of objective turret, filter cube turret and Z-motor for Nikon TI-E ASI XY stage with piezo focus insert (300 µm range) for TI-E; Bruker Slim Stage for FN-1 Internal focus motor and stage Z-piezo for TI-E; Bruker Z-focus motor and Bruker piezo-driven objective Z-focus for NI-E Photoactivation Scanner Optical Inputs Analog Inputs/Outputs Outputs Inputs Single-pair galvanometer (3 mm) scanner for high-speed photoactivation, photobleaching, and photoablation Helios launch with up to 5 diode laser lines; Ultra-fast IR laser input via table-mounted optics (includes Pockels cell) Up to 8 analog outputs for control of analog or TTL-controllable devices Up to 8 analog inputs for recording -10V to +10V signals Software Prairie View Imaging Software O/S Computer Complete image acquisition and photoactivation control software package Windows 7, 64-bit processor Intel processor Quad Core; Two 128GB SSDs (mirrored) for O/S and application software; 512GB SSD for data collection; 2TB platter drive data archival; 32GB RAM Bruker Nano Surfaces Division Middleton, WI USA Phone productinfo@bruker.com

Opterra II Multipoint Scanning Confocal Microscope. Innovation with Integrity

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