R DUAL CCD CAMERA Dual wavelength imaging Both images captured simultaneously Wide field of view Plus superior image alignment Dual focal plane imaging By adjustable CCD device position Max. QE over 70 % High sensitivity from VIS to NIR A One-of-a-kind Solution for Dual Wavelength Imaging With the ORCA-D2, you can now capture dual wavelength images simultaneously, at wavelength ranges of your choice, and with a wide field of view − a unique combination that simplifies the challenges of conventional FRET and ratio imaging techniques. The ORCA-D2 contains two CCD devices. Interchangeable optical blocks are used to set the wavelength ranges for each CCD. During image capture, the camera automatically corrects focus, alignment and color shifting to produce high-quality images. Other benefits of the ORCA-D2 include high sensitivity and low noise, courtesy of Hamamatsu's advanced CCD cooling technology; wide dynamic range; standard IEEE1394b interface; and Hamamatsu's proprietary vacuum-sealed chamber technology, which promotes long-term maintenance-free operation with even the most demanding applications. APPLICATIONS Ratio imaging Single and dual wavelength fluorescence microscopy, FRET Blue to NIR fluorescence applications Colocalization and FISH applications Dual wavelength TIRF microscopy, real-time confocal microscopy Combined transmission and fluorescence imaging Multi-focal point imaging microscopy FEATURES Simultaneous capture of wide-field dual wavelength images Dual wavelength imaging Selectable wavelengths The unique camera design makes it possible to capture simultaneous dual wavelength images with a full field of view, even when the intensities differ significantly. Such images are difficult to obtain with conventional FRET or ratio imaging techniques. By changing optical blocks, you can select your wavelengths of interest. Hamamatsu offers three types of optical blocks for different wavelength ranges. CCD device 2 Optical block Lens mount CCD device 1 Internal structure of camera head Comparison with conventional methods ORCA-D2 W-view optics Simultaneous capture at dual wavelengths Simultaneous capture at dual wavelengths Selectable wavelengths CFP image DM M Steps required to correct color or image CCD camera DM Wide field of view ORCA-D2 camera head Selectable wavelengths W-View optics A8509 Auto-correction of color or image shifting M Reduced field of view YFP image Aberration correction of color mismatch between wavelengths YFP image CFP image 3CCD color camera Filter wheel Simultaneous capture at dual wavelengths R-channel 3CCD color camera G-channel No simultaneous capture at dual wavelengths No selection of wavelengths Selectable wavelengths CCD camera No correction of color or image shifting Wide field of view No correction of color or image shifting Wide field of view System for changing wavelength B-channel CFP image YFP image CFP image YFP image Application example 1 Ca2+ measurement using YellowCameleon 3.6 (optical block: DM 510 nm) The following images are an example of ratio imaging. Separated CFP and YFP (FRET) are measured with dual CCD devices. This sequence observed the YellowCameleon 3.6 (Ca2+ sensor based on CFP-YFP FRET) expressed Ins-1 cell response with a depolarizing stimulus. CFP-channel (CCD1) YFP-channel (CCD2) Samples courtesy of Hideo Mogami, Ph.D. Dept. of Physiology, Hamamatsu University School of Medicine Dept. of Environmental Biology, Okazaki Inst. for Integrative Bioscience Temporal change of brightness Sample: Ins-1 cell (insulin-producing cell) ORCA-D2 optical block: A11400-03 (DM 510 nm, Em1 483 nm/32 nm, Em2 542 nm/27 nm) Microscope: Olympus IX71 Objective lens: Olympus LUCPlanFLN 60x, NA 0.70 Easy setup Auto-correction of focus and alignment Focus adjustment During dual wavelength image acquisition, the camera will automatically correct image alignment, color mismatch, etc. Hardware and special software work together to adjust focus* and alignment to your experimental setup. Calibration results are saved in the software, eliminating the need to readjust for the same setup. Images with different focal plane can be acquired at the same time through the use of the camera's special software and adjustable CCD device position. Pollen with a 10 μm diameter The following images are examples of using different focal plane imaging at the same time. CCD1 *Please note that auto-correction may not be possible in some experimental setups. Example of correction of focus and alignment ±0 μm ±0 μm ±0 μm ±0 μm -4 μm CCD2 Before After Sample: ORCA-D2 calibration plate ORCA-D2 optical block: A11400-02 +4 μm High-sensitivity imaging from visible to near-infrared Spectral response Dual light mode Quantum efficiency (%) Two modes (high light, low light) are available for different situations. Low light mode provides high sensitivity over a wide spectral range from visible to near-infrared. Max. quantum efficiency over 70 % The ORCA-D2 offers high sensitivity from visible to near-infrared with its ER-150 interline CCD. In low light mode, sensitivity at 700 nm is nearly twice that of the ICX285* device used in most other scientific CCD cameras. (This sample is typical of the CCD characteristics, not guaranteed.) 80 Low light mode 70 High light mode ICX285 60 50 40 30 20 10 0 300 400 500 600 700 800 900 1000 1100 Wavelength (nm) * ICX285 is a conventional high-sensitivity CCD device which is used for scientific instrumentation. ICX285* image Low light mode image (ORCA-D2) Brightness profile of area marked with white horizontal line Application example 2 Brightness profile of area marked with white horizontal line Membrane Potential Measurment using Mermaid (optical block : DM 550 nm) Spontaneous oscillation of membrane potential in mouse cardiomyocyte expressing Mermaid. Intensity changes of dual fluorescence from Mermaid were measured simultaneously at 50 frames/s, using ORCA-D2. Green and Red channel of camera were used for mUKG and mKO respectively. 0.75 1000 Images courtesy of Hidekazu Tsutsui,Ph.D. Laboratory of Integrative Physiology, Department of Physiology, Graduate School of Medicine, Osaka University Ratio 900 0.70 0.65 700 600 0.60 Green-channel 500 0.55 400 Red-channel 0.50 300 0 1 2 3 Sec Ratio RIKEN Brain Science Institute Advanced Technology Development Core Laboratory for Cell Function Dynamics Sample: Cardiomyocyte from mermaid tg-mouse ORCA-D2 optical block: A11400-04(DM550 nm,Em1 520 nm/35 nm, Em2 593 nm/40 nm) Microscope: Olympus IX71 Objective lens: UPlan FL 60x/1.25 oil Intensity 800 SYSTEM CONFIGURATION Optical block A11400-01 ORCA-D2 Camera head Microscope Digital camera interface kit Camera control unit Computer 2/3-inch bayonet mount TV camera adapter Standard Option HCImage 2/3-inch bayonet mount lens Commercially available software For details on microscope adapters and other optical blocks, please consult your local Hamamatsu sales representative. HCImage software provides standard image measurements functions. Upgrades to more feature-rich versions are available. SPECIFICATIONS Trigger output Programmable timing output, Trigger ready output Lens mount Interface External control 2/3-inch bayonet mount (flange back focus 48 mm) IEEE1394b-2002 IIDC 1394-Based Digital Camera Specification Ver.1.32 AC 100 V to AC 240 V, 50 Hz / 60 Hz Approx. 70 V • A Power requirements Power consumption Ambient storage temperature Ambient operating temperature Ambient storage / operating humidity Applications Em1 Em2 None Half 510 nm 550 nm 630 nm 570 nm 0% 50 % 483 nm / 32 nm 520 nm / 35 nm 593 nm / 40 nm 520 nm / 35 nm 100 % 50 % 542 nm / 27 nm 593 nm / 40 nm 692 nm / 40 nm 641 nm / 75 nm 1CCD camera Beam splitter CFP / YFP FITC / TRITC Cy3 / Cy5 GFP/mCherry A11400-01 is included with camera. Other optical blocks are optional. DIMENSIONAL DIMENSIONAL OUTLINES OUTLINES (Unit: mm) Camera head (Approx. 3.0 kg) 107 ±1 225.5 ±3 Camera control unit (Approx. 3.5 kg) - 10 ˚C to + 50 ˚C + 10 ˚C to + 35 ˚C 70 % max. (no condensation) 74 ±1 Dynamic range typ. Cooling method Cooling temperature Dark current A/D converter Exposure time Sub-array Contrast intensification External trigger mode A11400-01 A11400-02 A11400-03 A11400-04 A11400-05 A11400-08 Optical block Dichroic 262 ±2 6.9 ±1 Readout noise (r.m.s.) typ. 1×1 Full well capacity typ. 2×2 Binning Type number 8 ±1 2✕2 4✕4 8✕8 6.45 μm (H) × 6.45 μm (V) 8.26 mm (H) × 6.19 mm (V) 20.00 MHz/pixel 11.2 frames/s 20.2 frames/s 33.6 frames/s 50.5 frames/s 8 electrons 18000 electrons 36000 electrons 2250 : 1 typ. (1 × 1) Peltier device + Forced-air cooled - 10 ˚C (Ambient temperature: + 20 ˚C) 0.01 electrons/pixel/s (- 10 ˚C) 12 bit 117 μs to 60 s Yes Analog gain (10 times max.), offset Edge trigger, Level trigger, Synchronous readout trigger, Start trigger Optical blocks 105 ±1 Binning C11254-10B (ORCA-D2) Hermetic vacuum-sealed head ER-150 progressive scan interline CCD 1280 (H) × 960 (V) × 2CCD chip 2.8 ±1 Type number Camera head type Imaging device Effective number of pixels Cell size Effective area Pixel clock rate Readout speed 1×1 333 ±2 Calculated from the ratio of the full well capacity and the readout noise ORCA is registered trademark of Hamamatsu Photonics K.K. (France, Germany, Japan, U.K., U.S.A.) HCImage is registered trademark of Hamamatsu Photonics K.K. (Australia, China, EU, Japan, Norway, Singapore, Switzerland, U.S.A.) Product and software package names noted in this documentation are trademarks or registered trademarks of their respective manufacturers. ● ● Subject to local technical requirements and regulations, availability of products included in this promotional material may vary. Please consult your local sales representative. Information furnished by HAMAMATSU is believed to be reliable. However, no responsibility is assumed for possible inaccuracies or omissions. Specifications and external appearance are subject to change without notice. © 2010 Hamamatsu Photonics K.K. HAMAMATSU PHOTONICS K.K. www.hamamatsu.com HAMAMATSU PHOTONICS K.K., Systems Division 812 Joko-cho, Higashi-ku, Hamamatsu City, 431-3196, Japan, Telephone: (81)53-431-0124, Fax: (81)53-435-1574, E-mail: [email protected] U.S.A.: Hamamatsu Corporation: 360 Foothill Road, P. O. Box 6910, Bridgewater. 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No. SCAS0067E07 NOV/2010 HPK