User manual LEICA TCS STED TECHNICAL DOCUMENTATION

[. . . ] Leica TCS STED The Fast Track to Superresolution Technical Documentation 1 8 2 5 9 3 10 6 4 7 11 1 2 3 4 5 6 7 8 9 10 11 Inverted research microscope Leica DMI6000 CS Scan head Laser and power supply Computer table Air damped optical table Control panel Supply control STED module Housing for 592 nm STED depletion laser APD detection unit (optional, recommended) Microscope control STED Module Mechanics STED Laser: Optomechanics excitation: depletion: ultra stable and compact device, firmly fixed to scanner internal Argon gas lasers (continuous wave), variable excitation wavelength 592 nm visible fiber laser (continuous wave) used imaging port UV-port (no UV available) modulation of depletion PSF, automated beam adjustment for perfect alignment of excitation and depletion laser, average duration: < 1 min. , alignment inside the scanhead, no illumination by lasers of the sample during alignment 2 Specifications* Microscope Z-drive inverted SuperZ galvanometer stage motorfocus (stand) Leica DMI6000 CS Trino/Bino 1500 m travel range/3 nm stepsize travel range depending on mechanics of microscope/15 nm step size diode 18 mW 442 nm Ar 100 mW 458, 476, 488, 496, 514 nm HeNe 1 mW 543 nm HeNe 2 mW 594 nm HeNe 10 mW 633 nm DPSS 10 mW 561 nm VFL depletion 1. 5 W 3 (STED, VIS, IR) up to 8 Acousto Optical Beam Splitter (AOBS®) or dichroic mirrors 400. . . 800 nm alignment stable single pinhole motorized by software, automatic mode available optically correct scanning at low inertia conventional and resonant scanner in one system (optional) active Lasers VIS STED depletion Optics number of laser ports for imaging number of lasers for imaging excitation ­ emission splitting detection range pinhole pinhole diameter control Scanner scanning concept switch conventional ­ resonant scanner vibration insulation Conventional (C) max line frequency min line frequency scan speed granulation max frame rate 512 x 512 max frame rate 512 x 16 beam park max frame resolution scan zoom pannning field rotation field diameter max line frequency min line frequency scan speed granulation max frame rate 512 x 512 max frame rate 512 x 16 2800 Hz 1 Hz 1400 5 Hz 50 Hz yes [no] 8192 x 8192 pixel 1. 0 . . . [. . . ] 64x] yes 200° optical [no] 15 mm [75 m] Resonant (R) * Text in brackets: deviant specifications when system is used in STED mode 3 Specifications* Scan modes t xt x xyt xy xz xz xyz xyz xyt xzt xyzt xytz emission separation max spectral confocal channels tunability of emission bands tuning steps of emission bands sensors yes (conventional scanner) yes yes yes yes yes yes yes yes yes yes yes yes highly sensitive prism spectral detector 5 yes 1 nm high sensitivity low noise PMT: R 9624 ultra sensitive Avalanche Photo Diodes (APD): 2 [1] (optional) high speed CCD Camera: Leica DFC360 FX, fully integrated (optional) 12 or 8 bit per channel 16 bit imaging 40 MHz optional, allowing BF, DIC, Ph etc 1 ­ 2 (optional) 1 ­ 2 (optional) digitally at high performance (FPGA, field programmable gate arrays) yes high performance PC workstation programmable control panel with LCD function & value display intuitive and guiding user interface context sensitive online help system multi-dimensional data acquisition region of interest (ROI) scan excitation line/frame sequential scan emission spectrum recording quantification tools multi-color restoration, spectral unmixing general time lapse experiment control tile scanning (mosaic scan) fully integrated STED control Interactive data recording also allowing job-sequencing and online evaluation combines Mark & Find with sophisticated 3D recordings, Live Data Mode etc. maximum and other projections, simulated fluorescence process, rotation animations, stereo pairs, red-green anaglyphes, height color coded extended depth of focus images etc. histogram based colocalization and area measurements deconvolution option for widefield images, confocal images FRAP wizard, FRAPxt wizard, FLIP wizard, FRET SE wizard, FRET AB wizard etc. Detection digitization max grey resolution read out frequency transmitted light detector non descanned transmitted light channels non descanned reflected light channels Electronics scanner control trigger out functions computer general Software Software options Live Data Mode Advanced Mark & Find 3D visualisation Colocalization Deconvolution MicroLab * Text in brackets: deviant specifications when system is used in STED mode 4 System Components 20 18 17 16 15 APD 2 19 APD 1 13 10 10 10 6 5 9 7 21 3 10 10 14 11 4 12 IR- La ser VIS- La s er 2 8 LEICA ST ED CW 1 1 2 3 4 5 6 7 8 9 10 11 12 Housing for 592 nm depletion laser, AOTF, electronics Fiber Helical vortex phase filter Incoupling STED dichroic Tandem Scanner Field rotation optics Quarter wave plate Transmitted light detector Reflected light detectors Photomultipliers Multi-function port IR EOM 13 14 15 16 17 18 19 20 21 Visible range AOTF AOBS Confocal detection pinhole Filter- and polarizer wheel incl. notch filters X1 emission port APD filter cubes Avalanche photodetectors Spectrophotometer prism STED objective lens visible and ultraviolet radiation: infrared radiation: 5 Installation Requirements Including optical bench & IR laser system Heat load max Electric supply 500 kg 3. 5 W Minimum three separate phases: VIS lasers: 100. . . 240 V AC ± 10% 2 x 1600 VA, 50/60 Hz (Power Input 1 + 2) 592 nm depletion laser: 100. . . 240*-10, 2. . . 1A Environment Temperature: 23°C ± 1°C Humidity: approx. 40 ­ 50% 1013 hPa Max floor vibration amplitudes: ­ frequency range 5. . . 12. 5 Hz: 30 m/sec rms ­ frequency range > 30 Hz: 12. 5 m/sec rms Internet access for advanced remote diagnostics Room must comply with country-specific regulations for laser class IV Room darkening recommended Protect from dust (fiber length: 3 m) to minimize vibrations, noise and heat approx. 1400 mm 4' 7'' Distance from wall 600 mm 2' Width approx. [. . . ] 1500 mm 4' 11'' www. leica-microsystems. com LEICA and the Leica Logo are registered trademarks of Leica Microsystems IR GmbH. [. . . ]