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OPERATING INSTRUCTIONS

Lattice Lightsheet 7

System Operation

ZEISS

04/2021 V_01

000000-2324-960

Auto Immersion

Time interval setting

Immersion supply mode

Fig. 55

Auto Immersion tab

Lattice Lightsheet / Aberration Control

Movement of sheet along optical axis of

illumination objective lens axis

Movement of sheet perpendicular to illumination

objective lens axis

Compensation of aberrations in detection

objective lens

Tilt compensation along X direction

Tilt compensation along Y direction

Fig. 56

Lightsheet/Aberration Control tab

Time series

Duration in cycles or period

Intervall between cycles

Fig. 57

Time Series tab

Summary of Contents for Lattice Lightsheet 7

Page 1: ...Lattice Lightsheet 7 Safety Information Setup Requirements and Instructions April 2021 ...

Page 2: ...t to alterations The manual is not covered by an update service Unless explicitly authorized dissemination or duplication of this document and commercial exploitation or communication of its contents are not permitted Violations will entail an obligation to pay compensation All rights reserved in the event of granting of patents or registration as a utility patent Issued by Carl Zeiss Microscopy G...

Page 3: ...the Customer 22 1 12 4 Maintaining the Liquid Cooling System LCS BU of the Camera 23 1 12 5 Count on Service in the True Sense of the Word 23 1 13 User Interface Mounting and Dismounting Components 24 1 13 1 Changing the Meniscus Lens Relay Optics 25 1 13 2 Removing the Immersion Supply System 28 1 13 2 1 Removing the Immersion Container 28 1 13 2 2 Immersion Priming 29 1 13 3 Mounting and Dismoun...

Page 4: ...64 2 10 System Overview 65 3 OPERATING INSTRUCTIONS 66 3 1 Switching on and off the Lattice Lightsheet 7 System 67 3 1 1 Starting and Stopping the ZEN Software 68 3 1 2 Presetting the System 70 3 1 2 1 Switching to Operation Mode Obligatory 70 3 1 2 2 Selecting the Carrier Obligatory 70 3 1 2 3 Calibration of the Stage 71 3 1 2 4 Priming immersions 71 3 1 2 5 Environmental Control Optional 72 3 2 ...

Page 5: ...nded for volumetric imaging of biological samples at high speed whereby the specimen is illuminated by a structured sheet of light Installation and commissioning of the Lattice Lightsheet 7 systems must be performed by authorized ZEISS service staff The system should not be used prior to instruction by a ZEISS representative The manufacturer will not assume liability for any malfunction or damage ...

Page 6: ...also observe all and any relevant statutory accident prevention regulations The user is referred to the safety data sheet provided together with the manual 1 2 Explanation of Symbols CAUTION Faults and hazards that might arise during operation which might cause damage to the unit or injury to the user Observe the operating manual CAUTION Laser radiation hazards possible when operating the system D...

Page 7: ...g to DIN EN 61326 1 1 Noise emission according to CISPR 11 DIN EN 55011 2 Noise immunity according to table 2 industrial sector In the rare case of a voltage surge on the wired data lines such as USB LVDS Sub LVDS e g from indirect lightning stroke a momentary interruption of the acquisition system s functionality is possible The imaging can stop or a short interruption of the camera image is poss...

Page 8: ...irective 2014 35 EU EMC directive 2014 30 EU KN 61000 6 2 KN 11 EMC User notice for Korea only 사용자안내문 기 종 별 사 용 자 안 내 문 A급 기기 업무용 방송통신기자재 이 기기는 업무용 A급 전자파적합기기로서 판매자 또는 사용자는이 점을 주의하시기 바라며 가정외의 지역에서 사용하는 것을 목적으로 합니다 The company works according to a certified Environment Management System according to ISO 14001 The product was developed tested and produced in accordance with the valid regulations and...

Page 9: ...or control computer system PC sCMOS camera stage motion controller camera liquid cooling system The following components must be inserted in the 3 point power strip multipoint connector 2 or incubation components power strip see Fig 1 gas mixer controller temp module controller The folowing components must be connected to the 3 point power strip multipoint connector 3 or storage components power s...

Page 10: ...the Lattice Lightsheet 7 are displayed in Fig 2 and Fig 3 respectively Line voltage 230 V AC 10 120 V AC 10 Line frequency 50 60 Hz 50 60 Hz Lattice Lightsheet 7 system incl lasers Max current 1 phase at 4 5 A 1 phase at 9 A Power Power consumption 900 VA continuous operation maximum 280 VA standby operation 0 025 VA off mode 900 VA continuous operation maximum 260 VA standby operation 0 011 VA of...

Page 11: ...SAFETY INFORMATION Lattice Lightsheet 7 Power Requirements ZEISS 04 2021 V_01 000000 2324 960 11 Fig 1 Schematic drawing of power connections ...

Page 12: ... LVDS 2 OVERVIEW CAMERA 11 CAMERA ADAPTER 3 SLM 12 GAS CAN Bus 4 Slit recess for loop through of tubing 13 GAS Supply line 5 TRIGGER STAGE 14 TEMP Supply line 6 TRIGGER CAM 1 15 TEMP CAN Bus 7 TRIGGER OUT 16 MICO F 1 8 TRIGGER CAM 2 17 POWER CONNECTOR 9 LVDS Fig 2 Connectors and fuses of the Lattice Lightsheet 7 ...

Page 13: ...h a protective contact The protective effect must not be overridden by an extension cable without a protective conductor Detachable power cords should not be replaced with inadequately assessed power cords Only the specified power cords may be used Only the connection cables supplied with the device may be used The use of commercially available cables for use with the LLS7 is prohibited A safe dis...

Page 14: ...g acclimation For operation the system must be placed in a closed room 1 Operation specified performance without incubation T 22 C 0 5 C without interruption 24 h a day independently whether system is operated or switched off It has to be ensured that the air flow of the air conditioning is not directed at the system 2 Operation reduced performance without incubation T 15 C to 30 C any conditions ...

Page 15: ...e power plug must be disconnected Do not set up the unit in the proximity of heat sources such as radiators or direct sunlight To avoid heat build ups the ventilation louvers or slots on the microscope system must not be covered up Do not position the electronics parts in a closed container The system must not be set up in areas with potential danger by explosives The unit must be connected to a p...

Page 16: ...after conversion of the Lattice Lightsheet 7 system authorized specialized staff must carefully check that it is in a proper condition and particularly that covers protecting against laser radiation are provided By establishing a corresponding workplace environment please ensure that the formation of electrostatic charges is avoided To avoid vibrations during operation the Lattice Lightsheet 7 sys...

Page 17: ... attached on the Lattice Lightsheet 7 system Fig 4 and Fig 5 must be observed Check whether all of the labels shown below are provided on your instrument and contact ZEISS Germany or one of the service agencies if you should discover that any of the labels should be missing You will receive a free replacement Fig 4 Warning and information labels of a Lattice Lightsheet 7 system ...

Page 18: ...ISS Warning and Information Labels Lattice Lightsheet 7 18 000000 2324 960 04 2021 V_01 1 Laser ON LED 2 US version only Fig 5 Warning and information labels of a Lattice Lightsheet 7 system front view top rear view bottom ...

Page 19: ...he laser The laser module is equipped with a key switch The laser module is equipped with an electric interlock at the rear of the laser module see Fig 4 This can be used to connect remote interlock Always keep the keys for the laser module in places which are not accessible to persons who are not authorized to operate the laser A white LED Fig 5 1 on the front of the component rack lights up once...

Page 20: ...of the disc Do not apply paper labels or write on any part of the disc data side or label side Wipe dust or fingerprints on the disc with a soft cloth from the center to the edge Do not use benzene thinner record cleaner or antistatic clothes This may damage the disc Do not expose the disc to direct sunlight or high temperatures Backup your data on a regular basis Do not install any other software...

Page 21: ...copy Please contact your ZEISS service representative to check the safety of the device Please keep the service logs of your device 1 12 1 Notes on Maintenance and Care by the Manufacturer Service To ensure proper functioning of the system in the long term we recommend entering into a service agreement with your closest ZEISS representative You are well advised to arrange a service agreement with ...

Page 22: ...der push it in and lock it by turning 90 anti clockwise 1 12 3 Notes on Maintenance by the Customer Care operations that may be carried out by operating staff are limited to cleaning painted and glass surfaces Before cleaning the instrument make sure the main power supply is disconnected Cleaning painted surfaces To do this use a clean cloth that has been moistened in a mixture of water and some d...

Page 23: ...highly qualified ZEISS specialists who will support you long beyond the purchase of your system Our aim is to enable you to experience those special moments that inspire your work Repair Maintain Optimize Attain maximum uptime with your microscope A ZEISS Protect Service Agreement lets you budget for operating costs all the while reducing costly downtime and achieving the best results through the ...

Page 24: ...er The following interfaces are no user interfaces when used with a Lattice Lightsheet 7 pco edge sCMOS cameras and their adapters Additional port and ist cover Other interfaces which are not described here are service interfaces and are only to be operated by authorized ZEISS service personnel The following devices can be mounted and dismounted by the user or are accessible by the user Relay Opti...

Page 25: ...60 25 1 13 1 Changing the Meniscus Lens Relay Optics The meniscus lens relay optics is mounted on a metal support Meniscus lens plus support must be exchanged as a whole 1 Move the transmission arm to the up position Fig 8 1 2 Open the front door Fig 8 2 1 Transmission arm 2 Front door Fig 8 Transmission arm and front door ...

Page 26: ... meniscus lens holder Fig 9 5 6 Insert meniscus lens holder Fig 10 5 into the hole Fig 10 6 7 Grasp the meniscus lens in the recessed grips Fig 10 8 1 and 8 2 lift it out and place it in the holder Fig 10 5 3 Incubator lid 4 Sample mounting frame 5 Meniscus lens holder Fig 9 Incubator lid sample mounting frame and meniscus lens holder 5 Meniscus lens holder 6 Hole 7 Meniscus lens relay optics 8 1 ...

Page 27: ...1 Connect the Luer Lock couplings 12 Insert the meniscus lens into the incubator Fig 12 13 Turn the meniscus lens until the three point support is in place 14 The orientation groove Fig 12 7 1 must point to the left 15 Reinsert the sample mounting frame 16 Reinsert the incubator lid 17 Lower the transmission arm 5 Meniscus lens holder 7 Meniscus lens 9 Luer Lock coupling 10 Luer Lock coupling Fig ...

Page 28: ...aching into the liquid 4 Close the front door We recommend always closing the front door to avoid dust entering the system The front door is not connected to the laser safe system so it can be operated with the door open As the container does not contain a lid and can receive light when the door is open there is a risk of bacterial or algae contamination which might lead eventually to a clogging o...

Page 29: ...er should be used as immersion Do not use oil or glycerol based immersion media as these more viscous solution will clog the tubing system Fig 14 4 1 Water immersion drop 6 3 way valve arrow indicates flow direction 2 Relay optics meniscus lens 7 Pump arrow indicates flow direction 3 Relay optics meniscus lens holder 8 Immersion container water 4 Hydraulic tubing 9 Luer Lock stopcock 5 Luer Lock c...

Page 30: ...t the syringe empty it reconnect it and refill it 3 To remove any air bubbles from the tubing Fig 14 4 press water out by slowly pushing the syringe piston Note that due to the valve Fig 14 6 and pump Fig 14 7 water can only flow to the relay optics Fig 14 2 via the Luer Lock coupling Fig 14 5 Observe water level rising at the meniscus lens and stop pressing the piston before water spills from the...

Page 31: ...d and put it vertically on the stage Make sure to match the electronic contacts Fig 15 2 between lid and stage and that the recess of the lid fits into elevated part of the stage The lid will then snap onto the stage via its magnets Fig 15 3 3 Lower the transmission arm The transmission arm is connected to the laser safety system In order that the system can operate the transmission arm must be lo...

Page 32: ...st the incubator spring using the two handling screes and lift the frame up from the other side 3 Remove the frame 4 Insert the incubator lid Mounting 1 Remove the incubator lid see section 1 13 3 2 Take a suitable frame Fig 17 1 using the two handling screws and press with one side against the spring of the incubator stage Fig 17 2 3 Make sure the mounting frame is tightly inserted 4 Insert the i...

Page 33: ...Fig 18 8 with a hexagon screwdriver SW8 4 Fill the cooling unit with innovate Protect coolant if the liquid loss was due to leakage until the liquid is visible on the thread of the filler neck If no leakage is obvious and the liquid los was due to evaporation fill with distilled water 5 Open the vent valve Fig 18 9 left stop 6 Close the filling refilling port Fig 18 8 7 Connect the system to the p...

Page 34: ...e following instruction manuals ibidi Heating System Stage Top for Lattice Lightsheet 7 ibidi Gas Mixer System for Lattice Lightsheet 7 The control units or modules should be placed on a level surface next to the system and can be stacked on top of each other Most conveniently the Temperature Controller is placed on top of the Gas Mixer and the Humidifier is placed on their side 1 13 6 1 Heating s...

Page 35: ...ect the USB Fig 19 8 to an USB port of the system computer Connect the CAN bus 2 Fig 19 7 to the Gas CAN Bus 1 plug Fig 2 11 of the Gas Mixer On the CAN Bus 1 socket insert a CAN Bus blind plug 1 Power control LED for power channel 6 CAN Bus 2 Can Bus connection to Gas Mixer 2 Lid control LED for Lid channel 7 CAN Bus 1 Can Bus blind plug 3 Tray control LED for tray insert channel 8 USB for connec...

Page 36: ...lly via its electronic contacts Fig 20 5 to the contacts provided on the heating chamber frame Fig 21 6 The magnetic cover Fig 20 6 can be used to close the two smaller recesses holes used for perfusion tubing or cables Heated Base and Tray 1 Electric contact for lid 7 Chamber frame 2 Docking block 8 Mesh for relay optic frame 3 Electric socket 9 Heated plate 4 Electric cable plate and frame 10 El...

Page 37: ... amount of O2 in the gas mixture to create hypoxia via the N2 supply Gas Mixer The Gas Mixer Fig 22 is the gas mixing and control unit of the ibidi Gas Incubation System In addition to the gas mixture the humidity within the incubation chamber is controlled by a feedback loop using a humidity sensor located in the incubation chamber The LEDs on the front of the gas mixer Fig Fig 22 indicate the st...

Page 38: ...line Control the remaining pressure on the gas line s pressure reducing valve Then remove the tubing 1 Power Control LED for power channel 6 CAN Bus 2 Can bus connection to Lattice Lightsheet 7 Gas Can Bus 2 N2 Control LED for N2 supply 7 On Off On off switch 3 Air O2 Control LED for air O2 supply 8 Output 1 Connects to blue tubing with white label of the heated tubing package 4 CO2 Control LED fo...

Page 39: ...ify the mixed gas Part of the mixed gas output is guided through the column The water inside the column is warmed up for humidity uptake The Humidifying Column base heats up to 50 C 122 F when in operation which is indicated by the warning label attached to the base of the column Fig 23 6 The base will be heated as long as the gas mixer is switched on independent on any software settings It is rec...

Page 40: ... Re Filling the Humidifying Column Before the first use the Humidifying Column must be filled with 130 ml of ultra pure water During operation the column should always be refilled before the water level falls below the minimum mark For re filling proceed as follows 1 Remove the blue stopper Fig 23 8 from the top of the column 2 Use a squeeze bottle Fig 23 9 to fill the column through the filling p...

Page 41: ... Humidifying Column 6 Heated wire split off from electrical cable 2 Gas tubing to the Heated Chamber 7 Insulating jacket 3 Merging point for heating wire into the tubing 8 Mixed gas output white 4 Gas output from Humidifying Column 9 Mixed gas output yellow 5 Gas input to Humidifying Column Fig 24 Heated Tubing package Arrows indicate flow direction of the gas mixture Electrical connection 1 Cable...

Page 42: ...he Heated Tubing The heating wire inside the tubing may be damaged and become non functional Note that the tubing from the Gas Mixer yellow marked output 2 combines with the tubing from the Humidifying Column red marked output to the tubing connecting to the Heated Chamber 1 13 6 3 Operation The Gas Incubation System is combined with the Heating System providing a fully controlled stage top chambe...

Page 43: ...25 5b It is recommended to insert an air filter Fig 25 6b between the air supply and the Gas Mixer 5 Connect the Supply Tubing N2 Fig 25 4c to the Input 1bar N2 port Fig 25 7 and connect other end to an appropriate CO2 supply source e g lab line or gas bottle Fig 25 5c It is recommended to insert an air filter Fig 25 6c between the air supply and the Gas Mixer If N2 is not needed close the port wi...

Page 44: ...ctric and gas circuits of the heating and gas incubation systems 1 Lattice Lightsheet 7 2 Heated chamber 2a Gas connection tube 2b Electrical connection cable 3 Humidifying column 4 Heated Tubing Package 4a Tubing white plugged end to LLS 7 4b Heating cable connecting gas mixer to humidity column 4c Tubing yellow marked end gas mixer 4d Tubing white end of gas mixer blue end of humidifying column ...

Page 45: ...b the measurement After connecting all cables and tubing and after filling the Humidifying Column with ultra pure water the system is ready to start 1 Set the gas regulation valve to the recommended pressure of 1 bar 14 5 psi 2 Switch on the Gas Mixer and wait until bubbling is observed 3 Set the parameters in the ZEN Blue Lattice Lightsheet incubation tab Fig Fig 26 4 Give the system time to equi...

Page 46: ...tion In the ZEN software the following warning symbols are displayed For Temperature Actual Temperature corresponds to set temperature within a range within of 0 1 C actual temperature is below set temperature actual temperature is above set temperature Temperature setting not active unchecked box For Atmosphere Actual gas humidity concentration corresponds to set concentration within a range with...

Page 47: ...ig 28 4 4 Close the lid and ensure that the thermoelement wire is fed through one of the recesses in the stage lid Fig 16 2 5 Set Lid temperature to 40 C and both Tray and Base to 38 C as start values to reach a sample temperature of 3 C 1 Thermoelement sensor 2 Ibidi 35 mm dish 3 Ibidi 8 well chamber 4 Drill holes in the lids Fig 28 Temperature adjustment set 6 Adapt tray and base temperatures as...

Page 48: ...7 1 Stage Motion Controller The stage of the Lattice Lightsheet 7 is controlled by the Motion controller Fig 30 It can move on 5 axes X Y Z X tilt Y tilt These axes are regulated via 5 channels For each channel there are three LEDs at the front of the emotion controller Power LED on channel powered Enabled LED on channel active Fault LED on channel error ...

Page 49: ...l 1 2 USB port 10 Input Output channel 2 3 LED display channel 1 11 On Off switch 4 LED display channel 2 12 Power in 5 LED display channel 3 13 Ethernet LAN connector 6 LED display channel 4 14 Encoder 5 Z1 Axis 5 7 LED display channel 5 15 Encoder 4 Z2 Axis 4 8 LED display power 16 Encoder 3 Z3 Axis 3 17 Encoder 2 Y Axis 2 18 Encoder 1 X Axis 1 19 Motor 5 Z1 Axis 5 20 Motor 4 Z2 Axis 4 21 Motor ...

Page 50: ... 7 Y drive 8 Not used Fig 31 Ergo pointer front left and rear right view 1 13 7 3 Stage The stage is calibrated with the stage top incubator lid mounted In case the lid is not mounted stage travel can become inaccurate at extreme travel ranges For more information on calibrating the stage read the instruction manual of the stage In case the lid is not mounted light can escape to the right side of ...

Page 51: ...n The supplied safety data sheet with notes on hazards and safety measures must be observed when handling the cooling liquid The port for the camera is no user interface and for service only Never remove the camera or the blind cap that might cover the port if the camera was detached 1 Detector port 1 6 Power socket 2 Rotation guard plate with laser protection label 7 Trigger out BNC connecter 3 T...

Page 52: ... separate safety data sheet for the liquid cooling system The LCS is connected to the system s 6 point power strip The cooling hoses should be as short as possible The connection creates a closed circuit starting at the cooling unit leading to the detection module input connector Fig 32 4 approx 200 cm hose length From the latter the output connector Fig 32 5 approx 200 cm hose length leads back t...

Page 53: ...gned to the detection module of the camera port Cam1 CamLink A right and CamLink B left The lower slots are assigned to the unused port Cam2 CamLink A right and CamLink B left The power cable is connected to the 6 point power strip of the system The trigger cable is always connected to the detection module with 1 IN and on the rear side of the main system module Lattice Lightsheet 7 to the Camera ...

Page 54: ...ce to be set up near the system Do not place them on the system table to avoid vibrations The stage controller can also be mounted in the component rack Component rack and liquid cooling system LCS can alternatively be placed below the system table An extra computer table for the Storage PC would need additional space to the left of the System PC computer table It is recommended that the room is e...

Page 55: ...et system must be placed on an actively damped system table to sufficiently suppress vibrations Fig 35 Fig 35 Lattice Lightsheet 7 system footprint on small system table dim in mm The laser module and power supply unit for Lattice Lightsheet 7 can be housed inside the components rack which should be positioned beside the microscope stand on the floor ...

Page 56: ...tor control computer system PC sCMOS camera stage motion controller camera liquid cooling system The following components must be inserted in the 3 point power strip multipoint connector 2 or incubation components power strip see Fig 36 gas mixer controller temp module controller The folowing components must be connected to the 3 point power strip multipoint connector 3 or storage components power...

Page 57: ...the Lattice Lightsheet 7 are displayed in Fig 36 and Fig 37 respectively Line voltage 230 V AC 10 120 V AC 10 Line frequency 50 60 Hz 50 60 Hz Lattice Lightsheet 7 system incl lasers Max current 1 phase at 4 5 A 1 phase at 9 A Power Power consumption 900 VA continuous operation maximum 280 VA standby operation 0 025 VA off mode 900 VA continuous operation maximum 260 VA standby operation 0 011 VA ...

Page 58: ...SETUP REQUIREMENTS ZEISS Power Requirements Lattice Lightsheet 7 58 000000 2324 960 04 2021 V_01 Fig 36 Schematic drawing of power connections ...

Page 59: ...60 59 1 POWER 9 SUB LVDS 2 OVERVIEW CAMERA 10 CAMERA ADAPTER 3 SLM 11 GAS CAN Bus 4 TRIGGER STAGE 12 GAS Supply line 5 TRIGGER CAM 1 13 TEMP CAN Bus 6 TRIGGER OUT 14 TEMP Supply line 7 TRIGGER CAM 2 15 MICO IF 1 8 LVDS 16 POWER CONNECTOR Fig 37 Connectors and fuses of the Lattice Lightsheet 7 ...

Page 60: ...th a protective contact The protective effect must not be overridden by an extension cable without a protective conductor Detachable power cords should not be replaced with inadequately assessed power cords Only the specified power cords may be used Only the connection cables supplied with the device may be used The use of commercially available cables for use with the LLS7 is prohibited A safe di...

Page 61: ...bation Temp Control 17 x 22 5 x 9 2 0 Incubation Gas Mixer 17 x 22 5 x 9 2 0 Incubation Humidifier 7 x 11 x 30 1 6 Incubation Top Stage w lid 20 x 13 x 3 0 2 System PC 20 x 45 x 55 25 0 Storage PC option 22 x 64 5 x 42 40 0 Monitor HP Z32 32 71 5 x 56 4 x 24 8 12 2 Monitor HP Z27n G2 27 61 3 x 50 9 x 21 4 8 0 Camera pco edge 4 2 CLHS 12 3 x 7 x 7 6 1 5 Liquid Cooling System LCS 50 x 39 x 19 7 0 Co...

Page 62: ...ncubation System 67 5 x 33 5 x 18 5 2 Incubation Power Supply 29 x 29 x 13 7 0 6 Incubation Carriers 17 x 16 x 8 0 8 System PC w Keyboard Accessories 59 x 31 x 60 10 2 System PC Monitor 32 82 5 x 51 5 x 31 5 15 0 Storage PC w Ethernet Cable 80 5 x 62 8 x 35 1 37 6 Storage PC Monitor 27 74 x 43 5 x 28 10 2 Storage PC Keyboard 49 7 x 20 5 x 5 1 0 Storage PC Accessories 39 2 x 26 5 x 7 2 2 Liquid Coo...

Page 63: ...s different from item 1 and 5 3 Storage less than 16 h T 4 C to 30 C 4 Storage less than 6 h T 20 C to 55 C 5 Temperature gradient 0 5 C h 6 Warm up time 1 h 3 h for high precision and or long term measurements 7 Relative humidity 65 at 30 C 8 Operation altitude max 2000 m 9 Heat emission 0 8 kW 2 7 Vibrations According to ISO 10811 operation of the Lattice Lightsheet 7 system is recommended for v...

Page 64: ...0 04 2021 V_01 2 9 Components of Lattice Lightsheet 7 module 1 Transmission arm with LED 2 Scanning stage 3 Incubator 4 Slide in module for immersion 5 Immersion container 6 Syringe 7 Meniscus lens holder 8 Front door 9 LLS 7 module 10 Camera 1 module Fig 39 Components of Lattice Lightsheet 7 LLS 7 system ...

Page 65: ...SETUP REQUIREMENTS Lattice Lightsheet 7 System Overview ZEISS 04 2021 V_01 000000 2324 960 65 2 10 System Overview Fig 40 System overview of the Lattice Lightsheet 7 system ...

Page 66: ...stem as quick as possible to obtain some first images from his samples This short instruction does NOT replace the detailed information available in the full user manual For your safety observe the following instructions The Lattice Lightsheet 7 microscope including its original accessories and compatible accessories from other manufacturers may only be used for the purposes and microscopy techniq...

Page 67: ...ponents Fig 39 2 3 Switch on the Laser Fig 39 3 by turning the key to I position 0 position stands for Laser off The Laser ON LED Fig 39 4 will light up 4 Switch on the system PC and the storage PC if required 5 Switch on the incubation components if not already in on state 6 Start the application Switching off the System 1 Switch off the Lattice Lightsheet 7 system in reverse order It is not abso...

Page 68: ...are configuration without activating the system hardware for use but instead simulating the system hardware for training purposes ZEN image processing to ignore all hardware and to activate only data handling and image processing functionality for existing images After starting the system the ZEN Starting application will show up briefly followed by the ZEN Lattice Lightsheet Software GUI in initi...

Page 69: ... that the stage operates with absolute positions You can perform calibration also later by pressing the Calibration Home X Y Z button of the Stage LLS tool tab Fig 49 We recommend having no sample on the stage and the transmitted light arm in the upper position as the stage will travel a long distance at high speed This will prevent a shift of the sample a tear off the immersion and any potential ...

Page 70: ...operation mode 3 1 2 2 Selecting the Carrier Obligatory Fig 48 Sample carrier selection If no sample carrier has been selected the sample carrier display window will show None with a cross in a triangle with red background In this state it is not possible to run an experiment The Start Experiment button is greyed out In order to select a sample carrier press the Select button next to the sample ca...

Page 71: ...lly for z positioning using the same Carrier when cells are sparse 3 1 2 4 Priming immersions There are two possibilities to prime immersion manual with the syringe and by software via the pump For manual priming push the piston of the filled syringe Inspect the immersion media emerging on top of the meniscus lens If air bubbles are visible suck off immersion with a soft tissue paper and repeat un...

Page 72: ... Tray and Base We recommend starting with heating Lid and Tray only and set the Lid temperature 2 degrees above the wanted temperature Set the required atmosphere in the Atmosphere expansion field of the Incubation tab Fig 51 of the right tool area You can select O2 CO2 and Hum humidity We recommend starting with CO2 levels 2 higher than the wanted CO2 percentage and a humidity of 75 If heated to ...

Page 73: ...etup 1 Track Selection 2 Track Addition 3 LED illumination Transmitted Light 4 Stage with sample fixed 5 Laser illumination Epifluorescence 6 Illumination objective lens fixed 7 Detection objective lens fixed 8 Filter wheel 9 Camera pco edge 4 2 CLHS fixed 10 Microscope control 11 Incubation control 12 Input Output control Fig 52 Imaging Setup tab ...

Page 74: ... covered in y 4 Number of slices per stack 5 Step size Interval 6 Light sheet selection 7 Dither on off switch 8 Camera region of interest ROI in µm and pixel px Fig 53 Acquisition Mode tab Channels 1 Track selection and arrangement 2 Laser control 3 Camera Color selection 4 Camera exposure integration time 5 Display settings channel specific Fig 54 Channels tab ...

Page 75: ...Control 1 Movement of sheet along optical axis of illumination objective lens axis 2 Movement of sheet perpendicular to illumination objective lens axis 3 Compensation of aberrations in detection objective lens 4 Tilt compensation along X direction 5 Tilt compensation along Y direction Fig 56 Lightsheet Aberration Control tab Time series 1 Duration in cycles or period 2 Intervall between cycles Fi...

Page 76: ...e illumination as you will lose contrast Fig 58 2 Use None position in the filter wheel 3 Use 2048 x 2048 camera ROI 4 Set camera integration time to 5 ms 5 Press Continuous You will see an image with a sharp band Fig 58 Imaging Setup tab 6 Use the Ergopointer or the Stage Control Tab to focus the sample so that the sharp band is centered around the crosshair Fig 59 Fig 59 Transmitted image center...

Page 77: ...ight be two broad beam reflections be visible originating from the lower cover glass immersion and upper glass surface sample interfaces respectively Fig 60 You must focus on the upper of the two reflected beams In case of doubt move z until the lower cover glass reflex starts to appear then move back to the upper reflex Fig 60 Lower and upper glass reflection if not focused Fig 61 Reflected light...

Page 78: ...and optimize for the sharpest image 9 Position the beam waist of the illumination in the Field of view FoV by moving the Focus illumination to minimize out of focus blur It might be useful to first coarsely align to determine the region of focus and then fine focus by smaller steps Focus is where the line patten is sharpest You might also perform this step with Dither On and optimize for the sharp...

Page 79: ...nce the system is aligned only the stage is moved Lightsheet and detection objective are fixed focusing is done by the stage moving the sample If you move to different sample areas and the image gets blurry or defocused don t touch Lightsheet Aberration controls Instead just use the z wheel to bring back the fluorescent line to the center of the image Fig 66 Lightsheet aberration control Upper pan...

Page 80: ...of the stage automatically 2 Define the Scan range Range y or the number of slices Slices 3 Define the step size or Interval Default interval is 0 2 µm for Nyquist sampled imaging This should only be changed to coarser values if experiment data rate speed allows for 4 Limit acquisition to the minimal necessary camera sensor area to reduce camera read out time and data volume optional 5 Set time se...

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