Zeiss LSM 710 Manuel utilisateur

Carl Zeiss LSM710 /780 ZEN

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This Quick Guide describes the basic operation of the LSM 710, LSM 780, LSM 710 NLO,

LSM 780 NLO and ConfoCor 3 Laser Scanning microscopes with the ZEN 2012 software.

The purpose of this document is to guide the user to get started with the system as quick as

possible in order to obtain some first images from his samples.

This Quick Guide does NOT replace the detailed information available in the full user

manual or in the manual of the respective microscopes (Axio Imager, Axio Observer, Axio

Examiner).

Also, this Quick Guide is written for a user who is familiar with the basics of Laser Scanning

Microscopy.

For your safety! ……………………………………………………………………………… 1

1. Starting the System…………………………………………………………………………… 2

2. Turning on the Lasers………………………………………………………………………… 6

3. Setting up the Microscope ………………………………………………………………… 7

4. Configuring the Beam Path and Lasers …………………………………………………… 8

5. Scanning an Image ………………………………………………………………………… 12

6. Storing and Exporting Image Data …………………………………………………… 18

7. Z stack ……………………………………………………………………………………… 22

8. View Tabs …………………………………………………………………………………… 24

9. View Option Control Tab ……………………………………………………………… 27

10. Operation of Microscope ………………………………………………………………… 30

11. Switching off the System ………………………………………………………………… 32

1. Notes on Handling the Laser Components and Illumination Systems

･The LSM systems are laser hazard class 3B instruments. If equipped with a Ti:Sa Laser, the LSM systems are

devices that belong to laser hazard class 4.These moderate and high-risk classes embrace medium-power and

high power lasers. You must take care not to expose yourself to the radiation of such lasers. In particular,

never look into the laser beam!

･Do not remove cables and optical fibers connected to microscope, scan module and laser module.

2. Notes on Care, Maintenance

･If spilt some water-soluble buffer (culture medium, etc), remove it by wiping with a dust-free cotton cloth or

a moistened cloth immediately. Before cleaning, switch off the microscope and disconnect it from the main

power supply.

･Remove oily or greasy dirt (immersion oils, finger prints) using cotton wool buds or a dust-free cotton cloth

and Optical Cleaning Mixture L. (This cleaning mixture is manufactured from 85% vol n-hexane and 15 % vol

isopropanol)

･Do not touch lamp housing when it is hot. Always check that the instrument is switched off and cooled

down before covering.

3. Observe the following instructions:

･In the Operating Manual, read the chapter Safety Instructions carefully before starting operation.

･Follow the safety instructions described in the operating manual of the microscope and X-Cite 120 lamp /

HBO 100 mercury lamp.

Contents

Introduction

For your safety

Carl Zeiss ipjTNM L TUM=wbk=

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※Safety lock

1. Starting the System

1-1. Starting the System

1-1-1. Switching on the LSM system

1-1-2. Switching on the X-Cite 120 or the HBO 100 mercury lamp

1-1-3. Switching on the Ar Laser

1-1-4. Switching on the Ti:Sa Laser

1-1-1. Switching on the LSM system

①Switch on the Main Switch of the Power remote

switch.

Check the safety lock is ON.

②Switch on the SYSTEM/PC and Boot up the computer.

・SYSTEM/PC provides power to the computer. This

allows use of the computer and ZEN software offline

・ Log on as ‘LSM user’.

③Switch on the COMPONENTS.

・COMPONENTS starts the other components and the complete system is ready to be

initialized by the ZEN software.

1-1-2. Switching on the X-Cite 120 or the HBO 100 mercury lamp

・ Switch on the main switch of the X-Cite 120 / HBO 100 lamp for reflected light

illumination via the power supply as described in the respective operating manual.

Carl Zeiss LSM710 /780 ZEN

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1-1-3. Switching on the Ar Laser

①If the Ar laser is required, switch it on via the toggle switch (Fig.1/①) on the power supply

②Turn the key from (0) to (I) direction (Fig.1/②). The laser is automatically kept in standby

mode for 5 minutes to warm up.

③Set the idle-run-switch from idle (Fig. 1/③down) to run (up). It takes about 50s until the

laser has reached the set output power (green LED) provided the warm up time of 5

minutes is already completed.

④Adjust the required power level with the control knob (Fig. 1/④) (default position should

be 11 o’clock). The adjustment range is from standby (left stop) to 7 A (green LED); tube

currents greater 7 A will show a red LED.

※The red LED indicates the power range which

is critical for the lifetime of the laser!

1-1-4. Switching on the Chameleon Laser

・If the Chameleon (Coherent) laser is required, turn the key in front of the power supply

from Stand by to ON.

1-2. Starting ZEN software

Double click the ZEN icon on the desktop to start the Carl Zeiss LSM software. The

ZEN Main Application window and the Startup window appear on the screen. Choose

Start system to start the system hardware for acquiring new images.

Start System ：initializes the whole microscope system and activates the entire software package

for new image acquisition and analysis.

Image Processing ：ignores all hardware and activates only data handling and image processing

functionality for already acquired images.

※Toggle the little symbol to show the Boot Status display.

①

②

③

④

Fig.1 Power supply of Ar laser

Carl Zeiss LSM710 /780 ZEN

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《Introduction to ZEN –Efficient Navigation 》

The ZEN 2012 interface is clearly structured and follows the typical workflow of the experiments

performed with confocal microscopy systems:

Left Tool Area（Fig.3-①,②,③）

The user finds the tools for sample observation, image acquisition, FCS, image processing and

system maintenance, easily accessible via four Main Tabs.

①Main Tabs ②Action Tabs ③Tool Group

Center Screen Area（Fig.3 - ④,⑤,⑥）

This area is for viewing. Each displayed image can be displayed and/or analyzed with many view

options available through view tabs which can be found on the left side of the image.

④Image Window ⑤View Tabs ⑥View Controller

Right Tool Area

File management and data handling tools.

⑦Open Images list (Thumbnail view)

Show manual tools

With Show manual tools activated, the

Setup Manager tools are displayed. The user

can edit the configuration from Light Path

tool.

Fig.4 Show manual tools

①

②

③

④

⑥

⑦

⑤

Fig.3 ZEN Main Application Window after Startup

Carl Zeiss LSM710 /780 ZEN

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Fig.5 Show all

Show all

With Show all de-activated, the most

commonly used tools are displayed. For each

tool, the user can activate Show all mode to

display and use additional functionality

More features of ZEN 2012 include:

−The user can add more columns for tools to the Left Tool Area or detach individual tools to position them

anywhere on the monitor. To add a column, drag a tool group by the title bar (e.g., Online Acquisition) to

the right and a new tool column automatically opens. Alternatively use the context menu "move tool group

to next column". To detach a tool, click on the little icon on the right end of the blue tool header bar

−Another unique feature in the Imaging Software is the scalable ZEN interface. This Workspace Zoom

allows adjustment of the ZEN 2012 window size and fonts to the situational needs or your personal

preferences

−Setting up conventional confocal software for a specific experiment can take a long time and is often

tedious to repeat. With ZEN these adjustments have to be done only once –and may be restored with just

two clicks of the mouse. For each type of experiment one can now set-up and save the suitable

Workspace Layout. These configurations can also be shared among users.

※These are just some of the most important features of the ZEN interface. For a more detailed description of

the functionality for the ZEN 2012 software, please refer to the User Manual that is provided with your system.

Fig.6 ZEN Window Layout

configuration

Carl Zeiss LSM710 /780 ZEN

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2. Turning on the lasers

ZEN 2012 operates all lasers automatically. Whenever they are used (manually or by loading

configuration), the lasers are turned on automatically.

To manually switch lasers on or off:

Check the Show manual tools tick box and open the Laser tool. All available lasers can be

operated within this tool (Fig. 7), except Ar laser. Select the required Laser, and turn on it

from the pull down menu.

Fig.7 Laser Control tool

・Blue Diode 405 （405nm）

The Diode lasers is switched on automatically after Software start

・Argon（458/488/514nm）

The Argon laser has to be switched on by manually. (Page2)

・DPSS (561nm)、HeNe (543nm、594nm、633nm)

Switch the requested lasers on by using the pull down menus

・MaiTai (Spectra Physics), Chameleon (Coherent)

Switch the requested lasers on by using the pull down menus

1. Starting the system, and Turning on the Lasers

↓

2. Microscope Observation via Eye pieces

↓

3. Configuring the LSM Beam path for used Dye

↓

4. Scanning an Image

Work flow to acquire image

Carl Zeiss LSM710 /780 ZEN

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3. Setting up the microscope

The Locate and Acquisition buttons switch between the use of the LSM and the microscope:

• Click on the Locate tab for direct observation;

press the Online button for your actions to take

effect immediately. In this tab, lasers are blocked.

• To close the light shutters on the microscope, click

Offline.

Open the Ocular tool to configure the components of your microscope like filters,

shutters or objectives. Place specimen on the microscope stage. The cover slip must be facing the

objective lens. Remember the immersion medium if the objective lens chosen requires it!

Transmitted light

・Open the graphical pop-up menu by clicking on the

Transmitted Light icon. Click on the On button. Set the

intensity of the Halogen lamp using the slider.

Selecting an objective

・Click on the Objective icon and select the objective lens for

your experiment. The chosen objective lens will automatically

moves into the beam path.

Reflected light

・Click on the Reflected Light shutter to open the shutter of the

X-Cite 120 lamp / HBO100.

・Click on the Reflector button and select the desired filter set by

clicking on it.

Storing the microscope settings

Microscope settings can be stored as configurations. These configurations can be assigned to buttons

that are easier to set up the microscope.

Fig.9 Microscope Configuration panel

Changing between direct observation and laser scanning mode

Setting up the microscope and storing settings

Fig.8 Microscope Control window, e.g.: Axio Observer. Z1

Carl Zeiss LSM710 /780 ZEN

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4. Configuring the beam path and lasers

Click on the Acquisition button to change to the LSM mode.

There are 3 ways to configure the beam path and lasers.

We highly recommend to use ‘4-1.Settings for track configuration’ especially for a user

who is not familiar with LSM.

4-1. Settings for track configuration

For loading an existing configuration, click button in the Configuration and select

the appropriate one from the list box.

ex) Single staining -> ‘Ex 488 (GFP, FITC, Alexa488)’ for Green fluorescence, ‘Ex 561 (Cy3,

Alexa568, Rhodamine)’ for Red fluorescence, etc..

Multi staining -> ‘Ex 488/561’ for Green and Red double staining, ‘Ex488/561/633’ for

Green, Red, and Far-red triple staining, etc…

※Please do not overwrite / delete existing configuration!

Fig.10 Acquisition Window

Fig.11 Configuration list

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