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Leica TCS SP8

Quick Start Guide

Leica TCS SP8 System Overview

Start-Up Procedure

1. Turn on the CTR Control Box, EL6000 fluorescent light source for the

microscope stand.

2. Turn on the Scanner Power (1) on the front of the Compact Supply Unit (CSU)

3. Turn on the Laser Power (2)

4. Enable the lasers by turning the key

from the Off position to the On

position (3).

5. Turn on Workstation and Monitor

6. Log into user profile

7. Double click on the LAS X icon to start the software.

8. Select Machine for the Configuration and DMi8 for the Microscope. (the

simulator is used to view and analyze images and not acquistition)

10. Click OK to start the initialization of

the LAS AF (Leica Application Suite

for Advanced Fluorescence)

11. A message will appear asking whether you want to initialize the stage. Initializing

the stage is required to activate the Tiling and Mark & Find features in the software.

If you select yes, the stage is going to

move to all 4 corners to calibrate the

position of the stage.

Make sure there is nothing on the

stage that will hit the condenser

before you select yes!

Shut-Down Procedure

1. Turn off the lasers in the software

2. Lower the objective to the lowest position, remove specimen and clean all of the

objectives

3. Save all unsaved data and exit LAS X.

NOTE: Do not turn off scanner and/or CTR control box for the microscope before the

software is closed

4. Turn off the switches on the front of the laser supply unit in the reverse order (#3-#1)

of start-up (ie. turn key off, then turn off Laser Power and then turn off Scanner

Power)

5. Turn off microscope control box

The LAS X software will open in the Acquire tab. There are 3 portions to this window:

Scan Parameters Light Path Image

Turning on the Lasers:

1. Click on the “+” in the Laser Lines Box in the Beam Path Settings Window to directly

open the main Laser Control Window

2. Activate the lasers you require

3. Alternatively, click on the Configuration tab and the Laser icon to open the main

Laser Control Window

NOTE: Only turn on the laser(s) with the appropriate laser line(s) that will excite

the fluorophores you are using

Additional Tools in the Configuration Tab…

1. Customize the USB Control Panel -assign various parameter and sensitivities to

the various knobs of the control panel

2. Specifications of the Objectives equipped on the microscope

Save and Load the customized settings

Alternatively , click on the Control

Panel Icon in the Beam Path Settings

Window as a short cut to the USB

Control Panel Window

Additional Tools in the Configuration Tab Continued…

3. Dye Database with the excitation and emission spectra of common fluorochromes

Emission spectrum from a lambda scan can be added to the dye database.

Many manufacturer’s will also provide the data for fluorophores that can added to

the database (see LAS AF Help)

Another great resource is the Leica FluoScout

interactive tool

http://www.leica-microsystems.com/fluoscout/

Beam Path Setting

Option #1: Manual setting of the beam path configuration

1. Click On to activate

the lasers 2. Adjust the laser intensity

of the appropriate laser

line(s) by moving the

slider up or by directly

entering the level (start

low as a suggestion).

4. Use Autoselect to select the beam splitter

(inactivate and manually select if BSC is red)

3. Select the appropriate objective

6. Select the appropriate emission

spectrum (if available) from the

dye data base to use as a guide

8. Define the of emission to be collected with the sliders

5. Click On to activate the

appropriate detector(s)

Keep in mind that the emission spectra

displayed are to be used as a guide and are not

defining the wavelengths collected. The

specific wavelengths collected are determined

by the position of the gates.

Alternatively, double click on

the slider to open a window

that allows you to directly

enter the start and end

position of the gates.

7. Select the Pseudocolour