Leica Sp5 WLL STED Operation instructions

1

Wellcome on the STED Tutorial

Before using the Sp5 WLL STED microscope ,

Version 1 27.08.2013

You will need to put down your name on the reservation system =

https://sv-bbs.epfl.ch/ptbiop/

STED, on the SP5WLL.

2

Welcome to the Leica Sp5 WLL

Confocal Invert tutorial.

System start Up. (Page 3-8)

Laser switch and calibration (page 11-13)

Sted configuration(page 14-17)

System shut down. (page 18-22)

3

System Start up (step 1)

On the STED Tower, Switch “ ON ” the button

4

System Start up (step 2)

The SHG led turn green for 3sec,

then it’s turn orange for 3 minutes,

then it’s turn green again after 1 minutes.

5

System Start up (step 3)

Turn the key from the depletion laser 592-nm in

vertical position.

The SHG led keep green.

6

System Start up (step 4)

Turn on the PC/Microscope button

Turn on the Scanner Power button.

Turn on the Laser Power button (White laser is

“on”).

Turn the Key on the right position (Argon laser

is “on”).

7

Turn “on” the XCite Light.

System Start up (step 5)

8

System Start up (step 6)

When the computer is on

Introduce your Username

Introduce your Password

Introduce your login

Start the LASAF software.

Under “Configuration” select Machine STED.

Do OK

9

Choosing the objective

Under Acquire and Acquisition

Choose and select your objective

in the list.

To acquire a STED image you need to use only the

HCX PL APO CS 100x 1.40 OIL

10

Choosing the Filter

You need to introduice a Notch filter to stop the laser 592nm

Select “Analyzer Filter”

Deselect “ Auto Select”

Select “NF STED ”filter

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