2.1 Basic Confocal Operation - A Check List
Guide |
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The numbered sections (2.1.1-2.1.5) of this pilots' check list guide should be completed in
order. Numbers in parenthesis refer to items in this microscope
diagram. (This panel should open in another window. It
may be obscured by other windows)
2.1.1. Power Up the Confocal System
- In warm weather ensure that cooling system is on. Panel is on wall
just to the left of the electrical box behind the door to room 006.
- Remove dust cover gently from microscope
- Ensure that microscope is powered off (1)
- Switch on mercury arc lamp (if conventional fluorescence is required) let
lamp strike (about 15-30 seconds)
- Switch on microscope power (1)
- Turn laser power controls to minimum
- If 568 nm (green, Rhodamine, Texas Red, TRITC or CY3 like) excitation is
required switch on Krypton (Kr) laser fan and ignition key
- If 488 nm (blue, FITC , CY2 or Alexa 488 like) excitation is required
switch on Argon (Ar) laser fan and ignition key
- Switch on scan electronics
- If 633 nm (CY 5 like) excitation is required switch on Neon-Helium (HeNe)
laser key
- Switch on both computer monitors
- Switch on work station computer (\\Leica_tcs) if not already running
- Login on to WindowsNT with assigned name and password and Physiology
domain
- Click on TCS_RUN program icon (this
may take a few minutes to start) If errors occur, close program, do not save sheets and
then restart TCS_RUN
2.1.2. Setup the Microscope for Conventional Viewing
- Setup microscope for Kohler
illumination (see Smooth Starts for detailed
explanation)
- Close scan head shutter (17)
(top
right hand plunger rod in)
- Locate specimen and focus onto it using transmitted light
- Set upper limit. Do not
set lower limit. (Clear lower limit if it has been set.)
- Open fluorescent light diverter (a.k.a.
mercury lamp block off rod) (14)
(pull rod out on left side of scope firmly)
- Set fluorescent filter dichroic wheel (15)
to 3 for fluorescein, 2 for texas red, or scan (also for transmitted illumination)
- Locate and check specimen fluorescence by eye
2.1.3. Setup the Microscope for Confocal Scanning
- Fluorescence light diverter rod (14)
pushed in
- Filter/Dichroic wheel (15)
rotated to position 1 or 4
for scanning (the "scan" sticker has fallen off)
- DIC wave plate (16)
pulled out (unless doing
simultaneous DIC+confocal)
- Walliston prism wheel (13)
set to "H"
(unless doing simultaneous DIC+confocal)
- Scan head shutter (17)
rod out
2.1.4. Setup the Computer for Scanning
Setting
Microscope To Conventional Viewing (again)
- Pull Fluorescence illumination shutter (14) slider
out if using fluorescence
- Rotated in appropriate filter/dichroic wheel (15)
- Push Scan head shutter (17) lever in
- When looking down eye pieces re-adjust focus (galvo stage may have moved)
2.1.5.
Shut Down and Clean Up the Confocal System (see also Smooth Starts & Clean Finishes)
- Turn Argon and Krypton laser power controls to minimum (black knob)
-
Check
bookings for today
- Krypton (green) or Argon (blue) lasers should have been run at minimum power for 10
minutes if they are going to be keyed off (see next step)
- If no one is booked to use the confocal in next 3 hours turn off laser ignition keys. Important:
leave fans running for now
- If no one is expected to use mercury arc lamp in next 1 hour turn it off
- Only leave on lasers if the next user(s) are going to use in next 3 hours
- If system is to be left on please ensure that the next user is actually planning to
arrive. Contact information is provided on the
booking
page
- If next user's arrival is in doubt please favor turning off the system, especially if
next user is the last booked user of the day
- Exit LEICA LCS program (File -> Exit)
- Log off WindowsNT (control+alt+del -> Logoff). Please do not
shut down WindowsNT or turn off computer
- Turn off monitors if you are the last person using the system for the day or if you know
that no one else will be using the system in the next hour
- Turn off scan electronics
- Turn off NeHe (red) laser, if on
- Lower lens turret
- Remove slide
- Clean off immersion oil with lens tissue (never Kim
wipes etc. They contain scratchy fiber glass!)
- Delete upper (and lower) travel limits on microscope
- Swing 10X objective into position
- If mercury lamp has been turned off place dust cover over scope when arc lamp housing is
not hot
- When laser vent pipes behind laser units feel barely warm (~2 minutes after keying off
lasers) power off laser fans (red switches))
Last update 03-07-31 version
2.9
