CLSM (Leica SP5) organisation and rules (Room G03.058, Biocenter Martinsried)
First, three basic, but crucial rules to keep the 'hardware' running (not following = not allowed to work at the CLSM!):
Different rules exist for using FLIM and/or the multiphoton laser. If you intend to use these techniques, contact the responsible person.
- ONLY use the special optics-paper to clean objective-lenses. If empty, you organise new one, but you will NOT use Kleenex or any kind of other tissues. They may look clean and bright-white - and this often is the problem - may contain chalk or so >crystals< ... micro-scratches on the lens - serious loss of resolution.
As a lens cleaning solution, you may use the provided alcohol mixture, containing 70% isopropyl alcohol (= IPA, isopropanol, 2-propanol, propan-2-ol, ) and 0.1% butane-1,3-diol (= 1,3-butylenglycol) in distilled water.
- ALWAYS cover the microscopes when not in use. Dust is getting into the microscopes, and you will >never ever< be able to get rid of it again. The time a microscope is exposed to dust is directly proportional to the time you can use it with good resolution.
- CORRECT entries in the logbook are mandatory. When you enter your usage-times (1/4 h is exact enough, no need for writing down in minutes), do it in a way that a realistic number comes out at the end (e.g. do not always round down).
- Before using the CLSM for the first time, you will get an instruction by Thomas Ott.
- Always make a reservation for the CLSM via the booking system.
- At the machine: before (!) you switch on the CLSM, enter your name and the starting time in the logbook!
- For non-FLIM work, please only use the standard PMTs (No. 3, 5), not the FLIM PMTs (2, 4). The standard PMTs are more flexible and can be set to higher sensitivity, whereas the FLIM detectors may be photo-damaged - they do not have any advantage for standard CLSM, so only use the standard PMTs for standard CLSM. When using the HyD detektor (position 1), please mind the extra precuations that need to be taken to operate it.
Using the mercury burner lamp
- Before switching on, check the log-book and respect a 30 min cool down time after the lamp was switched off. Otherwise you damage the bulb, the lifetime decreases considerably, and you risk mercury (toxic!) releasing im-/explosion and (expensive) bulb-housing damage.
- Never look into the UV light spot on the object level without a UV protection screen.
- Never cover the air duct on the lamp housing - the bulb is extremely hot.
- Only authorized persons may replace high-pressure mercury bulbs. To minimize risk of explosion, it has to be replaced at the end of its rated life (250 h in our case).
- Remove your slides, and turn down the microscope stage before you switch of the microscope! We had a damaged glass slide stuck in the condenser in the past! If something like this happens, you HAVE TO tell us!
- If next user has not yet arrived and you are unsure whether she/he will come: switch off the microscope (don't forget what you learned in the CLSM intro) and the Hg-lamp. If another user follows - a rule: for a 1-2 h break, the system should not be switched off, but set to 'stand by'. For any longer breaks, switch off. However, you need to know for sure that somebody will indeed come, afterwards. If not sure, finish and switch off everything.
- Resord the times for the system, lasers, mercury lamp in the log-book. Also write down if only the computer was used, even if only for 1 min. Every usage has to be recorded.
- Clean up - remove any dirt or liquids form the microscope parts. Use paper and alcohol.
- For cleaning lenses never ever use normal paper or Kleenex! Only use the provided special optics paper! Never ever use acetone for cleaning! If paper is empty, ask the responsible persons, or have a look in the microscopy room, next door!
- Cover the microscope for dust protection (most important!).
- Dispose any used paper and glass in the respective bins. This has to be a clean room. Please, also clean up things from other persons in case they forgot to do so. You are responsible for leaving a clean room !
- Turn off the room light, lock the door (!).
- The doors of the microscopy room have to be locked while nobody is in it.
- Dust considerably shortens life and performance of the microscopes. Also the workbenches have to be clean. No water allowed to stand close to the microscope or electronics!
- Keep dirt out of the microscope room!
- Report any problems and damages immediately !!!