The LSM 510 META confocal microscope is located on the sixth floor of the Medical School in the Neuroscience Imaging Center, room E632f. This state of the art laser scanning system can be mounted on either a motorized inverted stand (Zeiss Axiovert 200M) or a manual upright stand (Zeiss AxioImager Z1). The system offers eight excitation wavelengths (405nm, 458nm, 477nm, 488nm, 514nm, 543nm, 594nm, and 633nm) and for detection, three separate reflected light PMTs, each with its own adjustable pinhole and emission filter wheel. A transmitted light PMT is able to acquire phase and DIC images as well.

A unique feature of the LSM 510 META scan head is its ability to to acquire lambda stacks in 10 nm increments over a broad spectral range (411-753nm). A lambda stack collected from individual reference dyes can be used to generate an emmission fingerprint that can subsequently be used for spectral un-mixing of closely related fluors in the same sample.

The four most commonly used software modules (Multi-time, Physiology, FRET and FRAP) are available for on-line imaging and for off-line analysis at no extra charge. To facilitate live cell imaging, a Bioptics Delta T4 heating stage and objective heater can be used with both the widefield and confocal systems within the facility. For additional information and training, please contact Maryanne Pendergast at 368-2575.

• Maryanne Pendergast
  Training and Facility Manager
  368-2575 (office)
  216-224-8649 (cell)
  mxp19@case.edu

• Zeiss LSM 510 META laser scanning module
  
• Zeiss Axiovert 200M (motorized) inverted microscope
  
• Zeiss Axio Imager.Z1 upright microscope
  
• Diode laser (405nm)
  
• Multi-line Argon laser (458nm, 477nm, 488nm, 514nm)
  
• HeNe laser (543nm)
  
• HeNe laser (594nm)
  
• HeNe laser (633nm)
  
• Three reflected light PMTs; one transmitted light PMT
  
• Wide assortment of high quality objectives with both short and long working distances
  
• Bioptechs Delta T4 temperature control system for live cell imaging
  

• All prospective users are expected to have experience with basic epi-fluorescence microscopy including standard fluorescent or GFP techniques.
• Everyone must be trained and approved for independent operation of the system by Maryanne. The average training time is approximately 2-3 hrs, and is related to the user’s experience with microscopes and PC computers.
• Everyone is expected to use and care for the equipment as demonstrated during the initial training session. Abuse of the system may result in the cancellation of user privileges; repair costs will be charged to the lab at fault.
• Confocal assistance is available at an hourly rate for one-time evaluation experiments and infrequent users.
• Data can be stored short-term on the NIC server which is accessible to all computers within the facility as a mapped drive (Z:\); the server is available to other computers through a web interface. Data can also be saved to CDs, DVDs, and USB drives.
• Sign up during the day is first come/first serve and may be restricted to two half days per week during exceptionally busy times.
• Additional time is available after hours and on the weekend for experienced users (reduced rate, first come/first serve).
• Sign up should include user and PI names, time, and phone number.

• After your session please check the sign-up calendar:
  1. If the next user is scheduled within one hour, please leave the HBO lamp on and turn the main argon laser to stand-by. The other lasers may be turned off.
  2. If the next user is scheduled to arrive after one hour, please turn the system off.
  3. If you are the last user of the day, and someone is scheduled to use the system after hours, please call to ascertain that he or she will actually be coming. This is especially important for Friday evenings.

• Want to know how a microscope works? About color and light? Here is a site that has it all, including interactive java tutorials that are both educational and FUN!
  http://micro.magnet.fsu.edu/primer/index.html

• Concerned about possible spectral overlap between fluorescent dyes and resultant bleed-through problems? This interactive site will allow you to superimpose the excitation and emission spectra of multiple fluorochromes on a normalized axis. Use this site to plan the fluorescent dyes for your experiments.
  http://www.microscopy.bio-rad.com/fluorescence/fluorophoreDatab.htm
  

• The confocal listserv is maintained by the University of Buffalo and has archived discussions dating from November 1991. Topics covered include anything and everything having to do with confocal microscopy. It’s great for trouble shooting problems, and also for the breadth of information available. A visit to this site is a must for all confocal users!
  http://listserv.acsu.buffalo.edu/archives/confocal.html

• Website for Carl Zeiss, North America
  http://www.zeiss.com

Zeiss LSM 510 META Guided Tour

Zeiss LSM 510 Beam Configuration Tutorial

Zeiss LSM 510 FRET Tutorial

Zeiss LSM 510 Multi-Time Tutorial

Zeiss LSM 510 META Complete Operation Manual (602 pp)

CWRU Laser Safety Manual

OSHA Laser Safety