Access to the Cryo-Electron Microscopy Facility

Training of new individuals outside of the Divisions of Biological and Physical Sciences is suspended for the near future

Facility Availability
Hours of Operation
Sphera Training
Polara Training
Costs of Training and Microscope Use
Acknowledgement in Publications
Contact for Training and Instrument Use

Facility Availability

The Cryo-Electron Microscopy facility is available to UCSD faculty, staff and students and also to members of the non-profit research institutions on the Torrey Pines Mesa. Outside work may be performed on a limited basis. The Cryo-Electron Microscopy Facility does not do sample preparation. Users are expected to determine what preparation techniques are needed for their own specimens and to do them in their own lab. The only exception is negative staining and plunge freezing of particulate samples for which some stains are available and a cold room with a plunge-freeze device may be used.

Users will also be expected, with the proper training, to operate the instruments themselves. The Facility Manager is readily available, however, to answer questions, fix problems and load samples into the Polara. Examination of samples and collection of data may be done by Facility staff on an initial basis or if the projects are directly related to Dr. Timothy S. Baker’s (PI of the facility) research.

Users who need assistance with sample preparation and instrument operation are encouraged to contact the Dr. Marilyn Farquhar laboratory on the UCSD campus. Scanning electron microscopy is done by the Scripps Institute of Oceanography (Dr. Evelyn York, (858)534-2438).
Top

Hours of Operation

The operating hours of the facility are from 9 a.m. to 5 p.m. Monday through Friday. Trained users who have demonstrated a proficiency with the Sphera and Polara, and who have been given permission by the Facility manager, are allowed unrestricted access seven days a week.
Top

Sphera Training

Training to use the Sphera is fairly intensive. It is expected that the user be proficient with the instrument before using it unsupervised and not just be able to push the buttons. Training averages about 25 – 30 hours per individual and it is usually scheduled with half day a week sessions. Accommodations in the training routine are made for those individuals who have had prior microscopy experience. Users are given a microscope account on the Sphera, access to the on-line Scheduler, access to the UCSD CryoEM listserve, and an account on other computers in the facility for the temporary storage of digital data. Sessions are broadly based around the following topics:

Initial meeting to describe the process and find out the research needs of the operator – 1 hour
Description of the column, control pads and the vacuum system – 4 hours
Description of the Tecnai operating system and sample holder insertion and removal – 3 hours
Microscope log-on procedure, personal customization of the Tecnai user account, insertion of the sample holder under supervision and initial microscope practice time – 3 hours
Condenser and objective aperture selection and alignment, condenser astigmatism correction, logging on, eucentric height adjustment, gun alignment, accessing saved alignments, and correcting rotation center – 4 hours
Introduction to scattering and phase contrast and focusing the image, introduction to the Gatan CCD camera, collection of images, computation of Fourier transforms, and saving images – 4 hours
Review and practice of log-on procedures and initial alignments, description of image astigmatism correction, practice time – 3 hours.
Supervised startup, operation, and shutdown, practice collecting images – 3 hours.
The student is then encouraged to set up an initial practice session where he/she independently operates the instrument. The Facility Manager remains in the area and helps as needed. Additional explanations and demonstrations will be given as necessary – 3 hours.
The student schedules time for themselves on the Scheduler during the normal working hours. When proficiency is demonstrated, 24 hour access may be granted.

Additional training sessions may be scheduled based on the needs of the user. Such sessions could include:

Data collection on film and use of the darkroom
Low dose training
Preparation of continuous carbon and holey carbon support films
Negative staining
Advanced microscope alignment
Cryo-microscopy (multiple sessions)

Top

Polara Training

Use of the Polara is restricted to those individuals who specifically need the features of that instrument, specifically the more stable stage, the field emission gun (FEG), the higher accelerating voltage, the tomography software or the automatic data collection software. It is expected that users of the Polara will already be accomplished cryo-microscopists and be familiar with the Tecnai operating system. Accommodations will be made for those individuals with extensive microscopy experience on other microscopes. Infrequent users of the microscope will not be allowed to load cartridges into the multi-specimen holder or insert them into the microscope. Training for this instrument will include operation of the FEG, adding nitrogen to the dewars, using the cryo-transfer station, and using the multi-specimen holder and insertion rod.
Top

Costs of Training and Microscope Use

As of this time (May 2007) there are no charges for training or for the use of the facility. Preparations are being made to institute a recharge system but it is not known at this time when that will be put in practice.
Top

Acknowledgement in Publications

It is requested that an acknowledgement be given in all published journal articles where data collected in the Facility is used. Please forward all journal citations to Norm Olson. We would also appreciate it if our funding sources were mentioned like this:

"We acknowledge the use of the UCSD Cryo-Electron Microscopy Facility which is supported by NIH grants 1S10RR20016 and GM033050 to Dr. Timothy S. Baker and a gift from the Agouron Institute to UCSD."
Top

Contact for Training and Instrument Use

Please contact Norm Olson via email or at (858)822-6718.