2021B CryoTEMs for public use at SPring-8

Contents

1. Outline

CryoTEMs are offered for the contribution and the promotion of research and the enhancement of the competitiveness of users of the SPring-8 public beamline facilities in the field of structural biology and drug discovery. CryoTEMs will be available for use as ancillary facilities and will be treated in analogy to X-ray beamlines for research proposals in structural biology. Therefore, the expenses for CryoTEM use will be charged in the same way as it is for the use of SPring-8 public beamlines.
Proposals approved for the 2021A term (2021A2700 to 2021A2899) are valid for the 2021B term, however, for the usage of BL38B1 and CryoTEMs during the 2021B term it is necessary to make separate applications.

1.1 Mode of Operation

The following operation procedures will be applied to General Proposals, both Non-Proprietary and Proprietary. Budding Researchers Support Proposals will be treated the same as General Proposals (Non-Proprietary research).

1. CryoTEMs are operated as a part of proposals in the field of Life Science/Protein Crystal Analysis (L1).
2. CryoTEMs are only available for research proposal that use both X-ray beamlines and CryoTEMs.
3. Available proposal types for CryoTEMs are General Proposals (Non-Proprietary / Proprietary), Budding Researchers Support Proposals and Proprietary Time-Designated Proposals.
4. A questionnaire on the machine time allocation (to determine the number of shifts, date of use, desired microscope), termed “beamtime request survey”, will be completed by project leaders approx. 5 times a year. Prior consultation with CryoTEM staff regarding the schedule is necessary for time-designated use of CryoTEMs.
5. Public use of CryoTEMs will start from 2021B as trial operation.

1.2 CryoTEMs Available to Users

The following two CryoTEMs are available to users.

• EM01CT: CRYO ARM 300 (K3)
• EM02CT: CRYO ARM 200 (K2)

2. Application and Questionnaires for Machine Time Allocation

Applications of General Proposals (Non-Proprietary /Proprietary Research) and Budding Researchers Support Proposals should include the statement that the use of both, beamlines as well as CryoTEMs, is planned. In addition, a description of how protein sample and protein crystals, even if protein crystals have not been obtained yet, are to be prepared for measurements must be provided. Every submitted questionnaire should also include a description of the status of protein or complex preparation. This can be done by using results from size exclusion chromatography (SEC), crystallization trials, native and denaturing PAGE, negative staining EM or CryoEM.
Since applications of Time-Designated Proposals need to be accompanied by applications to other beamlines, please select the appropriate beamlines at application. In addition, please note that prior consultation about the experimental schedule with staff of the beamlines and CryoTEMs is necessary.

3. Notes on the Use of CryoTEMs

Participants in CryoTEM experiments are required to attend the pre-use training course*. It is free of charge.

• Users will carry out operation of CryoTEM during the allocated machine time (a unit of 3 shifts, starting at 10:00 a.m. and ending at 10:00 a.m. the next day) of the experiment.
• In general, the facility staff will handle the loading of samples into the CryoTEM. However, sample loading is performed at designated times as described below**.
• Vitrification equipment for cryo-specimen preparation is available for use in person at the facility.
• Together with automated data collection, we provide an environment for image processing (motion correction, and CTF estimation, which however, does not include full structural analysis, i.e. 3D reconstruction).

* This training course is intended for first-time users of CRYO ARM (JEOL) and teaches the minimum user requirements of operating the CryoTEMs in the facility.
Content: Software operation (JADAS: made by JEOL) for data acquisition after the sample has been loaded by the facility staff. This takes 1-3 days depending on prior experience.
Time period: In addition to the allocated machine time, courses are regularly held every other week during the operation period and can also be taken even before the approvals of the proposals.
** In principle, the timing of sample loading is assumed to be around 10:00, after 13:00, and before 17:00 during the allocated machine time. A maximum number of 12 grids can be loaded at one time. If you wish to perform independent sample loading and image acquisition, it is required to take the CryoTEM operation course for beginners (practical training for sample loading for independent on-site measurements) in addition to the pre-use training course described above.

4. Fees for Use of CryoTEMs

As with general proposals for other beamline usage, you are required to pay the following fees.

Machine Time Fee:
• General proposal (Non-Proprietary) and Budding Researchers Support Proposals: Exempted (Publication of results in peer-reviewed journal publications, etc. within 3 years after the end of the term of use is mandatory.)
• General proposal (Proprietary) 80,000 JPY/shift*
• Time-designated use proposal (Proprietary) 120,000 JPY/shift*
※ The fees for CryoTEM use may be revised based on actual usage results of the trial operation in 2021B.

User Fees:
• Fixed fee for consumables is 10,720 JPY/shift* (same as beamline use) for any types of applications.

* 1 shift = 8 hours operation

5. Contact

(1) Information Desk for CryoTEM

Center for Synchrotron Radiation Research
Protein Crystal Analysis Division, JASRI
Director, Takashi Kumasaka
E-mail: kumasaka＠spring8.or.jp / TEL: +81-(0)791-58-0833
Or
Hideki Shigematsu
E-mail: hideki.shigematsu＠riken.jp / TEL: +81-(0)791-58-2839

(2) Information Desk for Procedure of Proposal Application