Cryo grids quality control policy
For users sending in ready-to-image cryo grids, we need to see 1) a couple of high magnification images to show how the sample look like, particle distribution and presence/absence of aggregation, degradation and contaminants; 2) a few low magnification images covering multiple grid squares to show ice thickness, distribution and there are enough usable grid squares on one grid for high throughput data collection. This is to ensure that suitable cryo grids with proper ice thickness, good particle distribution and plenty of usable area on a single grid are available to best use valuable microscope time.
To collect data on one sample, the cryoEM facility should only load up to 8 cryo grids all during one load among which one good grid will be chosen to set up data collection (one good cryo grids can easily last for more than 1 week’s data collection). No significant amount of Titan Krios time or staff hours should be used for screening grids to make the best use of valuable resources here and to maximize throughput.
Requirements on sending in cryo grids
If a user needs a guideline on how to ship cryo grids, he/she can contact the facility for detailed instruction as well as the shipping address.
Our preferred grid is the Quantifoil 1.2/1.3 type. Our data collection protocol is optimized for this type of grid. If you plan to use a very different type of grid please contact the facility for a discussion.
For a scheduled data collection session, the cryo grids should arrive at the facility 1-5 business day before the start of the session if the user is shipping the grids. If the user is bringing the grids with him/her, it is fine for the grid to arrive on the morning on the first day of the scheduled session.
For one sample we typically load 6-8 cryo grids. To save sample handling and loading time, the 6-8 grids that will be loaded into the Krios should be stored in two cryo boxes instead of being scattered in multiple boxes. The two cryo boxes should be clearly labelled and tightened properly. User should choose cryoboxes that can be loosened with a commonly available tool such as a small Philip or slot screw driver.
For users with access to autogrids clipping tool (these are users with access to Titan Krios or Talos Arctica at their local cryoEM facility), they should clip the grids into autogrids (also called cartridges) before shipping. This will save sample loading time on the first day of the scheduled data collection session and thus the user gets more data collected instead.
Any projects where Janelia CryoEM staff prepare samples, grids, or do analysis of resulting images are considered to be collaborations. For collaboration work with the CryoEM facility that results in publication, CryoEM staff members should be listed as co-authors.
For data collection-only services, on user-provided ready-to-image cryo grids, if CryoEM staff enabled customization of approaches or made significant intellectual input or overcame extra difficulties (such as multiple loads of grids to find a usable one instead of a single load of up to 8 grids for one sample) to make the work possible, they should be listed as co-authors. Otherwise, the following acknowledgement should be included in the publication: 1, When describing data collection in the Materials and Methods or equivalent section, please mention that data was collected on a 300 kV FEI Titan Krios microscope located at the HHMI Janelia Research Campus. 2, In the Acknowledgement section of the paper, please acknowledge help from the Janelia CryoEM facility staff. Here is one possible example:
We thank Chuan Hong, Rick Huang and Zhiheng Yu at the HHMI Janelia CryoEM Facility for help in microscope operation and data collection.
After the paper is published, please send a link to the paper as well as one representative figure to the director of the Janelia CryoEM facility.