Tutorial: Manually Curate Exposures (≤v3.3)
Manually Curate Exposures is an interactive job in CryoSPARC that enables a user to visually inspect and curate a set of micrographs or movies. This tutorial refers to CryoSPARC v3.3 and earlier.
Manually Curate Exposuresis an interactive job in CryoSPARC that enables a user to visually inspect and curate a set of micrographs or movies, using either population-level statistics and adjustable thresholds, or individual inspection of diagnostic plots.
This interactive job allows you to:
- Load a stack of (i) micrographs, (ii) movies or (iii) micrographs and particles
- View and sort the stack by certain metrics associated with the data you loaded, e.g., average defocus, astigmatism, CTF fit, motion trajectories and power scores relating to picked particles
- Create and visualize interactive plots to view the distribution of micrographs/movies over different parameters, and identify outliers
- Adjust thresholds to bulk-exclude micrographs based on the statistic of interest
- View available CTF diagram (to inspect Thon rings), CTF fit, and rigid and local motion trajectory plots associated with an individual micrograph, all in one place
- Accept or reject micrographs/movies from a stack, for further processing
The overall goal of this interactive job is to aid in cleaning up your data so that low-quality micrographs, and their associated particles, do not detract from achieving the highest possible resolution refined structures.
After pre-processing: After
Patch Motion Correctionand
Patch CTF estimation, you can use
Manually Curate Exposuresto help you identify and exclude micrographs with sub-optimal characteristics, e.g., broken or thick ice, carbon edges, too much motion, etc.
After particle picking and
local motion correction: Once you have picked particles and estimated location motion trajectories, you can use this information in
Manually Curate Exposuresto identify and exclude micrographs associated with those picks whose local motion trajectories deviate from the norm for your dataset. In this case, the job will output a filtered set of micrographs as well as particles.
Manually Curate Exposuresfrom the Job Builder:
- Drag and drop the desired inputs from previous jobs. You can connect (a) micrographs only, (b) movies only, or (c) micrographs/movies and particles.
- There are no parameters to adjust for this interactive job, so simply click
Queueto launch the job.
Overview and Individual mode: The default view is
Overview. An explanation of the
Individualview can be found below.
Table of exposures: On the left, exposures are listed in a table with index values, and summary statistics are available at the top of the table. In this case, we are looking at a total of 1536 micrographs.
Selected: At the start of the job, all loaded micrographs are 'Selected', as indicated at the top of the table and their values in blue in the table. Once we begin adjusting the interactive plot and thresholds, micrographs whose statistics do not fall within the selected thresholds will become un-selected, and their values will turn grey in the table.
Available columns: A number of columns are displayed depending on the type of input. For example, because we loaded micrographs and particles with local motion trajectories, we see columns displaying not only those statistics associated with micrographs (e.g., average defocus, astigmatism, CTF fit, etc.) but also those corresponding to the picks and local motion trajectories (pick CC, pick power, local motion distribution, etc.) Click on a statistic to sort the table.
Create plots: The remainder of the window is dedicated to an interactive plot. All micrographs in the table are displayed in the plot. The
Y-Axisdrop-down menus directly below allow you to select different statistics by which to plot the micrographs. Selecting a different value from the drop-down will automatically refresh the plot. Common plots include:
Interact with plots: Hover over the plot area and a toolbar will appear at the top. These tools allow you to: download the plot as a PNG file, zoom, pan, and reset the axes. Note: Selections in the plot currently do NOT affect selection or acception/rejection of micrographs. Use the threshold sliders below the plot to make a selection and then the buttons above the plot to accept/reject/invert your selection.
Adjust thresholds to select micrographs: Once you have explored the distribution of micrographs on one or more plots and have identified outliers, you can adjust the various thresholds below the plot to filter the micrographs. The corresponding values in the table will turn grey. (Note: The plot does not currently reflect your selection - all micrographs will be shown on the plot.) For example, here we plotted CTF fit resolution (A) on both the x-axis and y-axis, and then adjusted the CTF fit resolution (A) slider to keep only those micrographs with a value of less than 8.48 A. In the table, 3 micrographs are greyed out, indicating they do not fall within the threshold values.
Accept selection: To keep all micrographs with CTF values falling within the desired range, click 'Accept Selection' at the top of the plot. Corresponding micrographs will then be highlighted green in the table.
Reject selection: Alternatively, you can reject micrographs with statistics falling within a certain range, following the same process as above, but click 'Reject selection' above the plot. Rejected micrographs will turn red in the table.
To switch to `Individual' mode, click on the button at the top, or by clicking on an individual row in the table:
Select a row to view details for that micrograph:
You can then view CTF plots, motion trajectories and other available plots corresponding to the selected exposure. It is also possible to move to the next exposure in the table (using the left/right arrows), as well as Accept or Reject the individual micrograph from this view. You can also click on any individual plot to enlarge it.
Done, the job will finish running and a list of outputs will become available. These can be used for further processing as required.
Common next steps include:
2D Classificationjob, and/or starting an
Ab-Initio Reconstructionusing only these particles, followed by
micrographs_acceptedto continue processing particle picking and local motion correction using only the good data