I am working with the filament structure which spans through the box but refinement is done in C1 symmetry. Because the density is not focused in the middle it makes problems with the local refinement (assertion error). I discovered that when the Window inner radius and Window outer radius are changed to ~0.6-0.7 the error does not occur and refinement works well. However, I noticed that the rest of the box outside the Window outer radius is not masked with 0 but there is still visible density. Is there a way to mask the volume close to the box boundaries like it is done in Relion with circular mask? Thanks!
The window inner radius and window outer radius in a refinement job control windowing that is applied the the particle images rather than the volume. So in your case setting them to 0.6/0.7 windows the particle images before using them for alignment/reconstruction. The 3D density however is always windowed by a soft spherical mask that just cuts out the corners of the box, along with a shaped mask (by default, based on dynamic masking). In the final refinement output, however, these masks are not applied - you are seeing the full result of reconstruction from the particle images directly. The masks are only intended to have an effect on alignment. So even though you see density outside of the window that you specified, this is normal and not a problem - since that density is not used for alignments.
Hope that helps!
This is great, thank you for the answer. However, this does not explain why during the NU-refinement there is an an assertion error when the inner and outer radius are set to default values. Do you have any idea why this happens? If you need more information I would be happy to provide them.
EDIT:
This error exactly:
Traceback (most recent call last):
File “cryosparc2_worker/cryosparc2_compute/run.py”, line 78, in cryosparc2_compute.run.main
File “cryosparc2_worker/cryosparc2_compute/jobs/nonuniform_refine/run.py”, line 645, in cryosparc2_compute.jobs.nonuniform_refine.run.run_non_uni_refine
AssertionError
Sorry for the late reply. No, I am using the dynamic masking for this job. I also tried with mask which was created in previous job but the error is persistent.
Hi @apunjani - sorry to drag this topic back from the dead - but is the soft windowing applied before or after CTF correction of the particle images? I am assuming after?
To your first question, the soft windowing on the particle images is actually applied before the CTF is used. That meaning, particles are windowed at the start, then are used for alignment/reconstruction along with the previously estimated CTF values.
For your second question, windowing is applied relative to the original center of the extracted particles, rather than the fulcrum. Right now, local refinement doesn’t recenter particles on the fulcrum, instead it modifies the shifts to achieve the same effect (as doing rotations around the fulcrum).