I am encountering a weird issue. I have two datasets of ~2 million particles each. Individually, they both refiine well (<3 Å). However, when I combine particles from both datasets, the resolution drops (> 4Å). In real and fourier space slices of reconstructions from combined particles, I also observe strange artefacts (see image attached). I have noticed that the maps reconstructed from dataset 2 are on a different absolute intensity scale when I open them in Chimera (Factor of 10 lower, so instead of the usual level of 0.5, the 2nd dataset is on a level of 0.05). I suspect that this might be the root cause of the issue.
Both datasets were from very similar grids collected on the same scope, mag and dose. For dataset 1, movies were pre-processed in Warp, the averaged micrographs were imported in cryosparc, CTF corrected, and particles exported using coordinates from Warp.
Dataset 2 was pre-processed with cryosparc live, then particles were picked in warp on the motioncorrected cryosparc live outputs, and finally particles were extracted in cryosparc using the CS live outputs and the warp coordinates.
Do the different pre-processing strategies explain this weird behaviour? is there a way to fix this without having to re-pre-process everything? Its ˜20 000 movies or each dataset…
Many thanks for your help,