Hello, everyone! I am dealing with data collected on K3 camera. I want to throw away 2 starting frames and 3 frames at the end during motion correction. Should I enter 2 in the Start frame (included, 0-based) option and 3 in the End frame (excluded, 0-based) option? BTW, I was told that the total dose is 50 e/Å^2 and each movie has 6 dose fractions containing 32 frames each. Is it reasonable to throw this number of frames or any good suggestion ?
Why do you want to throw away frames at this point? I would advise keeping all frames as a starting point and letting dose weighting take care of the later frames. If the first frame or two look particularly bad after patch motion you might consider discarding them, but I would not do so by default.
Hi @Verdandy! Thanks for your post!
As @olibclarke said, discarding frames is an older workflow that is less common now that we perform patch motion correction and dose weighting.
In the olden days, we discarded early frames because of the relatively high degree of anisotropic motion in those frames (which blurs particles) and we discarded late frames due to the high degree of radiation damage (which destroys high- and even medium-resolution information). Let’s approach both of those problems separately.
As I said, those early frames have a large degree of anisotropic motion. That is to say, the images might be expanding or contracting in different parts of the frame, or moving apart from each other. When we aligned the whole frame as a single unit that rendered these unusable, but now that we use patch motion correction those early frames are usable again. In fact, since they have received very little radiation damage, they’re quite valuable!
The later frames have been bombarded by electrons for a long time, and so have a good deal of radiation damage. We used to discard these too, but in this paper Grant and Grigorieff suggested instead that we use what we know about these images to get as much information as we can out of them.
Early in the movie the frames have a good degree of information at all resolutions, while later in the movie the high-res information has been destroyed, but the low-res info is still there! So these frames are “dose-weighted” (exposure filtered in that paper). Later frames have their high-resolution signal “turned down” so that they only contribute their good low-resolution information, but not their high-resolution noise.
I hope that helps explain why we don’t really need to discard frames anymore for a normal workflow! There are still times it might be useful to only process a certain number of frames (for instance, you might want to know which parts of your protein are damaged by radiation). In that case, remember that you’re still counting from the beginning of the movie! If you put 2 for
Start Frame and 3 for
End Frame you’d only use a single frame, frame 2