CTF correction
Forums
Hi,
I'm having a strange problem with CTF correction during my Frealign runs. Without CTF correction (setting WGH to 1.0) the reconstruction looks perfectly fine but is at 25Å resolution -first 0. However the reconstruction becomes excessively smooth when I change WGH to 0.07 (the default value). I have done the CTF estimation with CTFFIND, and the defocus values reported by CTFFIND agree with my manual estimation using Bshow, so I don't believe that the estimated defocus values are incorrect. I use the same particle orientation parameters for creating a reconstruction with and without CTF correction (both done via Mode 0), so it's not a refinement issue. Am I making a fundamental mistake here?
I am not doing any FMAG/FDEF/FASTIG/FPART/IEWALD corrections since I'm so far from ~4.5Å. The only culprit that I can think of is a soft box mask that has been applied to my raw particles. Could this be the problem?
Thank you for the help.
The switch off CTF correction
The switch off CTF correction in FREALIGN, you need to set WGH to -1.0 (a negative value). The reason your reconstruction might look smooth when you set WGH back to 0.07 might have several reasons. If your reconstruction has a resolution of 25Å there will not be that much detail visible due to the limited resolution. If the reconstruction is even smoother than expected for 25Å, it might be due to a high B-factor. You should run the reconstruction through the BFACTOR program and see if you get more detail. Choose a B-factor of -3000 or thereabout and option 2 for the filter (cosine edge).