Tilt angle convention

Dynamo, IMOD and novaCTF, by default, all use the same convention for the sign of the tilt angle of an image within a tilt series: At positive tilt angles, the right side of the tilt images is expected to be more underfocused than the left side. It is important to ensure that the tilt angles of the corresponding tilt series follow this convention. This is to make sure that within those software packages the true handedness of the tomogram is not lost, the CTF corrections are done properly and missing wedge masks for incomplete tilt series are applied correctly.

To test if the convention is respected in your tilt series, the defocus on the left and right side of the tilt series can be estimated and plotted against the angles of the corresponding tilt angle file. The result should look more or less like this:

Plot of estimated defocus from full micrographs of a tilt series (blue), only left side of micrographs (red) and only right side of micrographs (yellow). If for the tilts 1-20 the left side has a higher defocus, the angles for these tilts should be negative. Analogously, the tilts 22-41 should have a positive angle if the right side has a higher defocus.

If this is not the case, the angles in the tilt file have to be adapted accordingly. To be completely certain, after correcting the signs of the angles, a rearrangement of the images in the stack could be made so that the first tilt image has the angle of -60 degrees.