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Dear Relion community, when doing particle alignment in 3D in Relion, does the expectation step take into account previous information from 2D classification? E.g. if 10,000 particles went into a single class in 2D, the likelihood for these 10,000 particles to be in a more or less similar orientation in 3D should be high, right? At least the psi angle for each of these particles should be the same / change the same way. Thank you for helping me understand better. If the answer to this question is yes, then it would always be useful to do a 2D classification before starting a 3D classification, I guess. Best, Thomas ######################################################################## To unsubscribe from the CCPEM list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCPEM&A=1