When collecting data I try to balance the number of particles collected per session with the expected maximum resolution of reconstruction based on the magnification used. I have relied on the simple assumption that if the local resolution of my map is not Nyquist-limited, then my data were collected at a high enough magnification.

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Other than these things, the next main limit is just the total SNR based on the number of particles you have and the information (SNR) in each one. If you can perfectly align, have perfect CTFs, and a rigid particle, then you just need to collect enough images to reach any resolution you desire (theoretically). At that point (which pretty much only simple samples like apoferritin can reach) radiation damage to the protein during exposure is the main limitation.

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This seems like a good forum for asking for advice in this area: Am I oversimplifying things by only considering the Nyquist limit? I have noted that the DQE for a given detector decreases as the resolution approaches Nyquist, but I’m unsure if the extent of variation is consequential in terms of a final reconstruction. I am also wondering if it is easier to align small particles with a lower magnification (and subsequently higher apparent contrast).

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The final reconstruction resolution is limited by several factors, with the hardest limit being the Nyquist resolution chosen by setting the magnification. However, long before the Nyquist resolution the following things can limit:

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The focal length refers to the distance between the optical center of the lens and the image sensor.

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