Counting particles in cryo-electron microscopy may result in incorrect population estimates

Counting particles is a common practice in estimating populations from cryo-electron microscopy data. Here, it is shown that counting particles is not robust to noise, whereas statistically rigorous population-estimation methods are more resilient and hold promise for future development. This comment delves into the need for statistically rigorous population estimation in cryo-electron microscopy. Examples are given illustrating the failure of particle counting and the robustness of principled techniques.