Pursuing the physics of cryo-EM image formation

Cryo-electron microscopy (cryo-EM) has revolutionized structural biology, enabling near-atomic resolution determination of biological macromolecules. The success of modern reconstruction algorithms relies heavily on simplified image formation models, particularly the projection and weak-phase approximations, which linearize the relationship between the measured images and the specimen's three-dimensional (3D) scattering potential - commonly referred to as the map in cryo-EM. While these approximations have proven remarkably effective, the underlying physics of electron imaging is considerably richer, more complex, and arguably under-exploited. This chapter revisits the foundational principles of image formation in cryo-EM from first principles, tracing the physical pathway from electron-matter interaction through wave propagation to final image detection. Here, we show how the contrast transfer function (CTF) model commonly used in cryo-EM arises directly from the underlying image formation model, under standard approximations. Finally, we connect these physical models to reconstruction by introducing our own reconstruction algorithm, Ghostbuster, which leverages a physically accurate forward model to go beyond conventional CTF-based approaches. By grounding contemporary cryo-EM practice in rigorous physical principles, this chapter aims to provide essential context for assessing the validity of standard assumptions and guiding future methodological developments as the field continues to push toward its ultimate resolution limits.