CryoFSL: An Annotation-Efficient, Few-Shot Learning Framework for Robust Protein Particle Picking in Cryo-EM Micrographs

Accurate identification of protein particles in cryo-electron microscopy (cryo-EM) micrographs is crucial for high-resolution structure determination, but remains challenging due to the heavy reliance on extensive annotated datasets and the difficulty of ensuring robustness under low signal-to-noise ratio (SNR) conditions. Current approaches require large annotations and exhibit poor generalization to new protein targets. We present CryoFSL ( Cryo -EM F ew S hot- L earning), a novel few-shot learning framework built upon Segment Anything Model 2 (SAM2) with lightweight adapters, enabling robust particle picking using as few as five labeled micrographs, significantly reducing annotation burden. The framework’s hierarchical adapter design supports dynamic feature modulation for low-SNR and heterogeneous conditions, resolving the trade-off between annotation burden and performance. CryoFSL surpasses both traditional template-based methods and state-of-the-art deep learning models across diverse proteins in the few-short learning setting, achieving superior recall, precision and 3D reconstruction resolution with minimal supervision. It maintains stability across heterogeneous micrographs and consistently detects high-quality particles with fewer false positives. Notably, CryoFSL achieves competitive density map reconstruction resolution with just a fraction of the particles picked by other methods, redefining efficiency and quality in cryo-EM analysis. This work paves the way for scalable, generalizable, and annotation-efficient particle picking pipelines. The code is available at GitHub .