Pretraining and Distillation Matter More Than Architecture Family for Label-Free Single-Cell Classification
Choosing a deep learning architecture for label-free single-cell classification remains an open question, with microscopy benchmarks reporting conflicting conclusions about CNNs versus transformers. We present a controlled benchmark on LIVECell phase-contrast microscopy data using source-image-disjoint train/validation/test splits to prevent parent-image leakage and matched optimisation, augmentation, and evaluation protocols across EfficientNet, Vision Transformer (ViT), and EVA-02 models. This allows the effects of architecture, pretraining, fine-tuning, tokenisation, and distillation to be disentangled. We find that the previously reported CNN advantage is largely explained by pretraining rather than architecture: the smallest pretrained model outperforms the strongest model trained from scratch despite far fewer parameters. Pretraining improves macro-F1 by 3-4 points, while the gap between the best pretrained CNN and transformer is below 0.5 points. Architectural choices nevertheless matter: ViT-S/8 outperforms ViT-S/16 and matches the four-times-larger ViT-B/16 at a quarter of the parameters, showing that finer tokenisation benefits small cell crops. Conversely, layer-wise learning-rate decay, central to the EVA-02 fine-tuning recipe, degrades performance, highlighting that transfer heuristics from natural-image recognition may not generalise to microscopy. Finally, knowledge distillation substantially improves the deployment frontier: compact EfficientNet-B0 students distilled from teacher councils outperform every individually trained backbone, including the EfficientNet-B5 and EVA-02 teachers. Overall, our results show that rigorous control of pretraining and evaluation is essential for interpreting biomedical architecture benchmarks, while distillation may be a more effective route to practical single-cell classification than architecture choice alone.