Deep Learning for BioImaging: What Are We Really Learning?

Microscopy images contain rich information about how cells respond to perturbations, making them essential to applications like drug screening. To quantify images, researchers often use representation extraction methods, and recent years have seen a proliferation of deep learning methods. While measuring the quality of these representations is essential, evaluation remains fragmented, with each proposed model evaluated on different tasks and datasets, using custom pipelines and metrics, making it difficult to fairly compare models. Here, we introduce MorphoHELM, a comprehensive open benchmark for evaluating feature extraction methods for Cell Painting, the most widely-used morphological profiling assay. MorphoHELM consolidates evaluation standards in the field, extends and corrects them to be more robust, and evaluates on the widest range of methods to date. A defining feature of the benchmark is that each task is evaluated at different degrees of batch effects (or technical noise), directly quantifying how the ability of methods to detect biological signal degrades as noise increases. Together, these properties enable MorphoHELM to detect trade-offs between methods, and we demonstrate that models that excel at certain kinds of biological signal are weaker at others. We show that no existing model outperforms classic computer vision analytic strategies across all settings, which remain the strongest general use-case representations. All datasets, code, and evaluation tools are publicly available at https://github.com/microsoft/MorphoHELM.

Deep learning underlies most modern approaches and tools in computer vision, including biomedical imaging. However, for interactive semantic segmentation (often called pixel classification in this context) and interactive object-level classification (object classification), feature-based shallow learning remains widely used. This is due to the diversity of data in this domain, the lack of large pretraining datasets, and the need for computational and label efficiency. In contrast, state-of-the-art tools for many other vision tasks in microscopy - most notably cellular instance segmentation - already rely on deep learning and have recently benefited substantially from vision foundation models (VFMs), particularly SAM. Here, we investigate whether VFMs can also improve pixel and object classification compared to current approaches. To this end, we evaluate several VFMs, including general-purpose models (SAM, SAM2, SAM3, DINOv3) and domain-specific ones ($μ$SAM, PathoSAM, KRONOS), in combination with shallow learning and attentive probing on five diverse and challenging datasets. Our results demonstrate consistent improvements over hand-crafted features and provide a clear pathway toward practical improvements. Furthermore, our study establishes a benchmark for VFMs in microscopy and informs future developments in this area.

Single-cell light microscopy images have become an important data source for characterizing cell phenotypes, but their complexity and heterogeneity pose challenges to high-throughput automated analysis. Existing representation learning methods mostly rely on task-oriented modeling, which is limited by specific datasets and predefined tasks, making them difficult to generalize across different cell types and microscopy modalities, and experimental conditions. Although general-purpose methods have improved the generalization ability of image representation in recent years, their limited utilization of experimental background and biological context information still poses challenges in complex phenotypic analysis. Here, we propose scMIR, a vision-language foundation model for single-cell light microscopy image representation. By synergistically combining self-supervised image reconstruction with text-guided cross-modal alignment, scMIR can simultaneously encode morphological and biological semantic information in a unified representation space. scMIR is pre-trained on 207,957 image-text pairs, covering various cell types, microscopy modalities, and perturbation conditions. scMIR outperforms existing general models and task-oriented methods as systematically evaluated on various complex tasks using 16 benchmark datasets, including cell classification, clustering, phenotype inference, and batch effect correction tasks. Furthermore, scMIR shows a strong generalization ability across various tasks without requiring task-specific fine-tuning. With its unique advantages, we envision scMIR may promote the standardization and automation of high-throughput phenotyping workflows through supporting various downstream analysis tasks.