With the rapid advancement of biomedical research, there is an increasing demand for imaging systems, such as high resolution, large field of view, and low phototoxicity. However, conventional single-modality optical microscopes often struggle to balance these requirements, limiting their applicability in complex biological studies.
This inherent limitation is evident in current mainstream modalities. For instance, among fluorescence-based methods, structured illumination microscopy (SIM) is widely utilized specifically due to its super-resolution capability and relatively low phototoxicity, making it highly suitable for live-cell imaging. Nevertheless, SIM faces several challenges in practice, including photodamage during long-term imaging, and a limited field of view. within the domain of label-free imaging, differential phase contrast (DPC) provides global structural information without the need for fluorescent labeling. It features low illumination intensity, reduced biological perturbation, and inherently captures a larger imaging field under identical optical conditions. These complementary characteristics make SIM and DPC ideal candidates for integration into a unified multimodal imaging platform.
In a new study published in Light: Advanced Manufacturing , Professor Peng Xi and collaborators report a highly integrated multimodal fluorescence-phase microscopy (MFPM) system. This system seamlessly combines a coherent structured illumination module for fluorescence excitation with a programmable partially coherent LED illumination module for label-free imaging into a unified wide-field detection architecture, achieving a remarkably compact 60 cm × 60 cm footprint. Crucially, by sharing a common detection pathway, this system inherently guarantees precise spatial co-registration across distinct modalities, which maximizes the data acquisition efficiency.
To further enhance imaging efficiency, the researchers utilized a computational framework incorporating a dark channel prior-based background removal method and a frame-reduction reconstruction strategy. As a result, the MFPM system requires only ten raw images to reconstruct five imaging modes, including optical-sectioning SIM (OS-SIM), super-resolution SIM (SR-SIM), polarization dipole analysis, fast DPC (fDPC), and quantitative DPC (qDPC). This approach substantially improves imaging throughput while expanding the dimensionality of biological information.
The researchers validated the system across diverse biological specimens.
At the cellular level, phase imaging provides global morphological information, while super-resolution fluorescence and polarization-resolved analysis reveal fine subcellular structures such as actin filaments and their anisotropy. Together, these modalities achieve a perfect complementation of overall morphology and microscopic details.
In pathological applications, the system enables quantitative analysis of cervical tissue by combining large-field DPC screening with fluorescence-based feature extraction, offering objective metrics such as fluorescence intensity distribution and nuclear density for assisted diagnosis.
Furthermore, in live model organisms such as zebrafish, the MFPM system enables multimodal cross-validation of cardiac activity, improving the accuracy of heart rate measurements. It also allows high spatiotemporal tracking of neutrophil migration within complex tissues, providing valuable insights into immune response dynamics.
The researchers emphasize that this multimodal integration strategy effectively overcomes the inherent limitations of single-modality imaging. By unifying structural and functional imaging capabilities within a single platform, MFPM offers a powerful tool for comprehensive biological investigations. Looking ahead, incorporating deep learning and refined physical models will further enhance the system's performance and automation. With strong scalability toward event-driven imaging and virtual staining, this integrated platform is poised to significantly advance drug discovery, pathological screening, and fundamental biomedical research.
Light: Advanced Manufacturing
Versatile biological imaging enabled by multimodal fluorescence phase microscopy