Development and Application of a High-Resolution Cryogenic STXM System at the Shanghai Synchrotron Radiation Facility

Scanning transmission X-ray microscopy (STXM) is an advanced technique for label-free chemical imaging with nanometer-scale spatial resolution which is well suited to studies of complex and heterogeneous materials. For biological and other radiation-sensitive specimens, however, the X-ray dose required for high-quality imaging often exceeds the damage threshold of the sample at room temperature. The resulting mass loss, chemical modification, and structural collapse corrupt the very signals under investigation. Cryogenic sample conditions can mitigate these effects by suppressing radiation-induced diffusion and secondary reactions. Yet their practical implementation is challenging. An instrument must simultaneously maintain stable low-temperature operation, isolate vibrations, and transfer samples without contamination under high vacuum. Satisfying all of these demands at once is difficult. Herein, we report the development and commissioning of a dedicated cryogenic STXM system installed on the beamline BL08U1A of the Shanghai Synchrotron Radiation Facility (SSRF). The instrument combines a mechanically decoupled cryostat architecture with a closed-loop ultra-high vacuum cryo-transfer system. It achieves a spatial resolution better than 30 nm and a long-term thermal drift below 0.05 nm/min. Validation on poly(methyl methacrylate) (PMMA) films shows that imaging at 120 K extends the critical radiation dose by a factor of ∼39 times compared with room temperature. Mass loss and secondary radical reactions are effectively suppressed. In situ imaging of frozen-hydrated mouse embryonic fibroblasts further resolves subcellular features such as filopodia, free from dehydration artifacts. Together, these results establish a robust platform for nondestructive, high-resolution chemical imaging of radiation-sensitive materials under near-physiological conditions, providing a foundation for correlative and multimodal studies of hydrated biological systems.