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A key obstacle arises from the trade\u2010off between staining efficiency and tissue autolysis. Traditional Golgi staining, typically conducted at room temperature or 37 \u00b0C to optimize staining efficiency, leads to rapid autolysis of brain tissue, resulting in the loss of fine structural details. Here, a near\u2010freezing temperature (NFT) staining strategy in post\u2010mortem frozen (PMF) human brain samples are presented, using a mercury chloride\u2010based method under ice\u2010water bath conditions. In contrast to the 37 \u00b0C Golgi staining, this NFT\u2010based method significantly reduces tissue autolysis, preserving fine neuronal structures. Notably, neuronal counts in the same field of view increased by 5.5\u2010fold, and dendritic spine density increases by 22\u2010fold. Using this approach, uniform staining of millimeter\u2010thick is achieved, centimeter\u2010scale human brain slices and integrated it with synchrotron\u2010based X\u2010ray microscopy to perform micrometer resolution 3D reconstructions of the cerebellum and frontal lobe. This novel technique offers a powerful tool for the fine\u2010structural imaging of large\u2010volume brain tissue, providing new insights into the intricate organization of neural networks.<\/jats:p>","DOI":"10.1002\/advs.202504468","type":"journal-article","created":{"date-parts":[[2025,5,28]],"date-time":"2025-05-28T08:21:31Z","timestamp":1748420491000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Near\u2010Freezing\u2010Temperature Golgi Neuronal Staining for X\u2010ray Imaging of Human Brain"],"prefix":"10.1002","volume":"12","author":[{"given":"Feng","family":"Zhou","sequence":"first","affiliation":[{"name":"CAS Key Laboratory of Interfacial Physics and Technology, Shanghai Institute of Applied Physics Chinese Academy of Sciences  Shanghai 201800 China"},{"name":"University of Chinese Academy of Sciences  Beijing 100049 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