{"doi":"10.1364/oe.570174","title":"Tilted thick light-sheet microscopy with context-aware 3D localization for RNA imaging in intact 50 μm-thick sections","abstract":"<jats:p>\n                    Neuron typing in intact brain sections demands 3D imaging reconciling thickness compatibility with large-area coverage, which is a persistent challenge for RNA imaging methods limited by either imaging area (&lt; 1 mm\n                    <jats:sup>2</jats:sup>\n                    ) or section thickness (&lt; 20 μm). Here we introduce tilted thick light-sheet microscopy integrated with a context-aware 3D astigmatic localization algorithm based on continuous frame filtering, enabling multiplexed RNA imaging across 50 μm-thick tissue sections and centimeter-scale regions. The 45° illumination geometry achieves optical-interference-free imaging in a 50 μm-thick sample, while a context-aware 3D localization algorithm recovers submicron resolution by pinpointing fluorescence in situ hybridization (FISH) spots within volumetric excitation. This strategy could reveal layer-spanning neuronal distributions and region-specific expression gradients unattainable with conventional RNA imaging approaches. By resolving the trade-off between imaging depth and spatial coverage, our platform advances whole-brain transcriptomics and has the potential to carry out 3D neuron typing, which is critical for deciphering brain-wide cellular architectures.\n                  </jats:p>","journal":"Optics Express","year":2025,"id":634335,"datarank":0.10397207708399181,"base_score":0.6931471805599453,"endowment":0.6931471805599453,"self_citation_contribution":0.10397207708399181,"citation_network_contribution":0.0,"self_endowment_contribution":0.10397207708399181,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":null,"is_data_producer":false,"deposit_databanks":null,"is_oa":false,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":null,"fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1645106,"name":"Qihang Song","orcid":"0000-0001-9345-0913","position":1,"is_corresponding":false},{"id":1645107,"name":"Dongjian Cao","orcid":null,"position":2,"is_corresponding":false},{"id":1645108,"name":"Qihuan Li","orcid":null,"position":3,"is_corresponding":false},{"id":1645109,"name":"Wancheng Wu","orcid":null,"position":4,"is_corresponding":false},{"id":1645110,"name":"Yinghao Xiao","orcid":null,"position":5,"is_corresponding":false},{"id":1645111,"name":"Xingguang Chen","orcid":null,"position":6,"is_corresponding":false},{"id":1645112,"name":"Shujing Huang","orcid":null,"position":7,"is_corresponding":false},{"id":1415080,"name":"Jie Yang","orcid":"0000-0002-6156-2694","position":8,"is_corresponding":false},{"id":1645113,"name":"Yingjun Zhang","orcid":"0000-0001-5703-3610","position":9,"is_corresponding":false},{"id":1645114,"name":"Zhen-Li Huang","orcid":"0000-0003-2400-966X","position":10,"is_corresponding":false},{"id":746429,"name":"Lin Cai","orcid":"0000-0002-2757-7809","position":0,"is_corresponding":false}],"reference_count":0,"raw_metadata":{"has_enrichment":true,"resolved":true,"title":"Tilted thick light-sheet microscopy with context-aware 3D localization for RNA imaging in intact 50 μm-thick sections","abstract":"<jats:p>\n                    Neuron typing in intact brain sections demands 3D imaging reconciling thickness compatibility with large-area coverage, which is a persistent challenge for RNA imaging methods limited by either imaging area (&lt; 1 mm\n                    <jats:sup>2</jats:sup>\n                    ) or section thickness (&lt; 20 μm). Here we introduce tilted thick light-sheet microscopy integrated with a context-aware 3D astigmatic localization algorithm based on continuous frame filtering, enabling multiplexed RNA imaging across 50 μm-thick tissue sections and centimeter-scale regions. The 45° illumination geometry achieves optical-interference-free imaging in a 50 μm-thick sample, while a context-aware 3D localization algorithm recovers submicron resolution by pinpointing fluorescence in situ hybridization (FISH) spots within volumetric excitation. This strategy could reveal layer-spanning neuronal distributions and region-specific expression gradients unattainable with conventional RNA imaging approaches. By resolving the trade-off between imaging depth and spatial coverage, our platform advances whole-brain transcriptomics and has the potential to carry out 3D neuron typing, which is critical for deciphering brain-wide cellular architectures.\n                  </jats:p>","is_dataset_classified":null,"base_score":0.6931471805599453,"endowment":0.6931471805599453,"datacite_reuse_total":0,"file_count":0,"downloads":0,"views":0,"has_version_chain":false,"is_dataset":false,"is_oa":false,"pmid":"40734022","pmcid":null,"openalex_id":"https://openalex.org/W4412403483","authors":[],"funders":[{"funder_name":"National Natural Science Foundation of China","grant_id":"81827901","title":null},{"funder_name":"National Natural Science Foundation of China","grant_id":"82260368","title":null},{"funder_name":"National Key Research and Development Program of China","grant_id":"2022YFC3400601","title":null},{"funder_name":"Innovational Fund for Scientific and Technological Personnel of Hainan Province","grant_id":"KJRC2023C03","title":null},{"funder_name":"Start-up Fund from Hainan University","grant_id":"KYQD(ZR)-20077","title":null}],"total_grants":5,"fwci":1.0095,"citation_percentile":0.80923975,"influential_citations":0,"citation_trend":[{"year":2025,"count":1}],"oa_status":"gold","license":"https://doi.org/10.1364/OA_License_v2#VOR-OA","oa_locations":[{"url":"https://doi.org/10.1364/oe.570174","host_type":"journal"},{"url":"https://doi.org/10.1364/oe.570174","host_type":"publisher"},{"url":"https://opg.optica.org/viewmedia.cfm?URI=oe-33-15-32850&seq=0","host_type":"publisher"},{"url":"https://pubmed.ncbi.nlm.nih.gov/40734022","host_type":"repository"}],"fields_of_study":["Advanced Fluorescence Microscopy Techniques","Cell Image Analysis Techniques","Image Processing Techniques and Applications","Imaging, Three-Dimensional","Animals","Brain","Algorithms","RNA","In Situ Hybridization, Fluorescence","Neurons","Mice"],"mesh_terms":["Algorithms","Animals","Brain","Neurons","RNA","In Situ Hybridization, Fluorescence","Imaging, Three-Dimensional","Mice"],"keywords":["Optics","Microscopy","Light sheet fluorescence microscopy","Materials science","Context (archaeology)","Optical microscope","Optical coherence tomography","Scanning confocal electron microscopy","Scanning electron microscope","Physics","Biology"],"sdg_mappings":[],"linked_datasets":[],"clinical_trials":[],"software_tools":[],"database_accessions":[],"source":"live","citation_network_status":"fetched"},"created_at":"2026-08-06T13:29:31.356333Z","pmid":null,"pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}