{"doi":"10.1101/2023.11.07.566086","title":"Widefield in vivo imaging system with two fluorescence and two reflectance channels, a single sCMOS detector, and shielded illumination","abstract":"SIGNIFICANCE: Widefield microscopy of the entire dorsal part of mouse cerebral cortex enables large-scale (mesoscopic) imaging of neuronal activity with fluorescent indicators as well as hemodynamics via oxy- and deoxyhemoglobin absorption. Versatile and cost-effective imaging systems are needed for large-scale, color-multiplexed imaging of multiple fluorescent and intrinsic contrasts. AIM: Develop a system for mesoscopic imaging of two fluorescent and two reflectance channels. APPROACH: Excitation of red and green fluorescence is achieved through epi-illumination. Hemoglobin absorption imaging is achieved using 525- and 625nm LEDs positioned around the objective lens. An aluminum hemisphere placed between objective and cranial window provides diffuse illumination of the brain. Signals are recorded sequentially by a single sCMOS detector. RESULTS: We demonstrate performance of our imaging system by recording large-scale spontaneous and stimulus-evoked neuronal, cholinergic, and hemodynamic activity in awake head-fixed mice with a curved crystal skull window expressing the red calcium indicator jRGECO1a and the green acetylcholine sensor GRABACh3.0 . Shielding of illumination light through the aluminum hemisphere enables concurrent recording of pupil diameter changes. CONCLUSIONS: Our widefield microscope design with single camera can be used to acquire multiple aspects of brain physiology and is compatible with behavioral readouts of pupil diameter.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2023,"id":397699,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":2,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9523,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":776401,"name":"Natalie Fomin-Thunemann","orcid":"0000-0003-0449-4645","position":1,"is_corresponding":false},{"id":714257,"name":"Rockwell P. Tang","orcid":"0000-0003-3044-0085","position":2,"is_corresponding":false},{"id":1173900,"name":"Dora Balog","orcid":null,"position":3,"is_corresponding":false},{"id":1173901,"name":"Bernhard Zimmerman","orcid":null,"position":4,"is_corresponding":false},{"id":256592,"name":"Kıvılcım Kılıç","orcid":"0000-0001-7248-8728","position":5,"is_corresponding":false},{"id":896129,"name":"Emily A. Martin","orcid":"0000-0001-5785-9105","position":6,"is_corresponding":false},{"id":328302,"name":"Sreekanth Kura","orcid":"0000-0002-8071-3259","position":7,"is_corresponding":false},{"id":266732,"name":"Harrison Fisher","orcid":"0000-0002-6800-3107","position":8,"is_corresponding":false},{"id":1173902,"name":"Grace Chabbott","orcid":null,"position":9,"is_corresponding":false},{"id":1173903,"name":"Joel Herbert","orcid":null,"position":10,"is_corresponding":false},{"id":932998,"name":"Bradley C. Rauscher","orcid":"0009-0008-1737-059X","position":11,"is_corresponding":false},{"id":296763,"name":"John Jiang","orcid":null,"position":12,"is_corresponding":false},{"id":374501,"name":"Sava Sakadžić","orcid":"0000-0001-6318-1193","position":13,"is_corresponding":false},{"id":256593,"name":"David A. Boas","orcid":"0000-0002-6709-7711","position":14,"is_corresponding":false},{"id":246429,"name":"Anna Devor","orcid":"0000-0002-5143-3960","position":15,"is_corresponding":false},{"id":328304,"name":"Anderson Chen","orcid":"0000-0001-6110-5285","position":16,"is_corresponding":false},{"id":524202,"name":"Martin Thunemann","orcid":"0000-0003-4139-079X","position":17,"is_corresponding":false},{"id":776402,"name":"Patrick Doran","orcid":"0000-0002-0374-0376","position":0,"is_corresponding":true}],"reference_count":37,"raw_metadata":null,"created_at":"2026-07-19T01:19:39.497368Z","pmid":"37986755","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":[]}