{"doi":"10.1101/2025.09.30.679660","title":"Segmentation-guided photon pooling enables robust single cell analysis and fast fluorescence lifetime imaging microscopy","abstract":"Fluorescence lifetime imaging microscopy (FLIM) can probe the metabolic environment of living cells in a label-free and non-invasive manner. However, endogenous fluorophores have low absorption and quantum yields, which necessitates long integration times to acquire the high photon counts needed for accurate pixel-wise multi-exponential decay fitting. Here, we present a 'region-of-interest' photon pooling technique to expedite label-free, single cell FLIM acquisition and analysis. As a result, we achieved single-cell metabolic information at intervals as low as one second and acquired large FLIM mosaics 15 times faster than would be possible with conventional pixel-level analysis. This technique is computationally light, does not require machine learning algorithms, and has been integrated with commonly used analysis software and file types.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":559198,"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":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.952,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":407597,"name":"Danielle E. Desa","orcid":"0000-0001-8975-0526","position":1,"is_corresponding":false},{"id":1460414,"name":"Xiaotian Zhang","orcid":"0000-0001-8601-4160","position":2,"is_corresponding":false},{"id":1328448,"name":"Dan L. Pham","orcid":"0000-0003-1901-2987","position":3,"is_corresponding":false},{"id":108078,"name":"Rupsa Datta","orcid":"0000-0002-6432-2389","position":4,"is_corresponding":false},{"id":108082,"name":"Melissa C. Skala","orcid":"0000-0002-6320-7637","position":5,"is_corresponding":false},{"id":744913,"name":"Kayvan Samimi","orcid":"0000-0002-2566-612X","position":0,"is_corresponding":true}],"reference_count":43,"raw_metadata":null,"created_at":"2026-07-19T02:55:34.849815Z","pmid":"41256575","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":[]}