{"doi":"10.1111/cyt.13304","title":"Pre‐analytical effects on whole transcriptome and targeted RNA sequencing analysis in cytology: The effects of prolonged time in storage of effusion specimens prior to preservation","abstract":"OBJECTIVES: To investigate the pre-analytics of the molecular testing of cytology specimens, we studied the effects of time in refrigerator storage (4°C) of malignant effusions on RNA sequencing (RNAseq) results. METHODS: Ten effusion specimens were stored in a refrigerator (4°C) for different durations (day 0, 1, 4, and 7). All specimens were prepared as cytospins fixed in either Carnoy's solution or 95% ethanol (EtOH) and in an RNA preservative for a fresh frozen (FF) high-quality reference. Whole transcriptome (wt) and targeted (t)RNAseq of two multigene expression signatures were performed. We then compared transcript expression levels (including mutant allele fraction) according to pre-analytical variables using a concordance correlation coefficient (CCC) and a mixed effect model. RESULTS: Sequencing results were mostly stable over increasing time in storage. Cytospins fixed in Carnoy's solution were more concordant with FF samples than cytospins fixed in 95% EtOH at all timepoints. This finding was consistent for both wtRNAseq (averages: day 0 CCC = 0.98 vs 0.91; day 7 CCC = 0.88 vs 0.78) and tRNAseq methods (averages: day 0 CCC = 0.98 vs 0.81; day 7 CCC = 0.98 vs 0.90). Cytospins fixed in Carnoy's solution did not show significant changes in expression over timepoints or between expression signatures, whereas 95% EtOH did. CONCLUSION: RNAseq can be accurately performed on effusion specimens after prolonged refrigerator storage. RNA extracted from scraped cytospin slides fixed in Carnoy's solution was marginally superior to 95% EtOH fixation, but either method had comparable analytic performance to high-quality FF RNA samples.","journal":"Cytopathology","year":2023,"id":355842,"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":5,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.8368,"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":673642,"name":"Kevin Tran","orcid":"0000-0002-8162-4477","position":1,"is_corresponding":false},{"id":762212,"name":"Chunxiao Fu","orcid":"0009-0000-6950-5355","position":2,"is_corresponding":false},{"id":619673,"name":"Lili Du","orcid":null,"position":3,"is_corresponding":false},{"id":98977,"name":"Michal Marczyk","orcid":"0000-0003-2508-5736","position":4,"is_corresponding":false},{"id":680237,"name":"Rebekah Gould","orcid":"0000-0001-7391-627X","position":5,"is_corresponding":false},{"id":1104199,"name":"Eveline Chen","orcid":"0000-0001-8967-3890","position":6,"is_corresponding":false},{"id":1104627,"name":"Amy Tasto","orcid":null,"position":7,"is_corresponding":false},{"id":881201,"name":"Agata A. Tinnirello","orcid":"0000-0002-2172-7263","position":8,"is_corresponding":false},{"id":242395,"name":"W. Fraser Symmans","orcid":"0000-0002-1526-184X","position":9,"is_corresponding":false},{"id":881199,"name":"Gloria H. Sura","orcid":"0000-0003-4817-458X","position":0,"is_corresponding":true}],"reference_count":21,"raw_metadata":null,"created_at":"2026-07-19T01:13:21.145955Z","pmid":"37712171","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":[]}