title: Optimized Collection Protocol for Plasma MicroRNA Measurement in Patients with Cardiovascular Disease.
abstract: Background. Various microRNAs (miRNAs) are used as markers of acute coronary syndrome, in which heparinization is considered mandatory therapy. Nevertheless, a standard method of handling plasma samples has not been proposed, and the effects of heparin treatment on miRNA detection are rarely discussed.Materials and Method. This study used quantitative polymerase chain reaction (qPCR) analysis to investigate how storage temperature, standby time, hemolysis, and heparin treatment affect miRNA measurement in plasma samples from 25 patients undergoing cardiac catheterization.Results. For most miRNAs, the qPCR results remained consistent during the first 2 hours. The miRNA signals did not significantly differ between samples stored at 4°C before processing and samples stored at room temperature (RT) before processing. miR-451a/miR-23a ratio < 60 indicated < 0.12% hemolysis with 100% sensitivity and 100% specificity. Pretreatment with 0.25 U heparinase I recovered qPCR signals that were reduced byin vivoheparinization.Conclusions. For miRNA measurement, blood samples stored at RT should be processed into plasma within 2 hours after withdrawal and should be pretreated with 0.25 U heparinase I to overcome heparin-attenuated miRNA signals. The miR-451a/miR-23a ratio is a reliable indicator of significant hemolysis.
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan.
authors.forename: Chi-Sheng
authors.initials: CS
authors.lastname: Wu
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan; Division of Cardiology, Department of Internal Medicine, Linkou Chang Gung Memorial Hospital, Tao-Yuan, Taiwan.
authors.forename: Fen-Chiung
authors.initials: FC
authors.lastname: Lin
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan.
authors.forename: Shu-Jen
authors.initials: SJ
authors.lastname: Chen
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan; Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
authors.forename: Yung-Lung
authors.initials: YL
authors.lastname: Chen
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan; Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
authors.forename: Wen-Jung
authors.initials: WJ
authors.lastname: Chung
authors.affiliation: Molecular Medicine Research Center, Chang Gung University School of Medicine, Tao-Yuan, Taiwan; Division of Cardiology, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan.
authors.forename: Cheng-I
authors.identifier: 0000-0002-3134-2447
authors.initials: CI
authors.lastname: Cheng
mesh_terms: D000328:Adult; D000368:Aged; D001800:Blood Specimen Collection / Q000379:methods*; D002318:Cardiovascular Diseases / Q000097:blood* / Q000235:genetics*; D017931:DNA Primers / Q000378:metabolism; D015342:DNA Probes / Q000378:metabolism; D003710:Demography; D005260:Female; D005786:Gene Expression Regulation; D006461:Hemolysis; D019763:Heparin Lyase / Q000378:metabolism; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000097:blood* / Q000235:genetics*; D008875:Middle Aged; D016133:Polymerase Chain Reaction; D011309:Preservation, Biological; D013696:Temperature
chemical_list: D017931:DNA Primers; D015342:DNA Probes; D035683:MicroRNAs; D019763:Heparin Lyase