Estefanía Lozano-Andrés PhD , Tina Van Den Broeck PhD , Lili Wang PhD , Majid Mehrpouyan PhD , Ye Tian PhD , Xiaomei Yan PhD , Ger J.A. Arkesteijn PhD , Marca H.M. Wauben PhD
{"title":"荧光校准器的内在可变性影响了流式细胞术对纳米颗粒和细胞外囊泡的MESF或ERF值的分配。","authors":"Estefanía Lozano-Andrés PhD , Tina Van Den Broeck PhD , Lili Wang PhD , Majid Mehrpouyan PhD , Ye Tian PhD , Xiaomei Yan PhD , Ger J.A. Arkesteijn PhD , Marca H.M. Wauben PhD","doi":"10.1016/j.nano.2023.102720","DOIUrl":null,"url":null,"abstract":"<div><p>Flow cytometry allows to characterize nanoparticles (NPs) and extracellular vesicles (EVs) but results are often expressed in arbitrary units of fluorescence. We evaluated the precision and accuracy of molecules of equivalent soluble fluorophores (MESF) beads for calibration of NPs and EVs. Firstly, two FITC-MESF bead sets, 2 and 6 um in size, were measured on three flow cytometers. We showed that arbitrary units could not be compared between instruments but after calibration, comparable FITC MESF units were achieved. However, the two calibration bead sets displayed varying slopes that were consistent across platforms.</p><p>Further investigation revealed that the intrinsic uncertainty related to the MESF beads impacts the robust assignment of values to NPs and EVs based on extrapolation into the dim fluorescence range. Similar variations were found with PE MESF calibration.</p><p>Therefore, the same calibration materials and numbers of calibration points should be used for reliable comparison of submicron sized particles.</p></div>","PeriodicalId":19050,"journal":{"name":"Nanomedicine : nanotechnology, biology, and medicine","volume":null,"pages":null},"PeriodicalIF":4.2000,"publicationDate":"2023-11-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S1549963423000710/pdfft?md5=7ce6e8120251cac93c9d5492d02b8d27&pid=1-s2.0-S1549963423000710-main.pdf","citationCount":"0","resultStr":"{\"title\":\"Intrinsic variability of fluorescence calibrators impacts the assignment of MESF or ERF values to nanoparticles and extracellular vesicles by flow cytometry\",\"authors\":\"Estefanía Lozano-Andrés PhD , Tina Van Den Broeck PhD , Lili Wang PhD , Majid Mehrpouyan PhD , Ye Tian PhD , Xiaomei Yan PhD , Ger J.A. Arkesteijn PhD , Marca H.M. Wauben PhD\",\"doi\":\"10.1016/j.nano.2023.102720\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Flow cytometry allows to characterize nanoparticles (NPs) and extracellular vesicles (EVs) but results are often expressed in arbitrary units of fluorescence. We evaluated the precision and accuracy of molecules of equivalent soluble fluorophores (MESF) beads for calibration of NPs and EVs. Firstly, two FITC-MESF bead sets, 2 and 6 um in size, were measured on three flow cytometers. We showed that arbitrary units could not be compared between instruments but after calibration, comparable FITC MESF units were achieved. However, the two calibration bead sets displayed varying slopes that were consistent across platforms.</p><p>Further investigation revealed that the intrinsic uncertainty related to the MESF beads impacts the robust assignment of values to NPs and EVs based on extrapolation into the dim fluorescence range. Similar variations were found with PE MESF calibration.</p><p>Therefore, the same calibration materials and numbers of calibration points should be used for reliable comparison of submicron sized particles.</p></div>\",\"PeriodicalId\":19050,\"journal\":{\"name\":\"Nanomedicine : nanotechnology, biology, and medicine\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":4.2000,\"publicationDate\":\"2023-11-23\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S1549963423000710/pdfft?md5=7ce6e8120251cac93c9d5492d02b8d27&pid=1-s2.0-S1549963423000710-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Nanomedicine : nanotechnology, biology, and medicine\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S1549963423000710\",\"RegionNum\":2,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"MEDICINE, RESEARCH & EXPERIMENTAL\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Nanomedicine : nanotechnology, biology, and medicine","FirstCategoryId":"3","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S1549963423000710","RegionNum":2,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"MEDICINE, RESEARCH & EXPERIMENTAL","Score":null,"Total":0}
Intrinsic variability of fluorescence calibrators impacts the assignment of MESF or ERF values to nanoparticles and extracellular vesicles by flow cytometry
Flow cytometry allows to characterize nanoparticles (NPs) and extracellular vesicles (EVs) but results are often expressed in arbitrary units of fluorescence. We evaluated the precision and accuracy of molecules of equivalent soluble fluorophores (MESF) beads for calibration of NPs and EVs. Firstly, two FITC-MESF bead sets, 2 and 6 um in size, were measured on three flow cytometers. We showed that arbitrary units could not be compared between instruments but after calibration, comparable FITC MESF units were achieved. However, the two calibration bead sets displayed varying slopes that were consistent across platforms.
Further investigation revealed that the intrinsic uncertainty related to the MESF beads impacts the robust assignment of values to NPs and EVs based on extrapolation into the dim fluorescence range. Similar variations were found with PE MESF calibration.
Therefore, the same calibration materials and numbers of calibration points should be used for reliable comparison of submicron sized particles.
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The mission of Nanomedicine: Nanotechnology, Biology, and Medicine (Nanomedicine: NBM) is to promote the emerging interdisciplinary field of nanomedicine.
Nanomedicine: NBM is an international, peer-reviewed journal presenting novel, significant, and interdisciplinary theoretical and experimental results related to nanoscience and nanotechnology in the life and health sciences. Content includes basic, translational, and clinical research addressing diagnosis, treatment, monitoring, prediction, and prevention of diseases.
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