@article{dc6d75f4283d4c55a23f0308905f9243,
title = "Irreversible alteration of extracellular vesicle and cell-free messenger RNA profiles in human plasma associated with blood processing and storage",
abstract = "The discovery and utility of clinically relevant circulating biomarkers depend on standardized methods that minimize preanalytical errors. Despite growing interest in studying extracellular vesicles (EVs) and cell-free messenger RNA (cf-mRNA) as potential biomarkers, how blood processing and freeze/thaw impacts the profiles of these analytes in plasma was not thoroughly understood. We utilized flow cytometric analysis to examine the effect of differential centrifugation and a freeze/thaw cycle on EV profiles. Utilizing flow cytometry postacquisition analysis software (FCMpass) to calibrate light scattering and fluorescence, we revealed how differential centrifugation and post-freeze/thaw processing removes and retains EV subpopulations. Additionally, cf-mRNA levels measured by RT-qPCR profiles from a panel of housekeeping, platelet, and tissue-specific genes were preferentially affected by differential centrifugation and post-freeze/thaw processing. Critically, freezing plasma containing residual platelets yielded irreversible ex vivo generation of EV subpopulations and cf-mRNA transcripts, which were not removable by additional processing after freeze/thaw. Our findings suggest the importance of minimizing confounding variation attributed to plasma processing and platelet contamination.",
author = "Kim, {Hyun Ji} and Rames, {Matthew J.} and {Tassi Yunga}, Samuel and Randall Armstrong and Mayu Morita and Ngo, {Anh T.P.} and McCarty, {Owen J.T.} and Fehmi Civitci and Morgan, {Terry K.} and Ngo, {Thuy T.M.}",
note = "Funding Information: The authors would like to acknowledge Joshua A. Welsh from National Cancer Institute at NIH for helpful discussion and implementation of FCMpass software. We are also grateful to Nick Wang of OHSU for facilitating access to Quantstudio 7 Flex Real-Time PCR systems. We are also grateful to Jeong Youn Lim for biostatistics advice and the Cancer Early Detection Advanced Research (CEDAR) center of the OHSU Knight Cancer Institute for assistance with obtaining blood samples. The authors also thank Pamela S Canaday, Brianna Garcia, and Dorian Latocha for flow cytometry training at Oregon Health and Science University. We acknowledge funding support from Cancer Early Detection Advanced Research (CEDAR) center at Oregon Health and Science University{\textquoteright}s Knight Cancer Institute, Cancer Research UK/OHSU Project Award (C63763/A27122) and by grants from Kuni Foundation, Susan G. Komen Foundation (CCR21663959), the Department of Defense (W81XWH2110853), and the National Institutes of Health (R01HL101972, R21HD16-037, R01GM116184 and R01HL047014). Funding Information: The authors would like to acknowledge Joshua A. Welsh from National Cancer Institute at NIH for helpful discussion and implementation of FCMpass software. We are also grateful to Nick Wang of OHSU for facilitating access to Quantstudio 7 Flex Real-Time PCR systems. We are also grateful to Jeong Youn Lim for biostatistics advice and the Cancer Early Detection Advanced Research (CEDAR) center of the OHSU Knight Cancer Institute for assistance with obtaining blood samples. The authors also thank Pamela S Canaday, Brianna Garcia, and Dorian Latocha for flow cytometry training at Oregon Health and Science University. We acknowledge funding support from Cancer Early Detection Advanced Research (CEDAR) center at Oregon Health and Science University?s Knight Cancer Institute, Cancer Research UK/OHSU Project Award (C63763/A27122) and by grants from Kuni Foundation, Susan G. Komen Foundation (CCR21663959), the Department of Defense (W81XWH2110853), and the National Institutes of Health (R01HL101972, R21HD16-037, R01GM116184 and R01HL047014). Publisher Copyright: {\textcopyright} 2022, The Author(s).",
year = "2022",
month = dec,
doi = "10.1038/s41598-022-06088-9",
language = "English (US)",
volume = "12",
journal = "Scientific Reports",
issn = "2045-2322",
publisher = "Nature Publishing Group",
number = "1",
}