Gutenberg Open Science: Validating quantitative PCR assays for cfDNA detection without DNA extraction in exercising SLE patients

Please use this identifier to cite or link to this item: http://doi.org/10.25358/openscience-7000

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DC Field	Value	Language
dc.contributor.author	Neuberger, Elmo W. I.	-
dc.contributor.author	Brahmer, Alexandra	-
dc.contributor.author	Ehlert, Tobias	-
dc.contributor.author	Kluge, Katrin	-
dc.contributor.author	Philippi, Keito F. A.	-
dc.contributor.author	Boedecker, Simone C.	-
dc.contributor.author	Weinmann-Menke, Julia	-
dc.contributor.author	Simon, Perikles	-
dc.date.accessioned	2022-05-19T09:09:07Z	-
dc.date.available	2022-05-19T09:09:07Z	-
dc.date.issued	2021	-
dc.identifier.uri	https://openscience.ub.uni-mainz.de/handle/20.500.12030/7012	-
dc.description.abstract	Circulating cell-free DNA (cfDNA) has been investigated as a screening tool for many diseases. To avoid expensive and time-consuming DNA isolation, direct quantification PCR assays can be established. However, rigorous validation is required to provide reliable data in the clinical and non-clinical context. Considering the International Organization for Standardization, as well as bioanalytical method validation guidelines, we provide a comprehensive procedure to validate assays for cfDNA quantification from blood plasma without DNA isolation. A 90 and 222 bp assay was validated to study the kinetics of cfDNA after exercise in patients with systemic lupus erythematosus (SLE). The assays showed ultra-low limit of quantification (LOQ) with 0.47 and 0.69 ng/ml, repeatability ≤ 11.6% (95% CI 8.1–20.3), and intermediate precision ≤ 12.1% (95% CI 9.2–17.7). Incurred sample reanalysis confirmed the precision of the procedure. The additional consideration of pre-analytical factors shows that centrifugation speed and temperature do not change cfDNA concentrations. In SLE patients cfDNA increases ~ twofold after a walking exercise, normalizing after 60 min of rest. The established assays allow reliable and cost-efficient quantification of cfDNA in minute amounts of plasma in the clinical setting. Additionally, the assay can be used as a tool to determine the impact of pre-analytical factors and validate cfDNA quantity and quality of isolated samples.	en_GB
dc.language.iso	eng	de
dc.rights	CC BY	*
dc.rights.uri	https://creativecommons.org/licenses/by/4.0/	*
dc.subject.ddc	610 Medizin	de_DE
dc.subject.ddc	610 Medical sciences	en_GB
dc.subject.ddc	796 Sport	de_DE
dc.subject.ddc	796 Athletic and outdoor sports and games	en_GB
dc.title	Validating quantitative PCR assays for cfDNA detection without DNA extraction in exercising SLE patients	en_GB
dc.type	Zeitschriftenaufsatz	de
dc.identifier.doi	http://doi.org/10.25358/openscience-7000	-
jgu.type.dinitype	article	en_GB
jgu.type.version	Published version	de
jgu.type.resource	Text	de
jgu.organisation.department	FB 02 Sozialwiss., Medien u. Sport	de
jgu.organisation.department	FB 04 Medizin	de
jgu.organisation.number	7910	-
jgu.organisation.number	2700	-
jgu.organisation.name	Johannes Gutenberg-Universität Mainz	-
jgu.rights.accessrights	openAccess	-
jgu.journal.title	Scientific reports	de
jgu.journal.volume	11	de
jgu.pages.alternative	13581	de
jgu.publisher.year	2021	-
jgu.publisher.name	Macmillan Publishers Limited, part of Springer Nature	de
jgu.publisher.place	London	de
jgu.publisher.issn	2045-2322	de
jgu.organisation.place	Mainz	-
jgu.subject.ddccode	610	de
jgu.subject.ddccode	796	de
jgu.publisher.doi	10.1038/s41598-021-92826-4	de
jgu.organisation.ror	https://ror.org/023b0x485	-
Appears in collections:	JGU-Publikationen

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