PMID- 38368382 OWN - NLM STAT- MEDLINE DCOM- 20240219 LR - 20240220 IS - 2047-783X (Electronic) IS - 0949-2321 (Print) IS - 0949-2321 (Linking) VI - 29 IP - 1 DP - 2024 Feb 17 TI - Concentration strategies for spiked and naturally present biomarkers in non-invasively collected first-void urine. PG - 131 LID - 10.1186/s40001-024-01719-5 [doi] LID - 131 AB - BACKGROUND: First-void urine (FVU) provides a non-invasive method for collecting a wide range of biomarkers found in genital tract secretions. To optimize biomarker collection in FVU, this study investigated the impact of naturally present and supplemented precipitating agents: uromodulin (UMOD) and polyethylene glycol (PEG), on the concentration of human papillomavirus (HPV) pseudovirions (PsV), cell-free DNA (cfDNA), and cellular genomic DNA (gDNA) through centrifugation. METHODS: FVU samples from ten healthy female volunteers, along with a control sample, were spiked with seal herpesvirus 1 (PhHV-1) DNA, HPV16 plasmid DNA, and HPV16 PsV with an enhanced green fluorescent protein (EGFP) reporter. The samples were subjected to various concentration protocols involving PEG precipitation, low-speed centrifugation (5 min at 1000xg), and medium-speed centrifugation (1 h at 3000xg). Subsequently, quantitative PCR (qPCR) was used to assess cellular and cell-free glyceraldehyde-3-phosphate dehydrogenase (GAPDH) DNA, cell-free PhHV-1 and HPV16 DNA, and PsV (EGFP) DNA. In addition, UMOD levels were measured. RESULTS: The findings revealed that PEG significantly increased the concentration of cfDNA and gDNA in the pellet after centrifugation, with the most pronounced effect observed for cfDNA. Moreover, low-speed centrifugation without PEG effectively depleted cellular gDNA while preserving cfDNA in the supernatants. Pseudovirions were consistently pelleted, even with low-speed centrifugation, and a positive but not significant effect of PEG on PsV (EGFP) DNA yield in the pellet was observed. Additionally, a significant correlation was observed between UMOD and GAPDH, HPV16, and PsV (EGFP) DNA quantities in the pellet. Furthermore, large variations among the FVU samples were observed. CONCLUSIONS: With this study, we provide novel insights into how various biomarker precipitation protocols, including both the properties of FVU and the use of PEG as a precipitating agent, influence the concentration of cfDNA, cellular gDNA, and pseudovirions. CI - (c) 2024. The Author(s). FAU - Teblick, Laura AU - Teblick L AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. laura.teblick@uantwerpen.be. FAU - Lipovac, Marijana AU - Lipovac M AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. FAU - Burdier, F Ricardo AU - Burdier FR AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. FAU - De Smet, Annemie AU - De Smet A AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. FAU - Bell, Margo AU - Bell M AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. FAU - van den Borst, Eef AU - van den Borst E AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. AD - Centre of Medical Genetics, Faculty of Medicine and Health Sciences, University of Antwerp and Antwerp University Hospital, 2650, Edegem, Belgium. FAU - Matheeussen, Veerle AU - Matheeussen V AD - Department of Microbiology, Antwerp University Hospital (UZA), 2650, Edegem-Antwerp, Belgium. AD - Department of Medical Microbiology (LMM), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. AD - Department of Medical Biochemistry, Faculty of Pharmaceutical, Biomedical and Veterinary Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. FAU - Vorsters, Alex AU - Vorsters A AD - Centre for the Evaluation of Vaccination (CEV), Vaccine & Infectious Disease Institute (VAXINFECTIO), Faculty of Medicine and Health Sciences, University of Antwerp, 2610, Wilrijk-Antwerp, Belgium. LA - eng GR - 101040588/ERC_/European Research Council/International PT - Journal Article DEP - 20240217 PL - England TA - Eur J Med Res JT - European journal of medical research JID - 9517857 RN - 0 (Cell-Free Nucleic Acids) RN - 0 (Biomarkers) RN - 9007-49-2 (DNA) SB - IM MH - Humans MH - Female MH - *Human papillomavirus 16/genetics MH - *Cell-Free Nucleic Acids MH - Biomarkers MH - DNA PMC - PMC10873940 OTO - NOTNLM OT - Biomarkers OT - Concentration OT - FVU OT - First-void urine OT - Human papillomavirus COIS- A.V. is a co-founder and former board member of Novosanis (Subsidiary of OraSure Technologies Inc, Wijnegem, Belgium), a spin-off company of the University of Antwerp, and was a minority shareholder until January 2019. The University of Antwerp received grants from Merck, GSK, Hologic, Abbott, Roche, and Cepheid to support the HPV Prevention and Control Board. The University of Antwerp received a project grant and honoraria fee for lectures, presentations, and speaker bureaus from Merck. Other authors declare that they have no conflict of interest. EDAT- 2024/02/18 00:42 MHDA- 2024/02/19 06:43 PMCR- 2024/02/17 CRDT- 2024/02/17 23:17 PHST- 2024/01/03 00:00 [received] PHST- 2024/02/08 00:00 [accepted] PHST- 2024/02/19 06:43 [medline] PHST- 2024/02/18 00:42 [pubmed] PHST- 2024/02/17 23:17 [entrez] PHST- 2024/02/17 00:00 [pmc-release] AID - 10.1186/s40001-024-01719-5 [pii] AID - 1719 [pii] AID - 10.1186/s40001-024-01719-5 [doi] PST - epublish SO - Eur J Med Res. 2024 Feb 17;29(1):131. doi: 10.1186/s40001-024-01719-5.