Concentration Strategies for Spiked and Naturally Present Biomarkers in Non-invasively Collected First-void Urine

Overview

Journal Eur J Med Res

Publisher Biomed Central

Specialty General Medicine

Date 2024 Feb 17

PMID 38368382

Authors

Laura Teblick

Marijana Lipovac

F Ricardo Burdier

Annemie De Smet

Margo Bell

Eef van den Borst

Veerle Matheeussen

Alex Vorsters

Affiliations

Soon will be listed here.

Abstract

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 1000×g), and medium-speed centrifugation (1 h at 3000×g). 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.

Citing Articles

HPV-specific antibodies in female genital tract secretions captured via first-void urine retain their neutralizing capacity.

Teblick L, Lipovac M, Molenberghs F, Delputte P, De Vos W, Vorsters A Hum Vaccin Immunother. 2024; 20(1):2330168.

PMID: 38567541 PMC: 10993920. DOI: 10.1080/21645515.2024.2330168.

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