A protocol for urine collection and storage prior to DNA methylation analysis

PloS One
J BosschieterJ A Nieuwenhuijzen

Abstract

Urine poses an attractive non-invasive means for obtaining liquid biopsies for oncological diagnostics. Especially molecular analysis on urinary DNA is a rapid growing field. However, optimal and practical storage conditions that result in preservation of urinary DNA, and in particular hypermethylated DNA (hmDNA), are yet to be determined. To determine the most optimal and practical conditions for urine storage that result in adequate preservation of DNA for hmDNA analysis. DNA yield for use in methylation analysis was determined by quantitative methylation specific PCR (qMSP) targeting the ACTB and RASSF1A genes on bisulfite modified DNA. First, DNA yield (ACTB qMSP) was determined in a pilot study on urine samples of healthy volunteers using two preservatives (Ethylenediaminetetraacetic acid (EDTA) and Urine Conditioning Buffer, Zymo Research) at four different temperatures (room temperature (RT), 4°C, -20°C, -80°C) for four time periods (1, 2, 7, 28 days). Next, hmDNA levels (RASSF1A qMSP) in stored urine samples of patients suffering from bladder cancer (n = 10) or non-small cell lung cancer (NSCLC; n = 10) were measured at day 0 and 7 upon storage with and without the addition of 40mM EDTA and/or 20 μl/ml Penicillin Strept...Continue Reading

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Citations

Jul 24, 2020·Current Opinion in Urology·Stephan BrönimannShahrokh F Shariat
Apr 7, 2020·PloS One·Elien AugustusPatrick Pauwels
Jan 29, 2021·Scientific Reports·S BachR D M Steenbergen
Nov 5, 2020·Clinical Epigenetics·Rianne van den HelderRenske D M Steenbergen
May 1, 2021·Molecules : a Journal of Synthetic Chemistry and Natural Product Chemistry·Laura TéblickAlex Vorsters
Jun 3, 2021·Cancers·Masanori OshiKazuaki Takabe
Oct 5, 2021·Epigenetics : Official Journal of the DNA Methylation Society·Sander BachRenske D M Steenbergen

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Methods Mentioned

BETA
collection
Assay
PCR
urine storage

Software Mentioned

FSA
PenStrep
SPSS
R

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