URINE CELL-FREE DNA AS A POTENTIAL BIOMARKER FOR OBSTETRIC COMPLICATIONS. AN ALTERNATIVE DETECTION METHOD

RACCA, MARÍA EMILIA; CEPEDA PAULA JULIETA; ROSSETTI, MARÍA FLORENCIA; RAMOS, JORGE GUILLERMO; CARDOZO, MARÍA ALEJANDRA; LUQUE, ENRIQUE H; MUÑOZ-DE-TORO, MÓNICA; MILESI, MARÍA MERCEDES; VARAYOUD, JORGELINA

Congreso; Reunión Anual de Sociedades de Biociencias. SAIC SAI&FAIC SAFIS; 2022

Sociedad Argentina de Investigación Clínica

Urine cell-free DNA (cfDNA) could serve as a biomarker for different diseases (e.g., obstetriccomplications). Noninvasive sampling and the possibility to analyze large volume samplesare advantages of urine-based protocols. Urine cell-free fDNA assessment usually includescommercial kits. We aimed to determine the efficiency of an alternative purification methodto extract and quantify urine cfDNA, comparing cfDNA levels on pregnant (P) andnon-pregnant (NP) women. cfDNA was isolated from urine samples of P (first, second andthird trimester, n=47) and NP (n=23) women. After urine pH neutralization andhigh-revolution centrifugation, cfDNA was purified from 800 μL supernatant applying anorganic extraction method. First, urine samples were lysed with 5M NaCl and 20% sodiumdodecyl sulfate solutions, then cfDNA was extracted with chloroform followed by ethanolprecipitation. Urine cfDNA was resuspended in 50 μL Tris-EDTA buffer and resultingconcentrations were measured by UV spectrophotometry. Purified cfDNA integrity wasanalyzed by agarose gel electrophoresis. Finally, cfDNA levels were quantified by real timePCR amplification of Actin-β gene (ACTB). cfDNA was detected in 69% urine samples andyielded specific ACTB amplification with low standard deviation between PCR duplicates (Ctdifference