Effects of Prolonged Storage of Whole Plasma or Isolated Plasma DNA on the Results of Circulating DNA Quantification Assays

Abstract

Analysis of molecular markers in biological fluids has been proposed as a tool for early detection and monitoring of cancer. Circulating plasma DNA concentrations have been found to be higher in cancer patients than in cancer-free control subjects, but little is known about the effect of specimen storage on plasma DNA concentrations. Here we investigated the impact of long-term storage of both plasma samples and purified plasma DNA on the reproducibility of plasma DNA quantification as determined using real-time polymerase chain reaction analysis. The analysis was performed on samples from a subset of 34 lung cancer patients and 28 matched control subjects selected from 200 subjects in our previously published case-control study and from 117 cancer-free smokers enrolled in a lung cancer screening program. Two samples of plasma and isolated DNA were assessed for each patient, with a median of 41 months between the first and second assessments for participants in the case-control study and 9 months for participants in the screening study. DNA levels declined substantially between the two assessments at an average rate of approximately 30% per year. These data provide valuable information for the rational planning of retrospective studies of banked series of biological samples, particularly if collected over a long period of time, as can occur in large clinical trials.

Detection of mutated BRAFV600E variant in circulating DNA of stage III–IV melanoma patients

“…31 Our data indicate that while BRAFV600E can be detected in both serum and plasma from melanoma patients, plasma provides the best source to detect circulating BRAFV600E, although a direct comparison between serum and plasma was not obtained. In addition, our experiments indicate that DNA should be isolated from plasma samples and frozen in the setting of clinical monitoring studies, because its amount decreases after protracted storage, probably for degradation, and that the PCR techniques lose reproducibility, as recently shown by Sozzi et al 37 In contrast to that of patients, plasma and serum from healthy donors were reproducibly negative for the BRAFV600E marker. These results indicate that although the BRAFV600E mutation occurs at a high frequency also in benign melanocytic nevi, 28 it is not detectable in circulating DNA of healthy subjects with nevi.…”

Detection of BRAF mutations in the tumour and serum of patients enrolled in the AZD6244 (ARRY-142886) advanced melanoma phase II study

“…Third, concern has been raised with regard to the reliability of cfDNA mutation detection in stored blood samples (Sozzi et al, 2005;Daniotti et al, 2007). Previous reports have demonstrated that cfDNA mutations are no longer detectable in previously positive serum and DNA samples after 12 months of storage (Daniotti et al, 2007).…”

Cell-Free DNA: An Upcoming Biomarker in Transplantation

“…Sample processing: within 6-8 h after collection (46)(47)(48) Use of specialized blood collecting tubes (Cell-Free RNA BCT 1 -Streck) that allow 7 days storage of blood by increasing cell stability and inhibiting nuclease mediated DNA degradation (48,49) 7-30% yearly DNA degradation during storage (50,51) cfDNA isolation:…”