Effect of sample type on plasma concentrations of cell‐free DNA and nucleosomes in dogs

Abstract

Cell-free DNA (cfDNA) and nucleosomes are two biomarkers of cell death and neutrophil extracellular trap formation that are increased in dogs with sepsis, immune-mediated haemolytic anaemia, cancer and following trauma and have diagnostic and prognostic values. cfDNA and nucleosomes are typically measured in plasma samples using DNA-specific fluorophores and ELISA assays, respectively, but their concentrations may be affected by pre-analytical variables such as sample type. The present study aimed to investigate the influence of sample type on the plasma cfDNA and nucleosome concentrations of a heterogeneous group of dogs presenting to an emergency room. Triplicate samples were collected into K2-ethylenediamine tetraacetic acid, 3.2% citrate and a specialised DNA stabilisation tube (Streck BCT), processed rapidly and frozen for batch analysis. Biomarker concentrations were compared between sample types by calculation of Spearman’s correlation coefficients, and with Deming regression, Bland-Altman plots and the Friedman test. Overall, biomarker concentrations were highly correlated between the three sample types. The most concordant results were obtained using citrate samples and the DNA stabilisation tube. Matched cfDNA concentrations between the different sample types were significantly different but there was no significant difference between the nucleosome concentrations in any of the sample types. The present study suggests that cfDNA and nucleosomes can be successfully measured in various sample types, but distinct sample types do not produce interchangeable results. This argues for use of a consistent sample type within studies and suggests standardisation may be useful for the field.

Evaluation of nucleosome concentrations in healthy dogs and dogs with cancer

“…This is significantly lower than the concentration seen in the dogs with lymphoma reported in this study with median of 590 ng/mL. Comparisons between the concentrations of nucleosomes reported here and those in other reports of inflammation and control groups are difficult due to the fact that nucleosome concentrations have previously been reported only in arbitrary units(7)(8)(9)(20)(21)(22).…”

Characterizing circulating nucleosomes in the plasma of dogs with hemangiosarcoma

“…Liquid biopsy (LB) techniques refer to the minimally invasive approach of sampling tumor specific biomarkers in the blood of diseased patients [ 1 ]. LB has been most studied in the field of oncology; however, literature also exists on the use of LB for the prognostication of inflammation, sepsis and trauma in humans and dogs [ 2 – 5 ]. In general, LB techniques are used to identify intact circulating tumor cells, extracellular nucleic acids such as cell free DNA (cfDNA), messenger RNA (mRNA), microRNA (miRNA), extracellular vesicles (exosomes), nucleosomes and glycoproteins or antigens in the circulation (e.g.…”

Characterizing Circulating Nucleosomes in the Plasma of Dogs with Hemangiosarcoma

“…Liquid biopsy (LB) techniques refer to the minimally invasive approach of sampling tumor speci c biomarkers in the blood of diseased patients(1). LB has been most studied in the eld of oncology; however, literature also exists on the use of LB for the prognostication of in ammation, sepsis and trauma in humans and dogs (2)(3)(4)(5). In general, LB techniques are used to identify intact circulating tumor cells, extracellular nucleic acids such as cell free DNA (cfDNA), messenger RNA (mRNA), microRNA (miRNA), extracellular vesicles (exosomes), nucleosomes and glycoproteins or antigens in the circulation (e.g.…”