{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,1,18]],"date-time":"2026-01-18T03:45:10Z","timestamp":1768707910623,"version":"3.49.0"},"reference-count":38,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2024,8,1]],"date-time":"2024-08-01T00:00:00Z","timestamp":1722470400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"NIH-T-32 training"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Veterinary Sciences"],"abstract":"<jats:p>Cell-free DNA (cfDNA) is fragmented extracellular DNA that is released from cells into various body fluids. Previously published data from adult horses supports cfDNA as a potential disease biomarker, but also shows that direct measurement in plasma is inaccurate due to matrix effect. It is currently unknown whether a similar matrix effect exists in foal plasma. Given this, the objectives of the current study were to investigate foal plasma for potential matrix effect during fluorescence measurement of cfDNA using a Qubit fluorometer, and to determine whether neat and\/or extracted plasma cfDNA concentrations are significantly different in healthy, sick non-septic (SNS) or septic foals. We hypothesized that matrix effect would interfere with accurate fluorescent measurement of cfDNA in foal plasma. Further, we hypothesized that mean extracted cfDNA concentrations, and\/or extracted cfDNA:neutrophil ratio, would be elevated in plasma of septic foals compared to healthy or SNS foals. Cell-free DNA was measured in neat plasma, and following DNA extraction with a commercial kit, from 60 foals. Foal plasma exhibited both autofluorescence and non-specific dye binding, confirming matrix effect. However, even with extraction, no significant difference was found in cfDNA concentrations, or cfDNA:neutrophil ratios, between healthy (sepsis score \u2264 5), SNS (sepsis score 6\u201311 and negative blood culture), or septic (sepsis score \u2265 12 \u00b1 positive blood culture) foals. Our data show that matrix effect interferes with accurate Qubit measurement of cfDNA in foal plasma and supports previous findings that plasma cfDNA concentrations are not associated with sepsis diagnosis in foals. Further research is needed to better understand neutrophil function and dysfunction in foal sepsis.<\/jats:p>","DOI":"10.3390\/vetsci11080346","type":"journal-article","created":{"date-parts":[[2024,8,1]],"date-time":"2024-08-01T08:14:46Z","timestamp":1722500086000},"page":"346","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Investigation of Extracted Plasma Cell-Free DNA as a Biomarker in Foals with Sepsis"],"prefix":"10.3390","volume":"11","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3226-2166","authenticated-orcid":false,"given":"Kallie J.","family":"Hobbs","sequence":"first","affiliation":[{"name":"Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Bethanie L.","family":"Cooper","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Katarzyna","family":"Dembek","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5270-4566","authenticated-orcid":false,"given":"M. Katie","family":"Sheats","sequence":"additional","affiliation":[{"name":"Department of Clinical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695, USA"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2024,8,1]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"660","DOI":"10.1111\/vec.12348","article-title":"Retrospective evaluation of blood culture isolates and sepsis survival rate in foals in the Czech Republic: 50 cases (2011\u20132013)","volume":"25","year":"2015","journal-title":"J. Vet. Emerg. Crit. Care"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"113","DOI":"10.1186\/s13054-019-2407-8","article-title":"Neutrophil extracellular traps (NETs) exacerbate severity of infant sepsis","volume":"23","author":"Colon","year":"2019","journal-title":"Crit. Care"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"34","DOI":"10.1159\/000440975","article-title":"The Role of Cell-Free Plasma DNA in Critically Ill Patients with Sepsis","volume":"41","author":"Clementi","year":"2016","journal-title":"Blood Purif."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"29","DOI":"10.1186\/s40635-019-0251-4","article-title":"Comparison of the source and prognostic utility of cfDNA in trauma and sepsis","volume":"7","author":"Coke","year":"2019","journal-title":"Intensive Care Med. Exp."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"402","DOI":"10.1177\/10406387221078047","article-title":"Investigation of plasma cell-free DNA as a potential biomarker in horses","volume":"34","author":"Bayless","year":"2022","journal-title":"J. Vet. Diagn. Investig."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Li, R.H., and Tablin, F. (2018). A comparative review of neutrophil extracellular traps in sepsis. Front. Vet. Sci., 5.","DOI":"10.3389\/fvets.2018.00291"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"e2206789","DOI":"10.1002\/advs.202206789","article-title":"Circulating Cell-Free DNAs as a Biomarker and Therapeutic Target for Acetaminophen-Induced Liver Injury","volume":"10","author":"Sun","year":"2023","journal-title":"Adv. Sci."},{"key":"ref_8","doi-asserted-by":"crossref","unstructured":"Fingerhut, L., Ohnesorge, B., von Borstel, M., Schumski, A., Strutzberg-Minder, K., Morgelin, M., Deeg, C.A., Haagsman, H.P., Beineke, A., and von Kockritz-Blickwede, M. (2019). Neutrophil Extracellular Traps in the Pathogenesis of Equine Recurrent Uveitis (ERU). Cells, 8.","DOI":"10.3390\/cells8121528"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Colmer, S.F., Luethy, D., Abraham, M., Stefanovski, D., and Hurcombe, S.D. (2021). Utility of cell-free DNA concentrations and illness severity scores to predict survival in critically ill neonatal foals. PLoS ONE, 16.","DOI":"10.1371\/journal.pone.0242635"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Panizzi, L., Dittmer, K.E., Vignes, M., Doucet, J.S., Gedye, K., Waterland, M.R., Rogers, C.W., Sano, H., McIlwraith, C.W., and Riley, C.B. (2023). Plasma and Synovial Fluid Cell-Free DNA Concentrations Following Induction of Osteoarthritis in Horses. Animals, 13.","DOI":"10.3390\/ani13061053"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"18","DOI":"10.1111\/j.2042-3306.1988.tb01445.x","article-title":"Development of a scoring system for the early diagnosis of equine neonatal sepsis","volume":"20","author":"Brewer","year":"1988","journal-title":"Equine Vet. J."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1111\/vec.12592","article-title":"Measurement of plasma cell-free DNA concentrations in dogs with sepsis, trauma, and neoplasia","volume":"27","author":"Letendre","year":"2017","journal-title":"J. Vet. Emerg. Crit. Care"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"278","DOI":"10.1111\/vcp.12478","article-title":"Evaluation of assays for quantification of DNA in canine plasma as an indirect marker of NETosis","volume":"46","author":"Smith","year":"2017","journal-title":"Vet. Clin. Pathol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"6499","DOI":"10.1007\/s00216-014-7835-3","article-title":"Towards standardisation of cell-free DNA measurement in plasma: Controls for extraction efficiency, fragment size bias and quantification","volume":"406","author":"Devonshire","year":"2014","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"665","DOI":"10.21037\/tlcr.2016.12.03","article-title":"Comparison of methods for circulating cell-free DNA isolation using blood from cancer patients: Impact on biomarker testing","volume":"5","author":"Torrente","year":"2016","journal-title":"Transl. Lung Cancer Res."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"86253","DOI":"10.18632\/oncotarget.21134","article-title":"Liquid biopsy in colon cancer: Comparison of different circulating DNA extraction systems following absolute quantification of KRAS mutations using Intplex allele-specific PCR","volume":"8","author":"Kloten","year":"2017","journal-title":"Oncotarget"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"6","DOI":"10.1177\/0300985819861721","article-title":"Neutrophil-Extracellular Traps, Cell-Free DNA, and Immunothrombosis in Companion Animals: A Review","volume":"57","author":"Goggs","year":"2020","journal-title":"Vet. Pathol."},{"key":"ref_18","doi-asserted-by":"crossref","unstructured":"Polatoglou, E., Mayer, Z., Ungerer, V., Bronkhorst, A.J., and Holdenrieder, S. (2022). Isolation and Quantification of Plasma Cell-Free DNA Using Different Manual and Automated Methods. Diagnostics, 12.","DOI":"10.3390\/diagnostics12102550"},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"22","DOI":"10.1111\/eve.36_13534","article-title":"Investigation of plasma cell-free dna (cfdna) as a novel biomarker of inflammation and disease severity in equine colic patients","volume":"33","author":"Bayless","year":"2021","journal-title":"Equine Vet. Educ."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"14","DOI":"10.1016\/j.cca.2018.01.007","article-title":"The value of fluorimetry (Qubit) and spectrophotometry (NanoDrop) in the quantification of cell-free DNA (cfDNA) in malignant melanoma and prostate cancer patients","volume":"479","author":"Ponti","year":"2018","journal-title":"Clin. Chim. Acta"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"19624","DOI":"10.1038\/s41598-023-46663-2","article-title":"Cell-free DNA as diagnostic and prognostic biomarkers for adult sepsis: A systematic review and meta-analysis","volume":"13","author":"Charoensappakit","year":"2023","journal-title":"Sci. Rep."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"524","DOI":"10.1177\/1753425917719995","article-title":"Elevated levels of circulating cell-free DNA and neutrophil proteins are associated with neonatal sepsis and necrotizing enterocolitis in immature mice, pigs and infants","volume":"23","author":"Nguyen","year":"2017","journal-title":"Innate Immun."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"267","DOI":"10.1111\/evj.12387","article-title":"The development of equine immunity: Current knowledge on immunology in the young horse","volume":"47","author":"Perkins","year":"2015","journal-title":"Equine Vet. J."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"676","DOI":"10.2746\/042516401776249327","article-title":"Neutrophil functions and serum IgG in growing foals","volume":"33","author":"Demmers","year":"2001","journal-title":"Equine Vet. J."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"797","DOI":"10.1111\/j.1939-1676.2007.tb03024.x","article-title":"Serum opsonization capacity, phagocytosis, and oxidative burst activity in neonatal foals in the intensive care unit","volume":"21","author":"Gardner","year":"2007","journal-title":"J. Vet. Intern. Med."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"4908","DOI":"10.1182\/blood-2009-09-242388","article-title":"Delayed but functional neutrophil extracellular trap formation in neonates","volume":"114","author":"Marcos","year":"2009","journal-title":"Blood"},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"182","DOI":"10.1159\/000452615","article-title":"Less Neutrophil Extracellular Trap Formation in Term Newborns than in Adults","volume":"111","author":"Lipp","year":"2017","journal-title":"Neonatology"},{"key":"ref_28","first-page":"1178","article-title":"NETosis: Molecular Mechanisms, Role in Physiology and Pathology","volume":"85","author":"Vorobjeva","year":"2020","journal-title":"Biochemistry"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"499","DOI":"10.1111\/j.1751-0813.2005.tb13304.x","article-title":"Effect of plasma transfusion on neutrophil function in healthy and septic foals","volume":"83","author":"McTaggart","year":"2005","journal-title":"Aust. Vet. J."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1517","DOI":"10.1111\/jvim.12208","article-title":"The prognostic utility of degenerative left shifts in dogs","volume":"27","author":"Burton","year":"2013","journal-title":"J. Vet. Intern. Med."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"912","DOI":"10.1111\/jvim.12338","article-title":"Degenerative left shift as a prognostic tool in cats","volume":"28","author":"Burton","year":"2014","journal-title":"J. Vet. Intern. Med."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"140","DOI":"10.1111\/j.1939-1676.1998.tb02109.x","article-title":"Factors associated with survival in septicemic foals: 65 cases (1988\u20131995)","volume":"12","author":"Gayle","year":"1998","journal-title":"J. Vet. Intern. Med."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"503","DOI":"10.1111\/vec.12773","article-title":"Determining prognosis in canine sepsis by bedside measurement of cell-free DNA and nucleosomes","volume":"28","author":"Letendre","year":"2018","journal-title":"J. Vet. Emerg. Crit. Care"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1016\/j.cveq.2019.11.003","article-title":"Clinical Pathology in the Foal","volume":"36","author":"Barton","year":"2020","journal-title":"Vet. Clin. N. Am. Equine Pract."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"275","DOI":"10.1111\/evj.12279","article-title":"Re-evaluation of the sepsis score in equine neonates","volume":"47","author":"Weber","year":"2015","journal-title":"Equine Vet. J."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1136\/vetrec.165.4.114","article-title":"Detection of bloodstream infection in neonatal foals with suspected sepsis using real-time PCR","volume":"165","author":"Pusterla","year":"2009","journal-title":"Vet. Rec."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"405","DOI":"10.1111\/evj.12307","article-title":"Detection of bacteraemia and host response in healthy neonatal foals","volume":"47","author":"Hackett","year":"2015","journal-title":"Equine Vet. J."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"1185","DOI":"10.1111\/jvim.15087","article-title":"Evaluation of updated sepsis scoring systems and systemic inflammatory response syndrome criteria and their association with sepsis in equine neonates","volume":"32","author":"Wong","year":"2018","journal-title":"J. Vet. Intern. Med."}],"container-title":["Veterinary Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2306-7381\/11\/8\/346\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T15:27:50Z","timestamp":1760110070000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2306-7381\/11\/8\/346"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2024,8,1]]},"references-count":38,"journal-issue":{"issue":"8","published-online":{"date-parts":[[2024,8]]}},"alternative-id":["vetsci11080346"],"URL":"https:\/\/doi.org\/10.3390\/vetsci11080346","relation":{},"ISSN":["2306-7381"],"issn-type":[{"value":"2306-7381","type":"electronic"}],"subject":[],"published":{"date-parts":[[2024,8,1]]}}}