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1.
Sci Rep ; 10(1): 8122, 2020 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-32415199

RESUMO

Multiple platforms are commercially available for the detection of circulating cell-free tumour DNA (ctDNA) from liquid biopsies. Since platforms have different input and output variables, deciding what platform to use for a given clinical or research question can be daunting. This study aimed to provide insight in platform selection criteria by comparing four commercial platforms that detect KRAS ctDNA hotspot mutations: Bio-Rad droplet digital PCR (ddPCR), BioCartis Idylla, Roche COBAS z480 and Sysmex BEAMing. Platform sensitivities were determined using plasma samples from metastatic colorectal cancer (mCRC) patients and synthetic reference samples, thereby eliminating variability in amount of plasma analysed and ctDNA isolation methods. The prevalence of KRAS nucleotide alterations was set against platform-specific breadth of target. Platform comparisons revealed that ddPCR and BEAMing detect more KRAS mutations amongst mCRC patients than Idylla and COBAS z480. Maximum sample throughput was highest for ddPCR and COBAS z480. Total annual costs were highest for BEAMing and lowest for Idylla and ddPCR. In conclusion, when selecting a platform for detection of ctDNA hotspot mutations the desired test sensitivity, breadth of target, maximum sample throughput, and total annual costs are critical factors that should be taken into consideration. Based on the results of this study, laboratories will be able to select the optimal platform for their needs.


Assuntos
Biomarcadores Tumorais/genética , DNA Tumoral Circulante/genética , Neoplasias Colorretais/diagnóstico , Análise Mutacional de DNA/classificação , Análise Mutacional de DNA/métodos , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Biomarcadores Tumorais/sangue , DNA Tumoral Circulante/sangue , Neoplasias Colorretais/genética , Humanos , Biópsia Líquida , Reação em Cadeia da Polimerase/métodos , Estudos Prospectivos
2.
J Clin Virol ; 84: 19-23, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27664778

RESUMO

BACKGROUND: Parvovirus B19 (B19V) DNA can be detected in blood over a long period after acute infection. Several reports associate the presence of B19V DNA with disease, irrespective of timing of the initial B19V infection. OBJECTIVES: This study aims to analyze the properties of B19V DNA in blood, differentiating between bare, non-infectious strands of DNA and B19V DNA in viable virions. STUDY DESIGN: Ten blood donors with asymptomatic acute B19V infection were followed and sampled up to 22 months after infection. The samples were treated with and without an endonuclease and tested for B19V DNA, to distinguish between DNA in virions and naked DNA. RESULTS: In the acute phase of infection, high levels of B19V DNA were detected, concurrent with B19V IgM antibodies. B19V DNA apparently was encapsidated, as indicated by resistance to endonuclease degradation. Subsequently, B19V DNA remained detectable for more than one year in all donors at low levels (<105 IU/mL). Approximately 150days after infection B19V DNA became degradable by an endonuclease, indicating that this concerned naked DNA. In some donors a second endonuclease-resistant peak occurred. DISCUSSION: Detection of B19V DNA in blood by PCR does not necessarily imply that B19V replication takes place and that infectious B19V virions are present. We propose that remnant B19V DNA strands can be released from tissues without active replication. This finding urges to reconsider an assumed role of B19V infection mainly based on B19V DNA detection in blood, a much debated subject in clinical syndromes such as myocarditis and arthritis.


Assuntos
Doadores de Sangue , DNA Viral/sangue , Infecções por Parvoviridae/diagnóstico , Parvovirus B19 Humano/genética , Parvovirus B19 Humano/isolamento & purificação , Anticorpos Antivirais/sangue , Artrite/diagnóstico , Artrite/virologia , DNA Viral/genética , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Miocardite/diagnóstico , Miocardite/virologia , Infecções por Parvoviridae/virologia , Parvovirus B19 Humano/imunologia , Reação em Cadeia da Polimerase , Fatores de Tempo , Replicação Viral
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