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1.
Sci Rep ; 13(1): 3075, 2023 02 22.
Artículo en Inglés | MEDLINE | ID: mdl-36813822

RESUMEN

In response to the SARS-CoV-2 pandemic, we developed a multiplexed, paired-pool droplet digital PCR (MP4) screening assay. Key features of our assay are the use of minimally processed saliva, 8-sample paired pools, and reverse-transcription droplet digital PCR (RT-ddPCR) targeting the SARS-CoV-2 nucleocapsid gene. The limit of detection was determined to be 2 and 12 copies per µl for individual and pooled samples, respectively. Using the MP4 assay, we routinely processed over 1,000 samples a day with a 24-h turnaround time and over the course of 17 months, screened over 250,000 saliva samples. Modeling studies showed that the efficiency of 8-sample pools was reduced with increased viral prevalence and that this could be mitigated by using 4-sample pools. We also present a strategy for, and modeling data supporting, the creation of a third paired pool as an additional strategy to employ under high viral prevalence.


Asunto(s)
COVID-19 , SARS-CoV-2 , Humanos , SARS-CoV-2/genética , Saliva/química , ARN Viral/genética , Reacción en Cadena de la Polimerasa , Sensibilidad y Especificidad , Prueba de COVID-19
3.
J Mol Diagn ; 19(3): 404-416, 2017 05.
Artículo en Inglés | MEDLINE | ID: mdl-28433077

RESUMEN

Nearly 80% of cancer patients do not have genetic mutation results available at initial oncology consultation; up to 25% of patients begin treatment before receiving their results. These factors hinder the ability to pursue optimal treatment strategies. This study validates a blood-based genome-testing service that provides accurate results within 72 hours. We focused on targetable variants in advanced non-small cell lung carcinoma-epidermal growth factor receptor gene (EGFR) variant L858R, exon 19 deletion (ΔE746-A750), and T790M; GTPase Kirsten ras gene (KRAS) variants G12C/D/V; and echinoderm microtubule associated protein like and 4 anaplastic lymphoma receptor tyrosine kinase fusion (EML4-ALK) transcripts 1/2/3. Test development included method and clinical validation using samples from donors with (n = 219) or without (n = 30) cancer. Clinical sensitivity and specificity for each variant ranged from 78.6% to 100% and 94.2% to 100%, respectively. We also report on 1643 non-small cell lung carcinoma samples processed in our CLIA-certified laboratory. Mutation results were available within 72 hours for 94% of the tests evaluated. We detected 10.5% mutations for EGFR sensitizing (n = 2801 samples tested), 13.8% mutations for EGFR resistance (n = 1055), 13.2% mutations in KRAS (n = 3477), and 2% mutations for EML4-ALK fusion (n = 304). This rapid, highly sensitive, and actionable blood-based assay service expands testing options and supports faster treatment decisions.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/genética , Análisis Mutacional de ADN/métodos , Quinasa de Linfoma Anaplásico , Proteínas de Ciclo Celular/genética , Receptores ErbB/genética , Exones/genética , Humanos , Neoplasias Pulmonares/genética , Proteínas Asociadas a Microtúbulos/genética , Mutación , Proteínas de Fusión Oncogénica/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Tirosina Quinasas Receptoras/genética , Serina Endopeptidasas/genética
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