RESUMEN
Infants are screened for cystic fibrosis (CF) in New York State (NYS) using an IRT-DNA algorithm. The purpose of this study was to validate and assess clinical validity of the US FDA-cleared Illumina MiSeqDx CF 139-Variant Assay (139-VA) in the diverse NYS CF population. The study included 439 infants with CF identified via newborn screening (NBS) from 2002 to 2012. All had been screened using the Abbott Molecular CF Genotyping Assay or the Hologic InPlex CF Molecular Test. All with CF and zero or one mutation were tested using the 139-VA. DNA extracted from dried blood spots was reliably and accurately genotyped using the 139-VA. Sixty-three additional mutations were identified. Clinical sensitivity of three panels ranged from 76.2% (23 mutations recommended for screening by ACMG/ACOG) to 79.7% (current NYS 39-mutation InPlex panel), up to 86.0% for the 139-VA. For all, sensitivity was highest in Whites and lowest in the Black population. Although the sample size was small, there was a nearly 20% increase in sensitivity for the Black CF population using the 139-VA (68.2%) over the ACMG/ACOG and InPlex panels (both 50.0%). Overall, the 139-VA is more sensitive than other commercially available panels, and could be considered for NBS, clinical, or research laboratories conducting CF screening.
Asunto(s)
Bioensayo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/genética , Fibrosis Quística/diagnóstico , Fibrosis Quística/genética , Mutación , Población Negra , Fibrosis Quística/etnología , Fibrosis Quística/patología , Pruebas con Sangre Seca , Femenino , Pruebas Genéticas , Técnicas de Genotipaje , Hispánicos o Latinos , Humanos , Lactante , Recién Nacido , Masculino , Tamizaje Neonatal , Sensibilidad y Especificidad , Población BlancaRESUMEN
A reversed-phase high performance liquid chromatographic (LC), tandem mass spectrometry (MS/MS) assay for the determination of tenofovir (TFV) and emtricitabine (FTC) in dried blood spots (DBS) from human whole blood was developed and validated. Whole blood samples were spotted, dried, and a 3mm punch was extracted with methanol for analysis by LC-MS/MS utilizing stable isotope labeled internal standards. The assay was validated over the range of 2.5-1000ng/mL for TFV and 2.5-5000ng/mL for FTC. The method was accurate (within ±15% of control) and precise (coefficient of variation ≤15%) for hematocrit concentrations ranging from 25% to 76%; using edge punches vs. center punches; and spot volumes of 10-50µL. Analytes were stable for five freeze/thaw cycles and up to 6 days at room temperature, whereas long-term storage required -20°C or -80°C. Comparison of TFV and FTC in DBS vs. plasma yielded r(2)≥0.96, indicating that DBS can be used as a plasma alternative for pharmacokinetic analyses in vivo.