RESUMO
The worldwide coronavirus disease 2019 pandemic has had devastating effects on health, healthcare infrastructure, social structure, and economics. One of the limiting factors in containing the spread of this virus has been the lack of widespread availability of fast, inexpensive, and reliable methods for testing of individuals. Frequent screening for infected and often asymptomatic people is a cornerstone of pandemic management plans. Here, we introduce 2 pH-sensitive "LAMPshade" dyes as novel readouts in an isothermal Reverse Transcriptase Loop-mediated isothermal AMPlification amplification assay for severe acute respiratory syndrome coronavirus 2 RNA. The resulting JaneliaLAMP assay is robust, simple, inexpensive, and has low technical requirements, and we describe its use and performance in direct testing of contrived and clinical samples without RNA extraction.
Assuntos
COVID-19 , RNA Viral , Corantes , Humanos , Concentração de Íons de Hidrogênio , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , RNA Viral/genética , SARS-CoV-2 , Sensibilidade e Especificidade , Estrutura SocialRESUMO
The outflow tract (OFT) is abnormal in many congenital heart defects. One critical mechanism for morphogenesis of this complex structure is apoptosis. Chicken embryos (stages 19-38; ED4-10) stained with a fluorescent supravital lysosomal dye (LysoTracker Red; LTR) revealed the three-dimensional relationship between structural changes and apoptosis. The LTR staining peaked in the OFT myocardium at stages 27-32, consistent with our previous analyses using other apoptosis assays. While LTR stained under both the pulmonary artery and the aorta, it was most prevalent in the subaortic myocardium before its elimination. Furthermore, LTR staining was most abundant in the myocardium under intensely cytokeratin-positive, thick epicardium. These data support the hypothesis that temporally and spatially restricted apoptosis in the OFT myocardium allows the aorta and pulmonary artery to dock at the appropriate angle and level with the proper ventricle. These data also support a relationship between the differentiating epicardium and cardiomyocyte apoptosis.