Automated reporter quantification in vivo: high-throughput screening method for reporter-based assays in zebrafish.
PLoS One
; 7(1): e29916, 2012.
Article
en En
| MEDLINE
| ID: mdl-22238673
ABSTRACT
Reporter-based assays underlie many high-throughput screening (HTS) platforms, but most are limited to in vitro applications. Here, we report a simple whole-organism HTS method for quantifying changes in reporter intensity in individual zebrafish over time termed, Automated Reporter Quantification in vivo (ARQiv). ARQiv differs from current "high-content" (e.g., confocal imaging-based) whole-organism screening technologies by providing a purely quantitative data acquisition approach that affords marked improvements in throughput. ARQiv uses a fluorescence microplate reader with specific detection functionalities necessary for robust quantification of reporter signals in vivo. This approach is 1) Rapid; achieving true HTS capacities (i.e., >50,000 units per day), 2) Reproducible; attaining HTS-compatible assay quality (i.e., Z'-factors of ≥0.5), and 3) Flexible; amenable to nearly any reporter-based assay in zebrafish embryos, larvae, or juveniles. ARQiv is used here to quantify changes in 1) Cell number; loss and regeneration of two different fluorescently tagged cell types (pancreatic beta cells and rod photoreceptors), 2) Cell signaling; relative activity of a transgenic Notch-signaling reporter, and 3) Cell metabolism; accumulation of reactive oxygen species. In summary, ARQiv is a versatile and readily accessible approach facilitating evaluation of genetic and/or chemical manipulations in living zebrafish that complements current "high-content" whole-organism screening methods by providing a first-tier in vivo HTS drug discovery platform.
Texto completo:
1
Base de datos:
MEDLINE
Asunto principal:
Procesamiento de Imagen Asistido por Computador
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Pez Cebra
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Genes Reporteros
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Dosificación de Gen
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Ensayos Analíticos de Alto Rendimiento
Tipo de estudio:
Diagnostic_studies
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Evaluation_studies
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Screening_studies
Idioma:
En
Revista:
PLoS One
Asunto de la revista:
CIENCIA
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MEDICINA
Año:
2012
Tipo del documento:
Article