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Automated Reporter Quantification In Vivo: High-Throughput Screening Method for Reporter-Based Assays in Zebrafish
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Automated Reporter Quantification In Vivo: High-Throughput Screening Method for Reporter-Based Assays in Zebrafish
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Journal Article
Automated Reporter Quantification In Vivo: High-Throughput Screening Method for Reporter-Based Assays in Zebrafish
2012
Overview
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.
Publisher
Public Library of Science,Public Library of Science (PLoS)
Subject
/ Animals
/ Animals, Genetically Modified
/ Assaying
/ Bacterial Proteins - genetics
/ Bacterial Proteins - metabolism
/ Biology
/ Cancer
/ Embryos
/ Gene Expression Profiling - methods
/ Gene Expression Regulation, Developmental
/ Genomics
/ High-Throughput Screening Assays - methods
/ Image Processing, Computer-Assisted - methods
/ Larvae
/ Luminescent Proteins - genetics
/ Luminescent Proteins - metabolism
/ Methods
/ Oxygen
/ Pancreas
/ R&D
