facilitating-drug-discovery-an-automated-high-content-inflammation

Facilitating Drug Discovery: An Automated High-content Inflammation Assay in Zebrafish

Zebrafish larvae are particularly amenable to whole animal small molecule screens1,2 due to their small size and relative ease of manipulation and ...

In this chapter we describe automated imaging methods used to measure the transport of an established membrane transport marker from the endoplasmic reticulum to the plasma membrane. The method is fast and significantly robust to be applied in systematic studies on a large scale such as genome-wide screening projects. We further describe the use of software macros and plugins in Image J that allow the quantification of the kinetics of membrane transport intermediates in fluorescence microscopy time-lapse sequences.

Imaging platforms for measurement of membrane trafficking.

The development of automated microscopy platforms has enabled large-scale observation of biological processes, thereby complementing genome scale biochemical techniques. However, commercially available systems are restricted either by fixed-field-of-views, leading to potential omission of features of interest, or by low-resolution data of whole objects lacking cellular detail. This limits the efficiency of high-content screening assays, especially when large complex objects are used as in whole-organism screening. Here we demonstrate a toolset for automated intelligent high-content screening of whole zebrafish embryos at cellular resolution on a standard wide-field screening microscope. Using custom-developed algorithms, predefined regions of interest-such as the brain-are automatically detected. The regions of interest are subsequently imaged automatically at high magnification, enabling rapid capture of cellular resolution data. We utilize this approach for acquiring 3-D datasets of embryonic brains of transgenic zebrafish. Moreover, we report the development of a mold design for accurate orientation of zebrafish embryos for dorsal imaging, thereby facilitating standardized imaging of internal organs and cellular structures. The toolset is flexible and can be readily applied for the imaging of different specimens in various applications.

Automated feature detection and imaging for high-resolution screening of zebrafish embryos.

Die Entwicklung der automatisierte Mikroskopie-Plattformen hat gro&szlig; angelegte Beobachtung biologischer Prozesse, damit erg&auml;nzt Genom Skala biochemische Techniken erm&ouml;glicht. Handels&uuml;bliche Systeme sind jedoch beschr&auml;nkt behoben-Feld-von-Ansichten, zu m&ouml;glichen Unterlassung der Funktionen von Interesse, oder durch niedrig aufl&ouml;sende Daten ganze Objekte zellul&auml;re Detail fehlt. Dies schr&auml;nkt die Wirksamkeit von hoch-Content Screening-Assays, insbesondere wenn gro&szlig;e komplexe Objekte wie im gesamten Organismus Screening verwendet werden. Hier zeigen wir eine Toolset f&uuml;r automatisierte intelligente hoch-Inhalt-Screening ganze Zebrafish Embryos bei zellul&auml;ren Aufl&ouml;sung am Mikroskop standard Breite-Feld-Vorf&uuml;hrung. Mit eigens entwickelte Algorithmen, vordefinierte Regionen von Interesse wie die Gehirn-werden automatisch erkannt. Die Regionen von Interesse sind anschlie&szlig;end automatisch bei hoher Vergr&ouml;&szlig;erung, damit schnellen Erfassung von zellul&auml;ren Aufl&ouml;sung Daten &uuml;berhaupt. Wir nutzen diesen Ansatz f&uuml;r den Erwerb von 3-d-Datasets von embryonalen Gehirn von transgenen Zebrab&auml;rbling. Dar&uuml;ber hinaus berichten wir die Entwicklung einer Werkzeugkonstruktion f&uuml;r genaue Ausrichtung der Zebrafisch-Embryonen f&uuml;r dorsalen Imaging, standardisierte Darstellung der inneren Organe und zellul&auml;re Strukturen erleichtern. Das Toolset ist flexibel und kann ohne weiteres f&uuml;r die Bildgebung von verschiedenen Exemplaren in verschiedenen Anwendungen angewendet werden.

Article	Year
FEBS letters\| PubMed ID: 14623100	2003
Bioinformatics (Oxford, England)\| PubMed ID: 14988114	2004
Genome research\| PubMed ID: 15466293	2004
Traffic (Copenhagen, Denmark)\| PubMed ID: 15941410	2005
Cancer research\| PubMed ID: 16140941	2005
Imaging platforms for measurement of membrane trafficking. Methods in enzymology\| PubMed ID: 16413253	2005
Methods in enzymology\| PubMed ID: 16413254	2005
Nature methods\| PubMed ID: 16628209	2006
Journal of biotechnology\| PubMed ID: 17275941	2007
Nature protocols\| PubMed ID: 17406600	2007
Biotechnology journal\| PubMed ID: 17680709	2007
The Journal of cell biology\| PubMed ID: 17954604	2007
Reproductive toxicology (Elmsford, N.Y.)\| PubMed ID: 19406227	2009
Nature methods\| PubMed ID: 19898487	2009
Nature\| PubMed ID: 20360735	2010
BMC biology\| PubMed ID: 21176202	2010
Nature methods\| PubMed ID: 21258339	2011
Bioinformatics (Oxford, England)\| PubMed ID: 21493657	2011
Automated feature detection and imaging for high-resolution screening of zebrafish embryos. BioTechniques\| PubMed ID: 21548893	2011
Journal of laboratory automation\| PubMed ID: 21609691	2011
Angewandte Chemie (International ed. in English)\| PubMed ID: 21751312	2011
Archives of toxicology\| PubMed ID: 21983885	2012
Trends in biotechnology\| PubMed ID: 22652049	2012

Urban Liebel

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