Name: live imaging of drosophila melanogaster embryonic hemocyte migrations
Uploaded: 2010-02-12T18:30:00
Description: Watch this Scientific Journal Video about Live Imaging Of Drosophila melanogaster Embryonic Hemocyte Migrations at JoVE.com

Dit protocol is ontwikkeld door middel van ons werk binnen en in samenwerking met de laboratoria van Paul Martin en Antonio Jacinto. Wij danken de Bloomington Stock Centrum voor zijn uitstekende service en de Drosophila gemeenschap voor de voortzetting van te vliegen lijnen te delen. BS wordt momenteel gefinancierd door een BBSRC projectsubsidie. WW wordt gefinancierd door een Wellcome Trust Career Development Fellowship.

Voorbereiding <ol><li> Verkrijgen van juiste Drosophila regels met een hemocyte-specifieke Gal4 driver (bv SRP-Gal4 2) en een genetisch gecodeerd fluorescerende reporter onder controle UAS (bijv. UAS-GFP). Vliegen homozygoot zijn voor SRP-Gal4, UAS-GMA 3 of CRQ-Gal4, UAS-GFP 4, 5 zijn vooral nuttig voor imaging doeleinden (nb GMA is GFP gefuseerd aan het actine-bindende domein van Moesin), zie hieronder voor een bespreking van het bereik van...

<ol>
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De belangrijkste elementen van deze procedure zijn de selectie van gezonde embryo&#39;s met duidelijk gelabeld hemocytes en zorgvuldig te monteren ze zonder ze te beschadigen. Zodra de embryo&#39;s worden in de halogene olie zijn ze bestand tegen uitdroging en eenmaal gemonteerd kunnen worden afgebeeld voor enkele uren. In onze handen kunnen wij image hemocytes drie uur lang, met een verwaarloosbare uitdroging van het embryo of de voor de hand liggende foto-schade, het nemen van een z-stack van beelden om de drie minute...

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<td>Cell strainer</td>
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<td>BD Falcon</td>
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<td>Halcarbon oil 700</td>
<td>&nbsp;</td>
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live imaging of drosophila melanogaster embryonic hemocyte migrations

Veel studies adres celmigratie met behulp van<em&gt; In vitro</em&gt; Methoden, terwijl de fysiologisch relevante omgeving is dat van het organisme zelf. Hier presenteren wij een protocol voor de montage van<em&gt; Drosophila melanogaster</em&gt; Embryo&#39;s en de daarop volgende live-beeldvorming van fluorescent gelabelde hemocytes, de embryonale macrofagen van dit organisme. Met behulp van de Gal4-UAS-systeem<sup&gt; 1</sup&gt; Rijden we de uitdrukking van een verscheidenheid van genetisch gecodeerd, fluorescent gelabelde markers in hemocytes om hun ontwikkelings-verspreiding te volgen in de embryo. Na de winning van embryo&#39;s bij de gewenste stadium van ontwikkeling, is de buitenste chorion verwijderd en de embryo&#39;s worden vervolgens gemonteerd in halogene olie tussen een hydrofoob, gas-permeabel membraan en een glazen dekglaasje voor live imaging. In aanvulling op de bruto trekkende parameters zoals snelheid en richting, een hogere resolutie imaging in combinatie met het gebruik van fluorescerende verslaggevers van F-actine en microtubuli kan meer gedetailleerde informatie over de dynamiek van deze cytoskelet componenten bieden.

Watch this Scientific Journal Video about Live Imaging Of Drosophila melanogaster Embryonic Hemocyte Migrations at JoVE.com

Live Imaging Of Drosophila melanogaster Embryonic Hemocyte Migrations

Drosophila hemocytes verspreiden over het geheel van de zich ontwikkelende embryo. Dit protocol laat zien hoe te monteren en het imago van deze migraties met behulp van embryo&#39;s met fluorescent gelabelde hemocytes.

Live-beeldvorming van Drosophila melanogaster Embryonale Hemocyte Migraties

Many studies address cell migration using in vitro methods, whereas the physiologically relevant environment is that of the organism itself. Here we ...

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Live Imaging Of Drosophila melanogaster Embryonic Hemocyte Migrations

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