Name: fluorescent labeling of drosophila heart structures
Uploaded: 2009-10-13T23:30:00
Description: Watch this Scientific Journal Video about Fluorescent Labeling of Drosophila Heart Structures at JoVE.com

作者感谢SI伯恩斯坦（圣迭戈州立大学）批判性阅读和有益的建议，关于这个手稿的准备。这项工作是由美国国立卫生研究院拨款支持SI伯恩斯坦，SDSU的，和R.博德默，BIMR;和一个博士后奖学金从西方国家的美国心脏协会联盟G. Vogler和A. Cammarato。

在您开始之前<ol><li>准备了以下解决方案： <ol class="lalpha"><li>放宽缓冲区（人工果蝇血淋巴（ADH）的（见“可视化在果蝇中跳动的心脏”），其中包含10 mM的EGTA） </li><li>固定液（4％甲醛1X PBS） </li><li> PBSTx（用含0.1％TRITON - X - 100的） </li><li>适当稀释的小学和物种特异性荧光标记PBSTx中学的抗体</li></ol></li><li>解剖果蝇暴露心脏管（成人苍蝇和/或幼虫） <ol class="lalp...

<ol>
	<li>Brent, J. R., Werner, K. M., McCabe, B. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+larval+NMJ+dissection.">Drosophila larval NMJ dissection.</a> J Vis Exp. , (2009).</li><li>Molina, M. R., Cripps, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ostia,+the+inflow+tracts+of+the+Drosophila+heart,+develop+from+a+genetically+distinct+subset+of+cardial+cells.">Ostia, the inflow tracts of the Drosophila heart, develop from a genetically distinct subset of cardial cells.</a> Mech Dev. 1, 51-59 (2001).</li><li>Dulcis, D., Levine, R. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glutamatergic+innervation+of+the+heart+initiates+retrograde+contractions+in+adult+Drosophila+melanogaster.">Glutamatergic innervation of the heart initiates retrograde contractions in adult Drosophila melanogaster.</a> J Neurosci. 2, 271-280 (2005).</li><li>Zeitouni, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Signalling+pathways+involved+in+adult+heart+formation+revealed+by+gene+expression+profiling+in+Drosophila.">Signalling pathways involved in adult heart formation revealed by gene expression profiling in Drosophila.</a> PLoS Genet. 10, 1907-1921 (2007).</li><li>Robertson, C. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+metamorphosis+of+Drosophila+melanogaster,+including+an+accurately+timed+account+of+the+principal+morphological+change.">The metamorphosis of Drosophila melanogaster, including an accurately timed account of the principal morphological change.</a> J. Morphol. 2, 351-399 (1936).</li><li>Miller, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The internal anatomy and histology of the imago of Drosophila melanogaster. , (1950).</li><li>Rizki, T. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+circulatory+system+and+associated+cells+and+tissues.">The circulatory system and associated cells and tissues.</a> , (1978).</li><li>Cammarato, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Myosin+transducer+mutations+differentially+affect+motor+function,+myofibril+structure,+and+the+performance+of+skeletal+and+cardiac+muscles.">Myosin transducer mutations differentially affect motor function, myofibril structure, and the performance of skeletal and cardiac muscles.</a> Mol Biol Cell. 2, 553-562 (2008).</li><li>Taghli-Lamallem, O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dystrophin+deficiency+in+Drosophila+reduces+lifespan+and+causes+a+dilated+cardiomyopathy+phenotype.">Dystrophin deficiency in Drosophila reduces lifespan and causes a dilated cardiomyopathy phenotype.</a> Aging Cell. 2, 237-249 (2008).</li><li>Kelso, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=a+database+documenting+a+GFP+protein-trap+insertion+screen+in+Drosophila+melanogaster.">a database documenting a GFP protein-trap insertion screen in Drosophila melanogaster.</a> Nucleic Acids Res. 32, D418-D420 (2004).</li><li>Dietzl, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+genome-wide+transgenic+RNAi+library+for+conditional+gene+inactivation+in+Drosophila.">A genome-wide transgenic RNAi library for conditional gene inactivation in Drosophila.</a> Nature. 7150, 151-156 (2007).</li><li>Mery, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Drosophila+muscle+LIM+protein,+Mlp84B,+is+essential+for+cardiac+function.">The Drosophila muscle LIM protein, Mlp84B, is essential for cardiac function.</a> J Exp Biol. 211, 15-23 (2008).</li></ol>

在这里，我们提出了一个协议， 准备和染色果蝇背血管和组织通过荧光或共聚焦显微镜成像。我们提供一个简洁的细化和有效的染色， 允许在幼虫和成虫果蝇心管原位成像解决通常在我们的实验室工作的步骤帐户。其他缩写形式2，3，4所描述的类似的方法。 实施染色过程时，重要的额外细节需要考虑。例如，与其他生物材料中发现，相当<...

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<td>Ethylene glycol-bis (2-amino-ethylether) -N,N,N&#x2019;,N&#x2019;-tetra-acetic acid (EGTA)</td>
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<td>Formaldehyde, 10%, methanol free, Ultra Pure</td>
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<td>Triton-X-100</td>
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<td>Sigma-Aldrich&reg;</td>
<td>9002-93-1</td>
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<td>Alexa Fluor&reg; 594 phalloidin</td>
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fluorescent labeling of drosophila heart structures

“<em&gt;果蝇</em&gt;背船只，或心脏，是一个收缩的心肌，心包细胞沿心墙，支持alary肌肉和在成人，腹纵肌细胞层两侧排列的单层组成的管状结构。收缩纤维的房子保守的肌肉细胞结构成分，包括肌原纤维和细胞骨架/ submembranous蛋白复合物，与同源的细胞外基质的组成部分相互作用束密密麻麻。在这里，我们描述了从解剖的幼虫和半完整的成人心中的一个固定的协议，特别是心肌元素的荧光标记<em&gt;果蝇</em&gt;。具体来说，我们表现出的肌节的F -肌动蛋白和α-辅肌动蛋白在幼虫心中的标签。此外，我们执行标签的F -肌动蛋白和α-辅肌动蛋白，肌球蛋白GFP表达成蝇和α-辅肌动蛋白和pericardin，IV型细胞外基质的胶原蛋白，在野生型成人的心。特别注意的是，以一个半完整的成人心中的安装策略，最大限度地减少了处理和优化的机会，为维护心管及相关组织的完整性。这些准备工作都适合通过荧光和共聚焦显微镜成像。总体而言，这个程序允许的心从一个强大的基因听话的模型系统的结构特点周密细致的分析。

Watch this Scientific Journal Video about Fluorescent Labeling of Drosophila Heart Structures at JoVE.com

Fluorescent Labeling of Drosophila Heart Structures

在这里，我们描述了从幼虫和成虫心管的不同元素的荧光标记的一个基本协议<em&gt;果蝇</em&gt;。这些标本非常适合通过荧光或共聚焦显微镜成像。这种技术允许一个强大的模式生物的心功能的详细的结构分析。

荧光标记的果蝇心结构

The Drosophila melanogaster dorsal vessel, or heart, is a tubular structure comprised of a single layer of contractile cardiomyocytes, pericardial cells ...

Research

JoVE Journal

Biology

Fluorescent Labeling of Drosophila Heart Structures

Labeling Stem Cells with Fluorescent Dyes for non-invasive Detection with Optical Imaging

与非侵入性检测荧光染料标记的干细胞与光学成像

Optical imaging (OI) is an easy, fast and inexpensive tool for in vivo monitoring of new stem cell based therapies. The technique is based on ex vivo ...

科研

生物学

Visualizing the Live Drosophila Glial-neuromuscular Junction with Fluorescent Dyes

Visualizing the Live Drosophila Glial-neuromuscular Junction with Fluorescent Dyes

可视化的Live果蝇胶质细胞神经​​肌肉交界处，与荧光染料

Our project identified GFP labeled glial structures at the developing larval fly neuromuscular synapse. To look at development of live glial-nerve-muscle ...

Visualizing the Beating Heart in Drosophila

Visualizing the Beating Heart in Drosophila

可视化心脏跳动果蝇</em

The Drosophila heart has recently emerged as a good model system for examining the genetic, cellular, and molecular mechanisms underlying function in ...

Monitoring Heart Function in Larval Drosophila melanogaster for Physiological Studies

Monitoring Heart Function in Larval Drosophila melanogaster for Physiological Studies

在幼虫的监测心脏功能果蝇生理研究

We present various methods to record cardiac function in the larval Drosophila. The approaches allow heart rate to be measured in unrestrained  and  ...

Preparation of embryos for Electron Microscopy of the Drosophila embryonic heart tube

Preparation of embryos for Electron Microscopy of the Drosophila embryonic heart tube

电子显微镜对胚胎的制备果蝇胚胎心管

The morphogenesis of the Drosophila  embryonic heart tube has emerged as a valuable model system for studying cell migration, cell-cell adhesion and cell ...

Preparation of Drosophila Polytene Chromosome Squashes for Antibody Labeling

果蝇聚乙烯南瓜抗体标记染色体的制备

Drosophila has long been a favorite model system for studying the relationship between chromatin structure and gene regulation due to the cytological ...

Fluorescent Labeling of COS-7 Expressing SNAP-tag Fusion Proteins for Live Cell Imaging

对活细胞成像的SNAP - Tag融合蛋白的荧光标记的COS - 7

SNAP-tag and CLIP-tag protein labeling systems enable the specific, covalent attachment of molecules, including fluorescent dyes, to a protein of interest ...

Whole Mount RNA Fluorescent in situ Hybridization of Drosophila Embryos

Whole Mount RNA Fluorescent in situ Hybridization of Drosophila Embryos

全山RNA荧光<em在原位</em&gt;杂交<em&gt;果蝇</em&gt;胚胎

Assessing  the expression pattern of a gene, as well as the subcellular localization  properties of its transcribed RNA, are key features for ...

Intravital Microscopy for Imaging Subcellular Structures in Live Mice Expressing Fluorescent Proteins

活体显微镜成像的亚细胞结构的活体小鼠表达荧光蛋白

Here we describe a procedure to image subcellular structures in live rodents that is based on the use of confocal intravital microscopy. As a model organ, ...

Drosophila Preparation and Longitudinal Imaging of Heart Function In Vivo Using Optical Coherence Microscopy (OCM)

Drosophila Preparation and Longitudinal Imaging of Heart Function In Vivo Using Optical Coherence Microscopy OCM

果蝇的制备及心功能体内使用光学相干显微镜成像纵向（OCM）

Longitudinal study of the heartbeat in small animals contributes to understanding structural and functional changes during heart development. Optical ...