免疫组化标记在感觉神经元树突的微管，气管，并在肌肉果蝇幼虫体壁
Summary
要理解的复杂的细胞形状，如神经细胞树突，如何在发展过程中取得的，重要的是要能够准确地检测微管组织。在这里，我们描述了一个强大的免疫组化标记法研究微管组织的感觉神经元树突树突分枝，气管，肌肉，及其他<em>果蝇</em>幼虫体壁组织。
Abstract
要了解如何实现复杂的细胞形态的差异，重要的是要准确地按照微管组织。 果蝇幼虫体壁含有几种类型的细胞模型来研究细胞和组织的形态发生。例如气管是用于检查1，管形态发生和感觉神经元的果蝇幼虫一般的澄清和神经树突状分化2-5和变性 6级的具体机制已成为一个主要系统的树突状树枝状（ DA） 。
之间的神经元类，树突分支的形状可以显著不同，甚至在单个神经元的不同分支 7,8中。 DA能神经元的遗传研究表明，差的细胞骨架的组织可以在树突分支的形状4,9-11形态差异的基础。我们提供了一个强大的免疫标记法到ssay 在体内 DA的感觉神经元枝蔓乔木微管组织（图1，2，电影1）。该协议说明了第一龄幼虫的解剖和免疫组化，一个阶段，积极的感觉神经元树突生长和分支组织时发生12,13。
除了染色的感觉神经元，这种方法实现了强大的标签（电影2，3）微管在肌肉组织，气管（图3，电影3），和其他的体壁组织。这是宝贵的希望分析在体壁的微管组织调查机制时，在原地调查控制的组织和细胞的形状。
Protocol
1。试剂的制备 注：在开始之前进行解剖和免疫组化染色在磁场室和幼虫是使用特殊形状的昆虫引脚牵制。磁性室的建设，编制这些引脚的详细说明， 可以发现在14,15相关文献。简单地说，一个1x1cm方孔切成磁性表贴表，使一个小室的后部盖玻片。双方室是密封的环氧树脂胶，这种胶后室是用70％乙醇使用前清洗几次。准备解剖昆虫引脚弯曲所需要的形状，…
Discussion
要理解复杂的细胞形态如何实现它重要的是要能够准确地检测微管组织。在这里，我们描述了一个强大的免疫组化标签方法检测树突状树枝状感觉神经元的树突的微管组织。除了染色的感觉神经元，这种方法实现了强大的气管，肌肉和其他体壁组织的免疫组化染色。
我们使用此协议，研究开发的DA能神经元的感官树突微管组织。这些树突是非常敏感，受伤或应力25，?…
Divulgazioni
The authors have nothing to disclose.
Acknowledgements
我们感谢理化学研究所的资金。 P10 – GAL4阿兰文森特（保罗萨巴蒂尔大学，法国图卢兹）是一种礼物。
Materials
| Name of the reagent | Company |
Catalogue number |
Comments (optional) |
|---|---|---|---|
| Forceps | Dumont | 11251-20 | |
| Microscissors | FST | 15000-08 | |
| Mouse anti-α-tubulin (Clone: DM1A) | Sigma | T9026 | Dilution 1/1000 |
| Mouse anti-Futsch (Clone: 22C10), supernatant |
Developmental Studies Hybridoma Bank |
22C10 | Dilution 1/1000 |
| Rat anti-CD8 (Clone: 5H10) | Caltag | MCD0800 | Dilution 1/1000 |
| Alexa Fluor 488 anti-mouse IgG | Invitrogen | A-11001 | Dilution 1/500 |
| Cy3 anti-Rat IgG | Jackson Immunoresearch | 712-166-150 | Dilution 1/200 |
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Tags
Immunohistological LabelingMicrotubulesSensory Neuron DendritesTracheaeMusclesDrosophila Larva Body WallCell ShapesMicrotubule OrganizationTube MorphogenesisDendritic ArborizationNeuron-class-specific MechanismsDendritic DifferentiationDegenerationCytoskeletal OrganizationImmunological Labeling MethodIn Vivo Microtubule OrganizationFirst Instar LarvaSensory Neuron Dendrite OutgrowthBranching OrganizationMuscles LabelingTrachea Labeling
Citazione di questo articolo
Yalgin, C., Karim, M. R., Moore, A. W. Immunohistological Labeling of Microtubules in Sensory Neuron Dendrites, Tracheae, and Muscles in the Drosophila Larva Body Wall. J. Vis. Exp. (57), e3662, doi:10.3791/3662 (2011).