Name: long-term live imaging of drosophila eye disc
Uploaded: 2017-05-06T14:00:00.000+00:00
Duration: 6 min 8 s
Description: Watch this Scientific Journal Video about Long-term Live Imaging of Drosophila Eye Disc at JoVE.com

我们感谢春兰Hsu和宇奇·扬准备飞食品和维护飞股票，并以苏平·李和IMB成像核心为他们与激光共聚焦显微镜的帮助。这项研究是由赠款YHS（NSC 101-2321-B-001 -004，NSC 100-2321-B-001 -012，NSC 102-2321-B-001 -002，MOST 103-2311-B-001支持-035 -MY3）来自美国国家科学理事会和共和国的中国科学技术部。

1.预实验装置<ol><li>收集，根据mδ0.5-GAL4核表达绿色荧光蛋白（GFP）的苍蝇卵和文化，在25℃下5天。 </li><li>微波琼脂糖在10毫升1×磷酸盐缓冲盐水（PBS），直到琼脂糖完全溶解0.7克低熔点的。保持0.7％低熔点凝胶在37℃培养箱中，直到使用。 </li><li>所有消毒设备，包括镊子，剪刀42毫米×0.17毫米盖玻片，9孔玻璃斑点板和磁培养室，用70％乙醇超过12小时。 </li><li>制备培养基3:补充有2％胎牛血清（FBS），0.5％青霉素-链霉素，和1.25 mg / mL的胰岛素施耐德的果蝇介质。为了避免污染，在制备细胞培养罩平台。存储所制备的培养基在4℃下，并在一个月内使用。 </li><li>清洁清扫平台和实验者"s的70％乙醇的手开始解剖前。 </li></ol> 2.光盘解剖<ol><li>空气从70％乙醇擦干所有的设备。 </li><li>选择从飞小瓶第三龄幼虫。为了避免飞瓶污染，洗1X PBS 3倍的幼虫以去除浪费。 </li><li>解剖解剖显微镜下在1mL在9孔玻璃斑点平板培养基（在RT）的清洁的幼虫。 </li><li> 掌握具有一对钳子的幼虫在约三分之一的从后端的方式和在嘴钩另一个钳。拉两个镊子在相反方向上，以释放内部组织。 <ol><li>从连接到嘴钩眼脑复杂取出唾液腺，脂肪体，和角质层。如果腹神经索仍然连接，用剪刀剪它拿走。 </li></ol></li><li>使用一对钳子的把握嘴钩和一对钳子的，以除去组织片的COnnects背侧部分大脑和眼睛光盘。 注意:如果这个组织不除，两个眼睛触角光盘将接近对方和大脑。眼睛光盘随后不会盖玻片上平躺，因为此组织，脑，和眼触角光盘将形成一个三角形。 </li><li> 旋转整个复杂的腹朝上的位置。卸下连接钩或光盘大脑中的灯丝。 注意:如果灯丝不被除去，当口钩被移除眼触角光盘将分成两片。 <ol><li>取下嘴钩。小心不要过程中损坏眼睛光盘。 注:眼睛光盘是现在在介质中自由和仅由光学秆连接到大脑。 </li></ol></li></ol> 3.安装<ol><li>空气干燥并用一台42毫米×0.17毫米盖玻片组装腔室的部件。附加的O形环构成的双层以盖玻片以保持第中心Ë琼脂糖。 </li><li>组装盖玻片室。把42毫米×0.17毫米盖玻片在舞台上的受体。放置在盖玻片硅O形环。将基地。锁定密封件锁定件和放置盖（ 图1）。 </li><li>使用20微升微量具有10-200μL尖端解剖眼脑复杂仔细转移到O形环的中心。 </li><li> 除去大部分的培养基中，并添加0.7％低熔点琼脂糖的12μL（在37℃）到样品中。 注意:眼睛光盘的位置可以用钳子琼脂糖固化之前被改变。琼脂糖将在5分钟内固化。 <ol><li>加入1 ml培养基在室温下的O形环的中心;每个O形环可以加载多达4个样品。确保琼脂糖粘在O型圈，以避免样品的漂移。 注:介质的无曝气或流通是必要的。 </li></ol></li></ol>4.共聚焦显微镜<ol><li>放置在倒置的共聚焦显微镜的载物台的腔室30分钟以使温度平衡。这可以在成像期间防止Z轴漂移。 </li><li>长期活体成像之前，请检查是否组织是微分干涉对比显微镜完好。 </li><li>为了避免光毒性，使用下面2毫瓦的激光功率。使用40X物镜成像。 </li><li>以1μm的光学间隔获取在12分钟内62微米Z堆叠图像。获得扫描的40个循环以上共计10小时。确保每个周期包含成像的12分钟和休息3分钟。 </li></ol>

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作者宣称，他们没有竞争的经济利益。

在这项研究中，我们提供了对果蝇成虫盘长期实时成像试验的详细方案。我们花了很多时间练习了仔细解剖，避免盘片损坏，并允许盘接近盖玻片的附件。我们试图聚-L-赖氨酸（PLL）和低熔点琼脂糖以保持组织;在年底，低熔点琼脂糖表现出较好的能力，以保持组织。尽管在本文中仅使用眼睛光盘以证明在10 H实时成像期间感光体分化和网膜旋转的正常发生，该方法也可以应用到至少一个其它盘时，翼盘，如在证实以前的研究3。此外，培养基的制备和实时成像的设置非常简单，不需要充气或介质循环。 连续实时成像可提供生物前夕的一个明确的时间序列NTS。在我们前面的神经胶质细胞分化和迁移的眼睛光盘的报告中，我们提供了直接的证据证明神经胶质包裹从现有的神经胶质细胞分化迁移到眼盘3的后前位。长期体外培养允许细胞，如枫荧光蛋白的光转换，具体激光活化标记按照它们的行为3。它也可以容纳被直接加入到培养基中的化学品的测试;因此，它可以被用来作为药物筛选平台。由于一些动态过程出现几分钟或几秒钟之内，需要高的时间分辨率。为了提高时间分辨率，光片显微镜15,16或旋转盘显微镜17可以是一个好的选择。 目前的培养条件是不完美的。它不支持光盘的增长。小区proliferation仅支持多达12小时3。在体外培养12个小时后，感光体分化的速率开始降低3。这可能是由于缺乏某种营养素或激素。由于在该培养基中的主要差别是胰岛素的高浓度，所述哺乳动物的胰岛素可以是部分模拟内源性胰岛素样肽的功能。加入蝇提取物，昆虫血糖海藻糖和蜕皮激素20-羟基的各种浓度的，不利于3。然而，在12-18小时培养期是足够的研究许多发育过程。培养幼虫成虫盘替代培养方法进行了比较3，我们的培养和拍摄条件提供实时成像时间最长的窗口和最清晰。虽然内活幼虫和蛹光盘可以直接成像18，19，20，21，22，用于观测分辨率和时间窗口是有限的。晚幼虫和蛹的碟片可以很长一段时间23,24进行培养，但是这些光盘进行显著形态。采取平面的2D幼虫盘的优点，我们的培养和成像方法是最好的解决方案为止。

果蝇幼虫成虫盘已经为各种各样的生物机制研究的青睐实验系统。这些光盘从胚胎上皮invaginate和成为由两个上皮层构成囊状结构，Peripodial上皮（PE）和光盘正确（DP）1,2。在成虫盘细胞增殖并在幼虫和蛹的阶段逐步分化，并最终发育成成年体结构。由于成虫盘的平坦和简单的双层结构，它们很容易从幼虫解剖时观察到。然而，尽管许多团体多次努力，为成虫盘的长期培养的现有方法都不令人满意。我们最近开发了能够承受多次成虫盘的正常发展长达18个小时的培养方法，并允许实时成像3 </s了&gt;。培养方法可以支持细胞分化和迁移，但是它可以支持细胞增殖为仅7-12小时，并不能支持盘的增长。在培养基中的主要差别是胰岛素3的高浓度。该方法操作简单，并且不需要特殊的通风或介质循环。 其中研究最多的成虫盘的是眼睛，触角光盘。 果蝇眼睛建成约800小眼。每个小眼是由8个感光细胞（R1-R8）的。在幼虫眼睛光盘，感光细胞分化后的形态发生沟（MF），其跨过眼睛光盘扫过从后部的通道到前4,5。在一个小眼的感光体分化在序列，R8，随后R2 / R5，R3 / R4，R1 / R6和R7 6开始。在背的小眼和五个眼睛盘的ntral半部经受在相反方向上旋转90°，从而产生相反手7,8。旋转过程发生在两个阶段:首先，一个45°旋转开始于并在后面MF第六小眼行被完成;第二45°旋转是在完成约16排9,10。本研究中使用旋转网膜，由R3 / 4特异性mδ0.5-GA4 11标记，以证明正常细胞分化和动力学可以通过该方法观察到培养和实时图像的眼睛光盘。

<table><tbody><tr><td>Low-melting agarose</td><td>AMRESCO&amp;nbsp;</td><td>0815-25G</td><td></td></tr><tr><td>Phosphate-buffered Saline (PBS)</td><td>AMRESCO</td><td>0780-2PK</td><td></td></tr><tr><td>42 mm x 0.17 mm cover slips&amp;nbsp;</td><td>PST</td><td>G401-42</td><td></td></tr><tr><td>Schneider&amp;rsquo;s &lt;em&gt;Drosophila&lt;/em&gt; medium&amp;nbsp;</td><td>Thermo</td><td>21720-024</td><td></td></tr><tr><td>Fetal Bovine Serum (FBS)</td><td>Sigma</td><td>F9665</td><td></td></tr><tr><td>Penicillin-streptomycin&amp;nbsp;</td><td>GIBCO</td><td>759</td><td></td></tr><tr><td>Insulin</td><td>Sigma</td><td>I9278</td><td></td></tr><tr><td>Culture chamber&amp;nbsp;</td><td>PECON</td><td></td><td>POC-R2 Cell Cultivation System</td></tr><tr><td>40X objective</td><td></td><td></td><td>C-Apochromat 40X/1.2W Korr objective&amp;nbsp;</td></tr><tr><td>Fly strain:nls.GFP</td><td>Bloominggton</td><td>4775</td><td></td></tr></tbody></table>

long-term live imaging of drosophila eye disc

实时成像提供了实时连续跟踪动态的细胞和发育过程的能力。 果蝇幼虫成虫盘已经被用于研究许多生物学过程，诸如细胞增殖，分化，生长，迁移，细胞凋亡，竞争，细胞-细胞信号传导和房室边界形成。但是，对于长期体外培养和成虫盘的实时成像方法尚未尽管作了许多努力是令人满意的。最近，我们制定了长期体外培养和长达18小时成虫盘的实时成像的方法。除了在培养基中使用高浓度的胰岛素，一种低熔点琼脂糖也被用来嵌入该盘，以防止它在成像期间漂移。本报告采用的眼睛触角光盘作为一个例子。感光体R3 / 4特异性mδ0.5-GA4表达随后证明感光differen可以在一个10 H活体成像期间可以观察到tiation网膜和旋转。这是描述这种简单的方法的详细协议。

<html>在这个例子中，R3 / R4感光体用mδ0.5-GA4标记来监控分化感光体。 mδ0.5-GA4驱动在R3 11 R4和强表达表达较弱，使其成为R3和R4优异的标记物，以及用于旋转网膜的过程。在电影1，R3和R4分别GFP标记。在10 H实时成像会话的开始，有8行簇网膜（ 图2A和电影1），并在结束时（ 图 2B）的行14网膜簇的。分化网膜的速率为每行1.67小时，这是接近1.5-2 H /行的速率在体内圆盘7，8，9，10，11，S = "外部参照"&gt; 12，13，14。根据R3和R4的相对位置，网膜旋转期间实时成像（ 图2A" A‘’，B"，和B‘’）发生在离体培养的光盘。单个R3 / R4簇标记有白色球体在Movie 2的第一帧。在开始时，R3 / R4轴似乎在垂直于赤道（ 图2C和电影2）。然后，它似乎在3（ 图2C &#39;），6（ 图2C &#39;&#39;）旋转15°，30°，和45°，和图9（ 图2C &#39;&#39;&#39;）H，分别。这个数据表明，该方法可以在10 H实时成像期间维持感光体分化。 <img alt="图1" src="/files/ftp_upload/55748/55748fig1.jpg" /> Figu重1:商会系统的示意图。腔室组件的示意图:（1）将双层O形圈连接的盖玻片在舞台上的受体。 （2）将盖玻片上的硅O形环。 （3）将碱。 （4）锁定在密封储物柜。 （5）加载在O形环的中心的样品并放置盖。 <a href="http://ecsource.jove.com/files/ftp_upload/55748/55748fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/55748/55748fig2.jpg" /> 图2:R3 / R4分化和旋转的实时成像。 R3 / R4标记mδ0.5-Gal4的期间的实时成像10个H表达核型GFP。所有这些数字是从电影1抓获。在开始拍摄（A）图像。 （ <strong<html> &gt; A&#39;-A"）的图像从左侧（A"）和右（A方框区域放大"）。 （B）图像在10小时期间的结束作出。 （B&#39;-B"）的图像从左侧（B"）和右（B方框区域放大"）。 （CC"""）A单一R3 / R4对（通过在箭头标记"）之后通过的时间，以显示R3 / R4对的旋转。比例尺=15μm时，为10μm，而在A，B，和C为5μm，分别。所有附图是最大强度的z突起。的时间尺度表示在H:分钟。 <a href="http://ecsource.jove.com/files/ftp_upload/55748/55748fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="电影1" src="/files/ftp_upload/55748/55748mov1.jpg" /> 电影1:rce.jove.com/files/ftp_upload/55748/Movie_1.avi"目标=‘_空白’&gt;请点击这里下载这部电影。 <img alt="电影2" src="/files/ftp_upload/55748/55748mov2.jpg" /> 电影1: <a href="http://ecsource.jove.com/files/ftp_upload/55748/Movie_2.avi" target="_blank">请点击这里下载这部电影。</a>

Watch this Scientific Journal Video about Long-term Live Imaging of Drosophila Eye Disc at JoVE.com

Long-term Live Imaging of Drosophila Eye Disc

这个协议描述了用于长期体外培养和果蝇成虫盘的实时成像的详细方法。它表明的眼睛光盘的实时成像的10小时期间内感光体分化网膜和旋转。该协议是简单，不需要昂贵的安装。

长期的实时成像<em&gt;果蝇</em&gt;眼睛光盘

Live imaging provides the ability to continuously track dynamic cellular and developmental processes in real time. Drosophila larval imaginal discs have ...

long-term-live-imaging-of-drosophila-eye-disc

Research

JoVE Journal

Developmental Biology

Long-term Live Imaging of Drosophila Eye Disc

8.4K 次观看.  National Yang-Ming University. 这个协议描述了用于长期体外培养和果蝇成虫盘的实时成像的详细方法。它表明的眼睛光盘的实时成像的10小时期间内感光体分化网膜和旋转。该协议是简单，不需要昂贵的安装。 

视频: Long-term Live Imaging of Drosophila Eye Disc

Live-imaging of the Drosophila Pupal Eye

Inherent processes of Drosophila pupal development can shift and distort the eye epithelium in ways that make individual cell behavior difficult to track during live cell imaging. These processes include: retinal rotation, cell growth and organismal movement. Additionally, irregularities in the topology of the epithelium, including subtle bumps and folds often introduced as the pupa is prepared for imaging, make it challenging to acquire in-focus images of more than a few ommatidia in a single focal plane. The workflow outlined here remedies these issues, allowing easy analysis of cellular processes during Drosophila pupal eye development. Appropriately-staged pupae are arranged in an imaging rig that can be easily assembled in most laboratories. Ubiquitin-DE-Cadherin:GFP and GMR-GAL4&#8211;driven UAS-&#945;-catenin:GFP are used to visualize cell boundaries in the eye epithelium 1-3. After deconvolution is applied to fluorescent images captured at multiple focal planes, maximum projection images are generated for each time point and enhanced using image editing software. Alignment algorithms are used to quickly stabilize superfluous motion, making individual cell behavior easier to track.

Live-imaging of the Drosophila Pupal Eye

This protocol presents an efficient method for imaging the live Drosophila pupal eye neuroepithelium. This method compensates for tissue movement and uneven topology, enhances visualization of cell boundaries through the use of multiple GFP-tagged junction proteins, and uses an easily-assembled imaging rig.

实时成像<em&gt;果蝇</em&gt;蛹眼

Inherent processes of Drosophila pupal development can shift and distort the eye epithelium in ways that make individual cell behavior difficult to track ...

科研

发育生物学

Live Imaging of Glial Cell Migration in the Drosophila Eye Imaginal Disc

Glial cells of both vertebrate and invertebrate organisms must migrate to final target regions in order to ensheath and support associated neurons. While recent progress has been made to describe the live migration of glial cells in the developing pupal wing (1), studies of Drosophila glial cell migration have typically involved the examination of fixed tissue. Live microscopic analysis of motile cells offers the ability to examine cellular behavior throughout the migratory process, including determining the rate of and changes in direction of growth. Paired with use of genetic tools, live imaging can be used to determine more precise roles for specific genes in the process of development. Previous work by Silies et al. (2) has described the migration of glia originating from the optic stalk, a structure that connects the developing eye and brain, into the eye imaginal disc in fixed tissue. Here we outline a protocol for examining the live migration of glial cells into the Drosophila eye imaginal disc. We take advantage of a Drosophila line that expresses GFP in developing glia to follow glial cell progression in wild type and in mutant animals.

Here we describe a protocol to examine the migration of glial cells into the developing Drosophila eye using live microscopic analysis paired with GFP tagged glial cells.

住在果蝇眼睛成虫光盘胶质细胞迁移成像

Glial cells of both vertebrate and invertebrate organisms must migrate to final target regions in order to ensheath and support associated neurons.  While ...

生物学

A Novel RFP Reporter to Aid in the Visualization of the Eye Imaginal Disc in Drosophila

The Drosophila eye is a powerful model system for studying areas such as neurogenesis, signal transduction and neurodegeneration. Many of the discoveries made using this system have taken advantage of the spatiotemporal nature of photoreceptor differentiation in the developing eye imaginal disc. To use this system it is first necessary for the researcher to learn to identify and dissect the eye disc. We describe a novel RFP reporter to aid in the identification of the eye disc and the visualization of specific cell types in the developing eye. We detail a methodology for dissection of the eye imaginal disc from third instar larvae and describe how the eye-RFP reporter can aid in this dissection. This eye-RFP reporter is only expressed in the eye and can be visualized using fluorescence microscopy either in live tissue or after fixation without the need for signal amplification. We also show how this reporter can be used to identify specific cells types within the eye disc. This protocol and the use of the eye-RFP reporter will aid researchers using the Drosophila eye to address fundamentally important biological questions.

A Novel RFP Reporter to Aid in the Visualization of the Eye Imaginal Disc in Drosophila

We describe a novel red fluorescent protein (RFP) reporter that is expressed specifically in the Drosophila eye. We detail a methodology for dissection of the eye imaginal disc and how this reporter can be used to aid in the dissection and identification of specific cell types in the developing eye.

一种新型的RFP的记者，以帮助在眼成虫光盘的可视化果蝇</em

The Drosophila eye is a powerful model system for studying areas such as neurogenesis, signal transduction and neurodegeneration. Many of the discoveries ...

Preparation of Developing and Adult Drosophila Brains and Retinae for Live Imaging

The Drosophila brain and visual system are widely utilized model systems to study neuronal development, function and degeneration. Here we show three preparations of the brain and visual system that cover the range from the developing eye disc-brain complex in the developing pupae to individual eye and brain dissection from adult flies. All protocols are optimized for the live culture of the preparations. However, we also present the conditions for fixed tissue immunohistochemistry where applicable. Finally, we show live imaging conditions for these preparations using conventional and resonant 4D confocal live imaging in a perfusion chamber. Together, these protocols provide a basis for live imaging on different time scales ranging from functional intracellular assays on the scale of minutes to developmental or degenerative processes on the scale of many hours.

Preparation of Developing and Adult Drosophila Brains and Retinae for Live Imaging

This protocol describes three Drosophila preparations: 1) adult brain dissection, 2) adult retina dissection and 3) developing eye disc- brain complexes dissection. Emphasis is laid on special preparation techniques and conditions for live imaging, although all preparations can be used for fixed tissue immunohistochemistry.

发展和成人的制备果蝇大脑和视网膜实时成像

The Drosophila brain and visual system are widely utilized model systems to study neuronal development, function and degeneration. Here we show three ...

Real-Time Imaging of Germline Stem Cells in Drosophila Ex Vivo Germaria Culture

Extended Live Imaging of Female Drosophila melanogaster Germline Stem Cell Niches

Live imaging methods allow the analysis of dynamic cellular processes in detail and in real-time. The Drosophila ovary represents an excellent model to explore the dynamics of a myriad of developmental processes, such as cell division, stemness, differentiation, migration, apoptosis, autophagy, cellular adhesion, etc., over time. Recently, we have implemented an extended ex vivo culture and live imaging of the female Drosophila GSC niche. Using a Drosophila line harboring a GFP::Par-1 transgene as an example, this method allows the visualization of the GSCs&#39; asymmetric division within their niche and the description of the changes in the spectrosome morphology along the cell cycle. Here, we present a detailed protocol for the ex vivo culture of Drosophila germaria, enabling prolonged visualization of the female GSC niche. Importantly, this protocol is broadly applicable to live imaging GSCs with multiple fluorescently tagged proteins of interest that are available in stock centers and/or in the Drosophila research community.

Here, we provide a detailed protocol to perform live imaging of the asymmetric division of germline stem cells (GSCs) in the Drosophila ovarian niche. We use a transgenic line that ubiquitously expresses a green fluorescent protein (GFP) fusion of the spectrosome protein Par-1.

Live imaging methods allow the analysis of dynamic cellular processes in detail and in real-time. The Drosophila ovary represents an excellent model to ...

<meta charset="utf8"></head><body>无蛹的 果蝇 Pupal 腿部形态发生的扩展实时成像

Long-Term Live Imaging of Drosophila Pupal Leg Development After Puparium Removal

Over the past decades, significant progress has been made in understanding the mechanisms of cell fate determination. However, the process by which fate-determined cells form three-dimensional organismal shapes remains unclear. Recent advances in confocal microscopy have facilitated efforts to observe cell dynamics during development through live imaging. The Drosophila melanogaster pupa is ideal for live imaging due to its immobility, transparent pupal cuticle, and the availability of fluorescent reporter lines. A primary challenge for imaging is the puparium, the cuticle surrounding the pupa, which obstructs optical imaging. While previous methods involved either partial or complete removal of the puparium, maintaining pupal viability for extended periods after this procedure has remained challenging. Here, a simple method is presented for days-long live imaging of the Drosophila leg during the pupal stage, involving complete puparium removal. The method includes removing the puparium from a pupa adhered to double-sided tape, followed by assembling a small chamber on a glass-bottom dish to enclose the pupa and a drop of water. This setup is straightforward, reliable, and supports extended pupal survival by preventing desiccation. Long-term live imaging of the Drosophila pupa has significantly contributed to capturing how the adult leg undergoes dramatic three-dimensional structural changes over 2-3 days. These changes include the transient formation of an intriguing structure (the Parthenon-like structure) by epithelial cells, rapid tissue narrowing, joint formation, and bristle elongation. This method is applicable to the observation of various tissues and can potentially be combined with other techniques, such as optical gene induction, to advance the understanding of cell dynamics during the final shape formation of tissues in the pupal stage.

<meta charset="utf8"></head><body>作者聚焦：揭示 果蝇 发育过程中的动态上皮结构

Here, a protocol is presented for long-term live imaging of the Drosophila leg during the pupal stage, involving complete removal of the puparium. This protocol can also be applied to other tissues.

果 蝇 蛹切除后蛹腿部发育的长期实时成像

Over the past decades, significant progress has been made in understanding the mechanisms of cell fate determination. However, the process by which ...

Live Imaging of GFP-labeled Proteins in Drosophila Oocytes

The Drosophila oocyte has been established as a versatile system for investigating fundamental questions such as cytoskeletal function, cell organization, and organelle structure and function. The availability of various GFP-tagged proteins means that many cellular processes can be monitored in living cells over the course of minutes or hours, and using this technique, processes such as RNP transport, epithelial morphogenesis, and tissue remodeling have been described in great detail in Drosophila oocytes1,2.
The ability to perform video imaging combined with a rich repertoire of mutants allows an enormous variety of genes and processes to be examined in incredible detail. One such example is the process of ooplasmic streaming, which initiates at mid-oogenesis3,4. This vigorous movement of cytoplasmic vesicles is microtubule and kinesin-dependent5 and provides a useful system for investigating cytoskeleton function at these stages.
Here I present a protocol for time lapse imaging of living oocytes using virtually any confocal microscopy setup.

Live Imaging of GFP-labeled Proteins in Drosophila Oocytes

A protocol for live imaging of GFP-tagged proteins or autofluorescent structures in individual Drosophila oocytes is described.

GFP标记的蛋白质在实时成像<em&gt;果蝇</em&gt;卵母细胞

The  Drosophila oocyte has been established as a versatile system for investigating  fundamental questions such as cytoskeletal function, cell ...

The Tomato/GFP-FLP/FRT Method for Live Imaging of Mosaic Adult Drosophila Photoreceptor Cells

The Drosophila eye is widely used as a model for studies of development and neuronal degeneration. With the powerful mitotic recombination technique, elegant genetic screens based on clonal analysis have led to the identification of signaling pathways involved in eye development and photoreceptor (PR) differentiation at larval stages. We describe here the Tomato/GFP-FLP/FRT method, which can be used for rapid clonal analysis in the eye of living adult Drosophila. Fluorescent photoreceptor cells are imaged with the cornea neutralization technique, on retinas with mosaic clones generated by flipase-mediated recombination. This method has several major advantages over classical histological sectioning of the retina: it can be used for high-throughput screening and has proved an effective method for identifying the factors regulating PR survival and function. It can be used for kinetic analyses of PR degeneration in the same living animal over several weeks, to demonstrate the requirement for specific genes for PR survival or function in the adult fly. This method is also useful for addressing cell autonomy issues in developmental mutants, such as those in which the establishment of planar cell polarity is affected.

The Tomato/GFP-FLP/FRT Method for Live Imaging of Mosaic Adult Drosophila Photoreceptor Cells

The Tomato/GFP-FLP/FRT method involves visualizing mosaic photoreceptor cells in living Drosophila. It can be used to follow individual photoreceptor cell fates in the retina for days or weeks. This method is ideal for studies of retinal degeneration and neurodegenerative diseases or photoreceptor cell development.

番茄/ GFP-FLP / FRT方法马赛克成人的实时成像<em&gt;果蝇</em&gt;感光细胞

The Drosophila eye is widely used as a model  for studies of development and neuronal degeneration. With the powerful mitotic  recombination technique, ...

长期的实时成像

摘要

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此视频中的章节

实时成像

住在果蝇眼睛成虫光盘胶质细胞迁移成像

一种新型的RFP的记者，以帮助在眼成虫光盘的可视化果蝇

发展和成人的制备果蝇大脑和视网膜实时成像

Extended Live Imaging of Female Drosophila melanogaster Germline Stem Cell Niches

果蝇蛹切除后蛹腿部发育的长期实时成像

果蝇蛹切除后蛹腿部发育的长期实时成像

果蝇蛹切除后蛹腿部发育的长期实时成像

GFP标记的蛋白质在实时成像

番茄/ GFP-FLP / FRT方法马赛克成人的实时成像

长期的实时成像

摘要

探索更多视频

此视频中的章节

实时成像

住在果蝇眼睛成虫光盘胶质细胞迁移成像

一种新型的RFP的记者，以帮助在眼成虫光盘的可视化果蝇

发展和成人的制备果蝇大脑和视网膜实时成像

Extended Live Imaging of Female Drosophila melanogaster Germline Stem Cell Niches

果 蝇 蛹切除后蛹腿部发育的长期实时成像

果 蝇 蛹切除后蛹腿部发育的长期实时成像

果 蝇 蛹切除后蛹腿部发育的长期实时成像

GFP标记的蛋白质在实时成像

番茄/ GFP-FLP / FRT方法马赛克成人的实时成像

果蝇蛹切除后蛹腿部发育的长期实时成像

果蝇蛹切除后蛹腿部发育的长期实时成像

果蝇蛹切除后蛹腿部发育的长期实时成像