这项工作是由美国国立卫生研究院支持的授予F32 NS078958（DTB）和R01 AG033620（BG）。

1。组装测试飞行<ol><li>安全飞行缸环站1用链钳。 （留约3厘米的圆柱体重量菜的下面。） （注:飞行汽缸我们用90厘米长，直径13.5厘米）。 </li><li>插入称量皿上一薄层矿物油的飞行筒的下方。 </li><li>安全漏斗环站2使用环钳爪和钳。调整漏斗的高度，使得漏斗的底部是与飞行圆筒的顶部齐平。 （注:该漏斗的前端的直径必须小于放入滴管，使得小瓶不会掉下通过小瓶的外径）。 </li><li>插入落料管进入漏斗的顶部上，并用一个卡爪夹紧。 （注:我们使用滴管即25厘米长滴飞含小瓶从这个高度可以让所有的苍蝇与受力均匀一致弹射落管的内径应为史赖。TLY比小瓶允许小瓶以自由落下的外径大。） </li><li>切割聚丙烯片材（S）到合适的大小。 （注:为了有助于插入和移除的薄片，其宽度应比飞行筒的内周面稍小）。 </li><li>涂抹薄薄的一层缠结陷阱的表。允许使用前坐了1小时。 （注:保留足够的空间，在表的顶部和底部（大约3公分）无涂层把握片插入/删除）。 </li><li>将飞行气缸内的聚丙烯酰胺片。 </li><li>用松木支架组装相机的轨道。 （注:确保轨道的底部可以支持摄像头没有挡住了镜头参见图1B。） </li><li>添加瓶塞和螺旋到位。 （注:发生在位置的限位器，将让相机以查看全景模式下，整个塑料片。） </li></ol> 2。运行试验<ol> &lt;李&gt;收集苍蝇被测试的小瓶。为了获得最佳效果，请使用不超过20苍蝇/瓶。 </li><li>轻轻拍打苍蝇小瓶的底部，拔出，再插入滴管和释放瓶中。 （注:将小瓶落下的落料管，直到它碰到漏斗窄口当小瓶击中漏斗，苍蝇被喷入气缸的飞行。） </li><li>提起落管，以去除空瓶。 （注:苍蝇相同的测试组的多个小瓶可以在一个单一的聚丙烯酰胺片被检测我们发现，高达200苍蝇（10瓶20苍蝇每个）可以用于测试和成像随手在一张纸上。 </li><li>取出塑料片，将它放在一个平坦的白色表面。 （注:白色的海报板可以使用，如果台面是深色。） </li><li>组装相​​机在轨道的塑料片。相机应足够高的片材具有在视场中的片材的顶部和底部之上。 </li><li>沿滑动个摄像头E导轨抱着"捕获"按钮获取全景图像时。 </li><li>苍蝇降落在油的数量可以手动对每个试验进行计数。 </li><li>用于在给定的实验条件下都重复步骤2.2-2.7。苍蝇可以从每次试验之间的片材被移除。可选地，几个薄片可以被使用，以对每个试验的新表。 </li></ol> 3。数据收集<ol><li>使用ImageJ软件打开图像文件。 </li><li>如果需要的话，只包括着陆表面积作物图像。 （这是涂在缠结陷阱的区域。） </li><li>图像转换成8位灰度。 </li><li>创建一个"门槛"，以筛选出的白色背景。 （图像→调整→阈值）。 </li><li>设置参数来识别每个使用"分析粒子"菜单苍蝇。 （分析→分析颗粒）定义用于识别粒子的参数。与我们建立，我们发现使用5-90像素2和0.4-1.0的圆形区域将准确地识别所有的样品。 </li><li>采用坐标为每个粒子的生成的列表中的每个测量苍蝇的位置。的像素的x坐标可以被转换成厘米来计算降落高度。 </li><li>表导入到电子表格（如Microsoft Excel）。 </li></ol>

<ol>
	<li>Benzer, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+dissection+of+behavior.">Genetic dissection of behavior.</a> Sci. Am. 229, 24-37 (1973).</li><li>Ali, Y. O., Escala, W., Ruan, K., Zhai, R. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assaying+locomotor,+learning,+and+memory+deficits+in+Drosophila+models+of+neurodegeneration.">Assaying locomotor, learning, and memory deficits in Drosophila models of neurodegeneration.</a> J. Vis. Exp. , e2504 (2011).</li><li>de Vries, S. E., Clandinin, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optogenetic+Stimulation+of+Escape+Behavior+in+Drosophila+melanogaster.">Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster.</a> J. Vis. Exp. , e50192 (2013).</li><li>Mundiyanapurath, S., Certel, S., Kravitz, E. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Studying+aggression+in+Drosophila+(fruit+flies).">Studying aggression in Drosophila (fruit flies).</a> J. Vis. Exp. , e155 (2007).</li><li>Nichols, C. D., Becnel, J., Pandey, U. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Methods+to+assay+Drosophila+behavior.">Methods to assay Drosophila behavior.</a> J. Vis. Exp. , e3795 (2012).</li><li>Shiraiwa, T., Carlson, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Proboscis+extension+response+(PER)+assay+in+Drosophila.">Proboscis extension response (PER) assay in Drosophila.</a> J. Vis. Exp. , e193 (2007).</li><li>Lloyd, T. E., Taylor, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Flightless+flies:+Drosophila+models+of+neuromuscular+disease.">Flightless flies: Drosophila models of neuromuscular disease.</a> Ann. N.Y. Acad. Sci. 1184, e1-e20 (2010).</li><li>Atkinson, N. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+separation+of+two+behavioral+phenotypes+by+a+mutation+affecting+the+promoters+of+a+Ca-activated+K+channel.">Molecular separation of two behavioral phenotypes by a mutation affecting the promoters of a Ca-activated K channel.</a> J. Neurosci. 20, 2988-2993 (2000).</li><li>Atkinson, N. S., Robertson, G. A., Ganetzky, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+component+of+calcium-activated+potassium+channels+encoded+by+the+Drosophila+slo+locus.">A component of calcium-activated potassium channels encoded by the Drosophila slo locus.</a> Science. 253, 551-555 (1991).</li><li>Elkins, T., Ganetzky, B., Wu, C. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+Drosophila+mutation+that+eliminates+a+calcium-dependent+potassium+current.">A Drosophila mutation that eliminates a calcium-dependent potassium current.</a> Proc. Natl. Acad. Sci. U.S.A. 83, 8415-8419 (1986).</li><li>Gargano, J. W., Martin, I., Bhandari, P., Grotewiel, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rapid+iterative+negative+geotaxis+(RING):+a+new+method+for+assessing+age-related+locomotor+decline+in+Drosophila.">Rapid iterative negative geotaxis (RING): a new method for assessing age-related locomotor decline in Drosophila.</a> Exp. Gerontol. 40, 386-395 (2005).</li><li>Fry, S. N., Rohrseitz, N., Straw, A. D., Dickinson, M. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Visual+control+of+flight+speed+in+Drosophila+melanogaster.">Visual control of flight speed in Drosophila melanogaster.</a> J. Exp. Biol. 212, 1120-1130 (2009).</li><li>Mronz, M., Lehmann, F. O. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+free-flight+response+of+Drosophila+to+motion+of+the+visual+environment.">The free-flight response of Drosophila to motion of the visual environment.</a> J. Exp. Biol. 211, 2026-2045 (2008).</li></ol>

该作者有没有利益冲突披露。

使用这里介绍的方法，我们已经能够快速评估大量的果蝇突变体的飞行性能，提供比以前更高的效率。对于我们的实验中，我们经常分开的男性和女性，并提高他们在密度低（小于20苍蝇/瓶），以限制攻击可能会破坏翅膀。另一个重要的考虑是控制适当的，由于遗传背景的差异在飞行性能上的差异。我们还发现它有助于让苍蝇至少24小时的麻醉与二氧化碳在飞行试验之前恢复。另外，苍蝇?...

果蝇一直被用来研究行为1的遗传基础，而且研究人员已经设计出多种方法来分析不同类型的行为2-6。苍蝇一直在提供神经肌肉疾病7的有用模型特别有用。用来研究自发行为的普遍分析是飞行性能。原飞航测试仪是用于识别飞行缺陷突变体和对飞行能力1量化考核有用的，但它有几个缺点，限制了它的高通量筛选应用:采用涂油气缸是凌乱和繁琐，某些功能如气缸和介绍的苍蝇入管具有可变的力的长度减少量化精度，以及它是很难从测试仪恢复活苍蝇。为了克服这些限制，我们修改了飞行测试，包括了一些改进。我们增加了一个"降恩被"引入果蝇消除实验和用户之间的可变性，我们使用涂有粘合剂，其允许更容易清洗和个人蝇恢复可拆卸的压克力板，我们已经增加了飞行管的长度，以提高定量精度和可靠性。最后，我们使用数码相机和成像软件计算苍蝇降落的高度，我们相信这些改进将是有益的兴趣在飞行性能缺陷进行大规模遗传筛选任何实验室。

<table border="0" cellpadding="0" cellspacing="0" width="704">
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			<td height="33" style="height:33px;width:216px;">Putty knife</td>
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			<td><a href="http://www.homedepot.com">www.homedepot.com</a></td>
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			<td height="33" style="height:33px;width:216px;">Tangle Trap Insect Trap Coating</td>
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			<td><a href="http://www.biconet.com">www.biconet.com</a></td>
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			<td><a href="http://www.apple.com/mac/">www.apple.com/mac/</a></td>
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			<td height="33" style="height:33px;width:216px;">mineral oil</td>
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			<td height="33" style="height:33px;width:216px;">white poster board</td>
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			<td><a href="http://www.staples.com">www.staples.com</a></td>
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			<td style="width:171px;">Fisher Scientific</td>
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			<td height="33" style="height:33px;width:216px;">ImageJ Software</td>
			<td style="width:171px;">National Institutes of Health</td>
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			<td>http://rsb.info.nih.gov/ij/</td>
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			<td height="33" style="height:33px;width:216px;">digital camera</td>
			<td style="width:171px;">Sony</td>
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			<td height="33" style="height:33px;width:216px;">fine forceps</td>
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			<td><a href="http://www.finescience.com">www.finescience.com</a></td>
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			<td height="33" style="height:33px;width:216px;">polycarbonate cylinder (drop tube)</td>
			<td style="width:171px;">mcMaster-Carr</td>
			<td style="width:132px;">8585K62</td>
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			<td height="33" style="height:33px;width:216px;">flight cylinder (acrylic)</td>
			<td style="width:171px;">mcMaster-Carr</td>
			<td style="width:132px;">8486K943</td>
			<td><a href="http://www.mcmaster.com">www.mcmaster.com</a></td>
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			<td height="33" style="height:33px;width:216px;">polycarbonate sheets</td>
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			<td style="width:132px;">85585K25</td>
			<td><a href="http://www.mcmaster.com">www.mcmaster.com</a></td>
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			<td height="33" style="height:33px;width:216px;">ring stand (x2)</td>
			<td style="width:171px;">Fisher Scientific</td>
			<td style="width:132px;">S47808</td>
			<td><a href="http://www.fishersci.com">www.fishersci.com</a></td>
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			<td height="33" style="height:33px;width:216px;">ring support</td>
			<td style="width:171px;">Fisher Scientific</td>
			<td style="width:132px;">S47791</td>
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			<td height="33" style="height:33px;width:216px;">Three-prong extension clamps (x2)</td>
			<td style="width:171px;">Fisher Scientific</td>
			<td style="width:132px;">05-769-7Q</td>
			<td><a href="http://www.fishersci.com">www.fishersci.com</a></td>
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			<td height="33" style="height:33px;width:216px;">funnel</td>
			<td style="width:171px;">Fisher Scientific</td>
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			<td height="33" style="height:33px;width:216px;">chain clamps (x2)</td>
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			<td height="33" style="height:33px;width:216px;">glass vials</td>
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</table>

an improved method for accurate and rapid measurement of flight performance in drosophila

果蝇已经被证明是对行为的分析，一个有用的模型系统，包括飞行。在初始飞行测试涉及的滴飞入的油包覆的量筒;降落高度产生足够的推力，使其与圆筒的内壁接触之前提供通过评估多远苍蝇落入飞行性能的量度。在这里，我们描述了飞行测试仪有四个重大改进的更新版本。首先，我们增加了一个"落管"，以确保所有的苍蝇进入飞行圆柱体试验之间类似的速度，用户之间消除了变异。其次，我们与涂在缠结陷阱，旨在捕捉活虫粘合剂可拆卸的塑料片代替油涂层。第三，我们使用一个较长的气缸以使飞行能力更精确鉴别。第四，我们使用数码相机和成像软件，可以自动飞行性能得分。这些改进允许说唱ID，的飞行行为的定量评估，对于大型数据集和大规模遗传筛选有用的。

<html> 图1A示出了更新后的航班仪组件的示意图。 图1B示出的轨道设计允许相机拍摄的全景图像而不阻塞的视野。代表性的结果示于图2，其中slowpoke突变体果 ​​蝇，它具有已知的飞行缺陷8-10的飞行性能，进行比较，野生型广-S苍蝇。控制苍蝇一贯降落靠近圆筒的顶部，与个体之间的非常小的扩散和73±2.0厘米平均降落高度。相比之下?...

Watch this Scientific Journal Video about An Improved Method for Accurate and Rapid Measurement of Flight Performance in Drosophila at JoVE.com

An Improved Method for Accurate and Rapid Measurement of Flight Performance in Drosophila

这里，我们描述了一种用于飞行性能果蝇快速和准确的测量，从而实现高通量筛选。

在对飞行性能的准确，快速测定方法的改进<em&gt;果蝇</em

Drosophila has proven to be a useful model system for analysis of behavior, including flight. The initial flight tester involved dropping flies into an ...

an-improved-method-for-accurate-rapid-measurement-flight-performance

Universidad de Cartagena UDEC

Research

JoVE Journal

Biology

An Improved Method for Accurate and Rapid Measurement of Flight Performance in Drosophila

11.7K 次观看.  University of Wisconsin-Madison. 这里，我们描述了一种用于飞行性能果蝇快速和准确的测量，从而实现高通量筛选。 

视频: An Improved Method for Accurate and Rapid Measurement of Flight Performance in Drosophila

Pyrosequencing: A Simple Method for Accurate Genotyping

Pharmacogenetic research benefits first-hand from the abundance of information provided by the completion of the Human Genome Project. With such a tremendous amount of data available comes an explosion of genotyping methods. Pyrosequencing(R) is one of the most thorough yet simple methods to date used to analyze polymorphisms. It also has the ability to identify tri-allelic, indels, short-repeat polymorphisms, along with determining allele percentages for methylation or pooled sample assessment. In addition, there is a standardized control sequence that provides internal quality control. This method has led to rapid and efficient single-nucleotide polymorphism evaluation including many clinically relevant polymorphisms. The technique and methodology of Pyrosequencing is explained.


Pyrosequencing(R) is one of the most thorough yet simple methods to date used to analyze polymorphisms. This method has led to rapid and efficient single-nucleotide polymorphism evaluation including many clinically relevant polymorphisms. The technique and methodology of Pyrosequencing is explained.

焦磷酸测序：一个准确的基因分型的简单方法

Pharmacogenetic research benefits first-hand from the abundance of information provided by the completion of the Human Genome Project. With such a ...

科研

生物学

A Magnetic Tether System to Investigate Visual and Olfactory Mediated Flight Control in Drosophila

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. As such, visual stabilization of upwind tracking directly aids in odor tracking. But do olfactory signals directly influence visual tracking behavior independently from wind cues? Also, the recent deluge of research on the neurophysiology and neurobehavioral genetics of olfaction in Drosophila has motivated ever more technically sophisticated and quantitative behavioral assays. Here, we modified a magnetic tether system originally devised for vision experiments by equipping the arena with narrow laminar flow odor plumes. A fly is glued to a small steel pin and suspended in a magnetic field that enables it to yaw freely. Small diameter food odor plumes are directed downward over the fly s head, eliciting stable tracking by a hungry fly. Here we focus on the critical mechanics of tethering, aligning the magnets, devising the odor plume, and confirming stable odor tracking.

Here we describe how to tether a fly in an olfactory magnetic-tether (OMT) apparatus. We describe how to align the rare-earth magnets and odor ports, and how to set mass flow rates for both the stimulus delivery and vacuum suction to achieve optimal odor tracking.

一个磁性的系绳系统，以调查在果蝇的视觉和嗅觉介导的飞行控制

It has been clear for many years that insects use visual cues to stabilize their heading in a wind stream. Many animals track odors carried in the wind. ...

Visually Mediated Odor Tracking During Flight in Drosophila

Flying insects use visual cues to stabilize their heading in a wind stream. Many animals additionally track odors carried in the wind. As such, visual stabilization of upwind tracking directly aids in odor tracking. But do olfactory signals directly influence visual tracking behavior independently from wind cues? Additionally, recent advances in olfactory molecular genetics and neurophysiology have motivated novel quantitative behavioral analyses to assess the behavioral influence of (e.g.) genetically inactivating specific olfactory activation circuits. We modified a magnetic tether system originally devised for vision experiments by equipping the arena with narrow laminar flow odor plumes. Here we focus on experiments that can be performed after a fly is tethered and is able to navigate in the magnetic arena. We show how to acquire video images optimized for measuring body angle, how to judge stable odor tracking, and we illustrate two experiments to examine the influence of visual cues on odor tracking.

Here we describe how to optimize the acquired video image for an olfactory magnetic-tether (OMT) apparatus. We also describe two sample experimental protocols for studying visuo-olfactory fusion.

在果蝇的飞行过程中可视介导的气味追踪

Flying insects use visual cues to stabilize their heading in a wind stream. Many animals additionally track odors carried in the wind. As such, visual ...

Endurance Training Protocol and Longitudinal Performance Assays for Drosophila melanogaster

One of the most pressing problems facing modern medical researchers is the surging levels of obesity, with the consequent increase in associated disorders such as diabetes and cardiovascular disease 1-3. An important topic of research into these associated health problems involves the role of endurance exercise as a beneficial intervention.
Exercise training is an inexpensive, non-invasive intervention with several beneficial results, including reduction in excess body fat 4, increased insulin sensitivity in skeletal muscle 5, increased anti-inflammatory and antioxidative responses 6, and improved contractile capacity in cardiomyocytes 7. Low intensity exercise is known to increase mitochondrial activity and biogenesis in humans 8 and mice, with the transcriptional coactivator PGC1-&alpha; as an important intermediate 9,10. 
Despite the importance of exercise as a tool for combating several important age-related diseases, extensive longitudinal genetic studies have been impeded by the lack of an endurance training protocol for a short-lived genetic model species. The variety of genetic tools available for use with Drosophila, together with its short lifespan and inexpensive maintenance, make it an appealing model for further study of these genetic mechanisms. With this in mind we have developed a novel apparatus, known as the Power Tower, for large scale exercise-training in Drosophila melanogaster 11. The Power Tower utilizes the flies' instinctive negative geotaxis behavior to repetitively induce rapid climbing. Each time the machine lifts, then drops, the platform of flies, the flies are induced to climb. Flies continue to respond as long as the machine is in operation or until they become too fatigued to respond. Thus, the researcher can use this machine to provide simultaneous training to large numbers of age-matched and genetically identical flies. Additionally, we describe associated assays useful to track longitudinal progress of fly cohorts during training.

Endurance Training Protocol and Longitudinal Performance Assays for Drosophila melanogaster

We describe the first endurance training protocol for an important genetic model species, Drosophila melanogaster, and outline several assays to chart improvements in mobility following training.

果蝇的耐力训练协议和纵向性能检测

One of the most pressing problems facing modern medical researchers is the surging levels of obesity, with the consequent increase in associated disorders ...

Measurement of Lifespan in Drosophila melanogaster

Aging is a phenomenon that results in steady physiological deterioration in nearly all organisms in which it has been examined, leading to reduced physical performance and increased risk of disease. Individual aging is manifest at the population level as an increase in age-dependent mortality, which is often measured in the laboratory by observing lifespan in large cohorts of age-matched individuals. Experiments that seek to quantify the extent to which genetic or environmental manipulations impact lifespan in simple model organisms have been remarkably successful for understanding the aspects of aging that are conserved across taxa and for inspiring new strategies for extending lifespan and preventing age-associated disease in mammals.
The vinegar fly, Drosophila melanogaster, is an attractive model organism for studying the mechanisms of aging due to its relatively short lifespan, convenient husbandry, and facile genetics. However, demographic measures of aging, including age-specific survival and mortality, are extraordinarily susceptible to even minor variations in experimental design and environment, and the maintenance of strict laboratory practices for the duration of aging experiments is required. These considerations, together with the need to practice careful control of genetic background, are essential for generating robust measurements. Indeed, there are many notable controversies surrounding inference from longevity experiments in yeast, worms, flies and mice that have been traced to environmental or genetic artifacts1-4. In this protocol, we describe a set of procedures that have been optimized over many years of measuring longevity in Drosophila using laboratory vials. We also describe the use of the dLife software, which was developed by our laboratory and is available for download (<a href="http://sitemaker.umich.edu/pletcherlab/software">http://sitemaker.umich.edu/pletcherlab/software</a>). dLife accelerates throughput and promotes good practices by incorporating optimal experimental design, simplifying fly handling and data collection, and standardizing data analysis. We will also discuss the many potential pitfalls in the design, collection, and interpretation of lifespan data, and we provide steps to avoid these dangers.

Measurement of Lifespan in Drosophila melanogaster

Drosophila melanogaster is a powerful model organism for exploring the molecular basis of longevity regulation. This protocol will discuss the steps involved in generating a reproducible, population-based measurement of longevity as well as potential pitfalls and how to avoid them.

寿命测量<em&gt;果蝇</em

Aging is a phenomenon that results in steady physiological deterioration in nearly all organisms in which it has been examined, leading to reduced ...

An Innovative Method for Exosome Quantification and Size Measurement

Although the biological importance of exosomes has recently gained an increasing amount of scientific and clinical attention, much is still unknown about their complex pathways, their bioavailability and their diverse functions in health and disease. Current work focuses on the presence and the behavior of exosomes (in vitro as well as in vivo) in the context of different human disorders, especially in the fields of oncology, gynecology and cardiology.
Unfortunately, neither a consensus regarding a gold standard for exosome isolation exists, nor is there an agreement on such a method for their quantitative analysis. As there are many methods for the purification of exosomes and also many possibilities for their quantitative and qualitative analysis, it is difficult to determine a combination of methods for the ideal approach. 
Here, we demonstrate nanoparticle tracking analysis (NTA), a semi-automated method for the characterization of exosomes after isolation from human plasma by ultracentrifugation. The presented results show that this approach for isolation, as well as the determination of the average number and size of exosomes, delivers reproducible and valid data, as confirmed by other methods, such as scanning electron microscopy (SEM).

A method for isolation of exosomes from whole blood and further analysis by nanoparticle tracking using a semi-automatic instrument is presented in this article. The presented technology provides an extremely sensitive method for visualizing and analyzing particles in liquid suspension.

对于外来体定量和尺寸测量新方法研究

Although the biological importance of exosomes has recently gained an increasing amount of scientific and clinical attention, much is still unknown about ...

Measurement of Metabolic Rate in Drosophila using Respirometry

Metabolic disorders are a frequent problem affecting human health. Therefore, understanding the mechanisms that regulate metabolism is a crucial scientific task. Many disease causing genes in humans have a fly homologue, making Drosophila a good model to study signaling pathways involved in the development of different disorders. Additionally, the tractability of Drosophila simplifies genetic screens to aid in identifying novel therapeutic targets that may regulate metabolism. In order to perform such a screen a simple and fast method to identify changes in the metabolic state of flies is necessary. In general, carbon dioxide production is a good indicator of substrate oxidation and energy expenditure providing information about metabolic state. In this protocol we introduce a simple method to measure CO2 output from flies. This technique can potentially aid in the identification of genetic perturbations affecting metabolic rate.

Measurement of Metabolic Rate in Drosophila using Respirometry

Metabolic disorders are among one of the most common diseases in humans. The genetically tractable model organism D. melanogaster can be used to identify novel genes that regulate metabolism. This paper describes a relatively simple method which allows studying the metabolic rate in flies by measuring their CO2 production.

代谢率的测量<em&gt;果蝇</em&gt;使用呼吸运动

Metabolic disorders are a frequent problem affecting human health. Therefore, understanding the mechanisms that regulate metabolism is a crucial ...

Filtration Isolation of Nucleic Acids:  A Simple and Rapid DNA Extraction Method

FINA, filtration isolation of nucleic acids, is a novel extraction method which utilizes vertical filtration via a separation membrane and absorbent pad to extract cellular DNA from whole blood in less than 2 min. The blood specimen is treated with detergent, mixed briefly and applied by pipet to the separation membrane. The lysate wicks into the blotting pad due to capillary action, capturing the genomic DNA on the surface of the separation membrane. The extracted DNA is retained on the membrane during a simple wash step wherein PCR inhibitors are wicked into the absorbent blotting pad. The membrane containing the entrapped DNA is then added to the PCR reaction without further purification. This simple method does not require laboratory equipment and can be easily implemented with inexpensive laboratory supplies. Here we describe a protocol for highly sensitive detection and quantitation of HIV-1 proviral DNA from 100 &#181;l whole blood as a model for early infant diagnosis of HIV that could readily be adapted to other genetic targets.

We describe here a simple and rapid paper-based DNA extraction method of HIV proviral DNA from whole blood detected by quantitative PCR. This protocol can be extended for use in detecting other genetic markers or using alternative amplification methods.

核酸的分离过滤:一种简单快速的DNA提取方法

FINA, filtration isolation of nucleic acids, is a novel extraction method which utilizes vertical filtration via a separation membrane and absorbent pad ...

Hybrid-Cut: An Improved Sectioning Method for Recalcitrant Plant Tissue Samples

Maintaining plant section integrity is essential for studying detailed anatomical structures at the cellular, tissue, or even organ level. However, some plant cells have rigid cell walls, tough fibers and crystals (calcium oxalate, silica, etc.), and high water content that often disrupt tissue integrity during plant tissue sectioning.
This study establishes a simple Hybrid-Cut tissue sectioning method. This protocol modifies a paraffin-based sectioning technique and improves the integrity of tissue sections from different plants. Plant tissues were embedded in paraffin before sectioning in a cryostat at -16 &#176;C. Sectioning under low temperature hardened the paraffin blocks, reduced tearing and scratching, and improved tissue integrity significantly. This protocol was successfully applied to calcium oxalate-rich Phalaenopsis orchid tissues as well as recalcitrant tissues such as reproductive organs and leaves of rice, maize, and wheat. In addition, the high quality of tissue sections from Hybrid-Cut could be used in combination with in situ hybridization (ISH) to provide spatial expression patterns of genes of interest. In conclusion, this protocol is particularly useful for recalcitrant plant tissue containing high crystal or silica content. Good quality tissue sections enable morphological and other biological studies.

This protocol describes a simple Hybrid-Cut tissue sectioning method that is useful for recalcitrant plant tissues. Good quality tissue sections enable anatomical studies and other biological studies including in situ hybridization (ISH).

混合式切割:用于顽拗植物组织样品的改进切法

Maintaining plant section integrity is essential for studying detailed anatomical structures at the cellular, tissue, or even organ level. However, some ...

Optical Cross-Sectional Muscle Area Determination of Drosophila Melanogaster Adult Indirect Flight Muscles

Muscle mass wasting, known as muscle atrophy, is a common phenotype in Drosophila models of neuromuscular diseases. We have used the indirect flight muscles (IFMs) of flies, specifically the dorso-longitudinal muscles (DLM), as the experimental subject to measure the atrophic phenotype brought about by different genetic causes. In this protocol, we describe how to embed fly thorax muscles for semi thin sectioning, how to obtain a good contrast between muscle and the surrounding tissue, and how to process optical microscope images for semiautomatic acquisition of quantifiable data and analysis. We describe three specific applications of the methodological pipeline. First, we show how the method can be applied to quantify muscle degeneration in a myotonic dystrophy fly model; second, measurement of muscle cross-sectional area can help to identify genes that either promote or prevent muscle atrophy and/or muscle degeneration; third, this protocol can be applied to determine whether a candidate compound is able to significantly modify a given atrophic phenotype induced by a disease-causing mutation or by an environmental trigger.

Optical Cross-Sectional Muscle Area Determination of Drosophila Melanogaster Adult Indirect Flight Muscles

We report a method to quantify muscle area, which is an indirect method to determine muscle mass in Drosophila adults. We demonstrate the application of our methodology by analyzing the indirect flight muscles in a Drosophila model of Myotonic Dystrophy disease.

果蝇成人间接飞行肌的光学横截肌面积测定

Muscle mass wasting, known as muscle atrophy, is a common phenotype in Drosophila models of neuromuscular diseases. We have used the indirect flight ...

在对飞行性能的准确，快速测定方法的改进

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