Name: two types of assays for detecting frog sperm chemoattraction
Uploaded: 2011-12-27T13:00:00
Description: Watch this Scientific Journal Video about Two Types of Assays for Detecting Frog Sperm Chemoattraction at JoVE.com

我们感谢他们的视频显微镜工作站使用的WM凯克生物成像实验室。支持这项研究是由NSF资助IBN - 0615435。

1。所使用的材料和缓冲区<ol><li>卵母细胞林格氏液（1.5 × OR2）包含124毫米氯化钠，3.75 mM的氯化钾，1.5毫米氯化钙2，1.5毫米氯化镁2，1.5毫米的Na 2 HPO 4，10毫米HEPES， pH值7.8。施肥缓冲器（F - 1）包含41.25 mM氯化钠，氯化钾1.25毫米，0.25毫米氯化钙2，0.06毫米氯化镁2，0.5毫米娜2 HPO 4，2.5毫米的HEPES，pH值7.8。 </li><li> ...

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Chem. 286, 17311-17325 (2011).</li><li>Tholl, N., Naqvi, S., McLaughlin, E., Boyles, S., Bieber, A. L., Chandler, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Swimming+of+Xenopus+laevis+sperm+exhibits+multiple+gears+and+its+duration+is+extended+by+egg+jelly+constituents.">Swimming of Xenopus laevis sperm exhibits multiple gears and its duration is extended by egg jelly constituents.</a> Biol. Bull. 220, 174-185 (2011).</li><li>Eisenbach, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sperm+chemotaxis.">Sperm chemotaxis.</a> Rev. Reprod. 4, 56-66 (1999).</li><li>Riffell, J. A., Zimmer, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sex+and+flow:+the+consequences+of+fluid+shear+for+sperm-egg+interactions.">Sex and flow: the consequences of fluid shear for sperm-egg interactions.</a> J. Exp. Biol. 210, 3644-3660 (2007).</li><li>Corkidi, G., Taboada, B., Wood, C. D., Guerrero, A., Darszon, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tracking+sperm+in+three-dimensions.">Tracking sperm in three-dimensions.</a> Biochem. Biophys. Res. Commun. 373, 125-129 (2008).</li><li>Himes, J. E., Riffell, J. A., Zimmer, C. A., Zimmer, R. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sperm+chemotaxis+as+revealed+with+live+and+synthetic+eggs.">Sperm chemotaxis as revealed with live and synthetic eggs.</a> Biol Bull. 220, 1-5 (2011).</li><li>Sun, F., Giojolas, L. C., Rovasio, R. A., Tur-Kaspa, I., Sanchez, R., Eisenbach, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lack+of+species-specificity+in+mammalian+sperm+chemotaxis.">Lack of species-specificity in mammalian sperm chemotaxis.</a> Dev. Biol. 255, 423-427 (2003).</li><li>Guidobaldi, H. A., Teves, M. E., U&#241;ates, D. R., Anastas&#237;a, A., Giojalas, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progesterone+from+the+cumulus+cells+is+the+sperm+chemoattractant+secreted+by+the+rabbit+oocyte+cumulus+complex.">Progesterone from the cumulus cells is the sperm chemoattractant secreted by the rabbit oocyte cumulus complex.</a> PLoS One. 3, e3040-e3040 (2008).</li><li>Oren-Benaroya, R., Orvieto, R., Gakamsky, A., Pinchasov, M., Eisenbach, 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+sperm+chemoattractant+secreted+from+human+cumulus+cells+is+progesterone.">The sperm chemoattractant secreted from human cumulus cells is progesterone.</a> Hum. Reprod. 23, 2339-2345 (2008).</li><li>Teves, M. E., Guidobaldi, H. A., U&#241;ates, D. R., Sanchez, R., Miska, W., Publicover, S. J., Morales Garcia, A. A., Giojalas, L. 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L., Kirichok, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progesterone+activates+the+principal+Ca2++channel+of+human+sperm.">Progesterone activates the principal Ca2+ channel of human sperm.</a> Nature. 471, 387-391 (2011).</li><li>Boyden, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+chemotactic+effect+of+mixtures+of+antibody+and+antigen+on+polymorphonuclear+leucocytes.">The chemotactic effect of mixtures of antibody and antigen on polymorphonuclear leucocytes.</a> J. Exp. Med. 115, 453-465 (1962).</li><li>Zigmond, S. H., Lauffenburger, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Assays+of+leukocyte+chemotaxis.">Assays of leukocyte chemotaxis.</a> Annu. Rev. Med. 37, 149-155 (1986).</li><li>Villanueva-Diaz, C., Vadillo-Ortega, F., Kably-Ambe, A., Diaz-Pérez, M. A., Krivitzky, S. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+that+human+follicular+fluid+contains+a+chemoattractant+for+spermatozoa.">Evidence that human follicular fluid contains a chemoattractant for spermatozoa.</a> Fertil. Steril. 54, 1180-1182 (1990).</li><li>Villanueva-Diaz, C., Arizs-Martinez, J., Bermejo-Martinez, L., Vadillo-Ortega, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Progesterone+induces+human+sperm+chemotaxis.">Progesterone induces human sperm chemotaxis.</a> Fertil. Steril. 64, 1183-1188 (1995).</li><li>Olson, J., Xiang, X., Ziegert, T., Kittleson, A., Rawls, A., Bieber, A., Chandler, D. E. 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要么变形虫运动或鞭毛动力的游泳运动细胞趋化在许多生物中发现和研究这一现象，需要提供实用，可靠的的检测。一些现象的例子，如精子海胆卵或粘菌细胞的聚集，形成子实体的吸引力，有直接的视觉冲击力。这种现象已艾森巴赫18所述的各种方法进行定量。这些实验包括充满毛细血管吸引精子，加入水库，以创建一个渐变和使用的一个Makler室中有精子和趋化井使用的毛细血管细胞趋...

<table border="1"><tbody><tr><td> 项目名称 </td><td> 公司 </td><td> 目录编号 </td><td> 评论 </td></tr><tr><td> 24孔板</td><td>流式细胞-迪肯森</td><td>一千一百四十七分之三十五</td><td></td></tr><tr><td> 12毫米，外径插入12μm的微孔滤膜</td><td> Millipore公司</td><td> PIXP01250 </td><td>我们以前使用中光学康宁Transwell小板＃3403已经停产</td></tr><tr><td> Zigmond室</td><td> Neuroprobe </td><td> Z02 </td><td></td></tr><tr><td>硅油</td><td>通用电气公司</td><td> SF1154 </td><td>道康宁550流体相当于</td></tr><tr><td> Image J软件</td><td>韦恩Rasband，研究服务处，国立精神卫生研究所</td><td>免费下载 http://rsbweb.nih.gov/ij </td><td> Java程序运行在Windows，林UX和Mac </td></tr><tr><td> MtrackJ软件</td><td>埃里克Meijering / Imagescience / <a href="http://www.bigr.nl/">生物医学成像集团</a> ，伊拉斯谟MC -鹿特丹大学医学中心</td><td>在<a href="http://www.imagescience.org/meijering/software/mtrackj/">http://www.imagescience.org/meijering/software/mtrackj/</a>免费下载</td><td> Java程序运行在Windows，Linux和Mac </td></tr><tr><td>虚拟配音软件</td><td> <a href="http://www.virtualdub.org/gpl.html">GNU通用公共许可</a>软件</td><td>通过<a href="http://www.virtualdub.org/index.html">http://www.virtualdub.org/index.html</a>免费下载</td><td>安装说明仅适用于Windows插件“下的”图片J网站; </td></tr><tr><td> cellSens软件</td><td>奥林巴斯</td><td>请参阅网站<a href="http://www.olympusamerica.com/seg_section/product.asp?product=1070">：</a> http://www.olympusamerica.com/seg_section/product.asp？产品= 1070 </td><td>控制和获取各种摄像头的图像。也有图像处理能力</td></tr></tbody></table>

two types of assays for detecting frog sperm chemoattraction

精子在无脊椎动物chemoattraction可以足够强大，可以将吸管将含有一个精子悬液的吸引力肽和微观可视精子积累周围移液器1。在脊椎动物如青蛙，啮齿动物和人类的精子chemoattraction是比较困难的检测和定量分析要求。这种检测两大类型 - 定量来源的趋化，所谓的精子积累分析，和实际跟踪单个精子的游泳轨迹的检测精子的运动检测。 精子积累实验较快，允许数十或数百个在一天内完成检测，从而使剂量反应曲线和时间课程进行较快。这些类型的检测已广泛用于表征许多行之有效的chemoattraction系统 - 例如，中性粒细胞趋化BACterial肽和精子的趋化卵泡液。精子的跟踪分析，可以更多的劳动力密集型，但对如何chemoattractancts实际上改变的游泳路径，精子采取提供额外的数据。这类型的检测是需要表现出精子的运动方向相chemoattrractant渐变轴和可视化特征或转方向的变化，使精子接近卵子。 在这里，我们描述了这两种类型的检测采用的方法。利用精子积累检测，被称为“两院”法。两栖类动物的精子被放置在一个组织文化与聚碳酸酯过滤器直径为12微米的毛孔地板的板插入。插入与精子放入组织培养板中含有缓冲区和底部以及地面符合墙上（见图1）。吸管小心地将一个chemoatttractant井。潜伏期后，顶端插入含有精子水库是经过精心řemoved，并在底部室，通过膜的传递的精子都将被删除，颗粒，然后通过血球计数或流式细胞仪。 精子跟踪检测，采用了Zigmond室最初观察中性粒细胞趋化，并观察精子的修改Giojalas 和同事2,3。该商会由一个厚厚的玻璃幻灯片已被加工成两纵槽。这些相隔1毫米宽的观测平台。经过一个玻璃盖，精子的应用程序加载到一个低谷，到其他运动和单个精子的趋化剂的可视化视频显微镜。录像分析软件，以确定时间（xyt数据集），每个精子的运动轨迹功能在XY平面二维的细胞运动。

Watch this Scientific Journal Video about Two Types of Assays for Detecting Frog Sperm Chemoattraction at JoVE.com

Two Types of Assays for Detecting Frog Sperm Chemoattraction

蛋和蛋的周围外涂料频频释放多肽，蛋白质和小分子，与精子的沟通，引导他们到卵子，从而促进受精。利用青蛙的精子，我们描述和比较两类检测，用于检测精子chemoattraction - 精子的积累检测和精子跟踪检测。

两种类型的检测，检测青蛙精子Chemoattraction

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A High-Throughput Method For Zebrafish Sperm Cryopreservation and In Vitro Fertilization

A High-Throughput Method For Zebrafish Sperm Cryopreservation and In Vitro Fertilization

对于斑马鱼精子超低温冷冻保存的一个高通量的方法和<em在体外施肥

This is a method for zebrafish sperm cryopreservation that is an adaptation of the Harvey method (Harvey et al., 1982).  We have introduced two changes to ...

Techniques for Imaging Ca2+ Signaling in Human Sperm

Techniques for Imaging Ca2+ Signaling in Human Sperm

技术成像钙 2 +信号在人类精子

Fluorescence microscopy of cells loaded with fluorescent, Ca2+-sensitive dyes is used for measurement of spatial and temporal aspects of Ca2+ signaling in ...

Analysis of Schwann-astrocyte Interactions Using In Vitro Assays

Analysis of Schwann-astrocyte Interactions Using In Vitro Assays

雪旺的星形胶质细胞使用相互作用分析<em在体外检测

Schwann cells are one of the commonly used cells in repair strategies following spinal cord injuries. Schwann cells are capable of supporting axonal ...

Isolation and In vitro Activation of Caenorhabditis elegans Sperm

Isolation and In vitro Activation of Caenorhabditis elegans Sperm

分离和<em在体外激活线虫精子

Males and hermaphrodites are the two naturally found sexual forms in the nematode C. elegans. The amoeboid sperm are produced by both males and ...

Extraction of Tissue Antigens for Functional Assays

提取组织抗原的功能分析

Many of the antigen targets of adaptive immune response, recognized by B and T cells, have not been defined 1. This is particularly true in autoimmune ...

Assessing Differences in Sperm Competitive Ability in Drosophila

Assessing Differences in Sperm Competitive Ability in Drosophila

评估精子竞争能力的差异<em&gt;果蝇</em

Competition among conspecific males for fertilizing the ova is one of the mechanisms of sexual selection, i.e. selection that operates on maximizing the ...

Viability Assays for Cells in Culture

活性检测在培养细胞

Manual cell counts on a microscope are a sensitive means of assessing cellular viability but are time-consuming and therefore expensive. Computerized ...

Development of an Insert Co-culture System of Two Cellular Types in the Absence of Cell-Cell Contact

在细胞间接触缺失的两个蜂窝型的插入共培养系统的开发

The role of secreted soluble factors in the modification of cellular responses is a recurrent theme in the study of all tissues and systems. In an attempt ...