作者感谢卢尔德·卡伦·谭女士在养鱼方面给予的帮助。这项工作由NMBU、日本科学促进会（JSPS）的助学金（赠款编号18H04881和18K19323）资助，并资助住友基金会至S.K.的基础科学研究项目。

所有实验和动物处理都是根据挪威生命科学大学关于实验动物福利的建议进行的。挪威食品安全局（FOTS ID 24305）批准了使用淋病切除术的实验。
注：实验采用成年男性和女性（6-7个月大，体重约0.35克，长度约2.7厘米）日本美的进行。性别是通过区分次要性特征，如背鳍和肛鳍的大小和形状，如42，47描述确定。
1. 仪器和解决方案准备
<ol><li>准备麻醉库存溶液（0.6% 三卡因）。<ol>
<li>稀释 0.6 克三甲酸酯 （MS-222） 在 100 mL 的 10 倍磷酸盐缓冲盐水 （PBS）.</li>
		<li>将 1 mL 的 Tricaine 库存溶液分发到多个 1.5 mL 塑料管中，并储存在 -20 °C 下，直至使用。</li>
	</ol>
</li>
	<li>将 18 克 NaCl 加入 2 升水族馆水中，准备回收水（0.9% NaCl 溶液）。将溶液存放在室温下，直到使用。</li>
	<li>通过对角线打破剃须刀来准备切口工具，以获得一个锐点（图1A）。</li>
	<li>通过将 25 μL 肝素钠稀释到 1x PBS 的 500 μL 中，准备血液抗凝血溶液（0.05 U/μL 肝素钠）。将抗凝血液存储在 4 °C 下，直到使用。</li>
	<li>根据制造商的指示，用拔针器（图1B）拉玻璃毛细圈，从90毫米长的玻璃毛细圈中准备两根玻璃针。 注：玻璃针的外径为1毫米，内径为0.6毫米。</li>
	<li>通过切割盖子来准备一个1.5mL塑料管盖，并制作一个与针外径相符的孔（图1C）。要打孔，加热 9 毫米玻璃毛细孔的一端，并通过盖子刺穿加热的玻璃毛细圈。或者，使用针刺穿盖子，直到孔的直径与 9 毫米玻璃毛细孔相符。</li>
</ol>2. 妇科切除术
<ol><li>在30mL的水族馆水中稀释一管三甲酸酯（0.6%），准备0.02%的麻醉液。</li>
	<li>准备解剖工具，包括一个超细和两个细钳（一个有相对宽的尖端），小剪刀，尼龙线和剃须刀，如第1.3步所述。</li>
	<li>将鱼放入 0.02% 麻醉液中 30-60 秒，使鱼麻醉。 注：麻醉的持续时间取决于鱼的大小和重量，必须加以调整。为了确保鱼完全麻醉，鱼体可以用钳子轻轻捏捏。如果鱼没有反应，就可以开始做腺切除术。</li>
	<li>从麻醉液中取出鱼，将鱼水平放在其一侧，在解剖显微镜下出水。</li>
	<li>女性卵巢切除术 （OVX）<ol>
<li>取出卵卵（挂在女性身体外的卵子），如果有的话，刮切口区域的鳞片（图2A）。</li>
		<li>使用剃须刀刀片，在肋骨之间、骨盆和鼻鳍之间轻轻切口约2-2.5毫米长（图2A）。然后，轻轻捏鱼腹部，同时用宽尖的细钳一点一点地取出卵巢。</li>
		<li>使用细钳切开卵巢末端，将卵巢放在一边（图2B）。 注意：如果可能，请注意不要折断卵巢囊。如果卵巢囊破裂，尽可能完全去除任何腺体痕迹，而不留下任何未排卵的卵子。</li>
	</ol></li>
	<li>男性兰花切除术<ol>
<li>轻轻地在肛门上方的肋骨之间切开（图2A），然后用细钳慢慢打开切口。</li>
		<li>用细钳轻轻抓住睾试，慢慢取出睾试。之后，切开睾片的末端，以完全去除睾测试（图2B）。对于男性兰花切除术，所有制剂都类似于女性，直到切口部分。抓睾考试时，有时会获得类似于睾子的脂肪。然而，在恢复脂肪后，可以尝试再次找到睾试（图2B）。 注：对于男性和女性来说，重要的是尽量减少腹部的切口大小，以防止可能导致死亡的过度损伤。有时肠道也可能通过切口和淋头出现，因此请确保在关闭前将它们正确返回切口内。先前对卵巢和睾睾门在梅达卡腹部的位置的了解是必不可少的。</li>
	</ol></li>
	<li>缝合切口同样在男性和女性（图3）。<ol>
<li>将尼龙线放在切口区旁边，用超细钳子将切口右侧的皮肤刺入内侧腔内，用细钳将线带入（图3：1-2）。</li>
		<li>通过外体腔从切口左侧刺出皮肤以取出线（图3：3-4）。</li>
		<li>关闭切口开口，打两个结，切开过多的线（图3：4-6）。 注：缝合线必须足够紧，鱼的剩余线必须足够长，以防止缝合线松动。从麻醉到缝注的整个过程通常需要长达6分钟。更长的时间可能导致死亡。</li>
		<li>将鱼放入回收水中，并放置至少24小时，然后再将其转移到水族馆系统。 注：果纳切除鱼类通常在回收水中1-2小时后表现出正常行为。因此，根据实验目的，一个人可以在这个时间间隔后对鱼进行采样。</li>
	</ol></li>
</ol>3. 血液采样程序
<ol><li>准备工具：玻璃针、硅胶毛细管、带孔的塑料管、空的1.5毫升塑料管、迷你中心和胶带。</li>
	<li>使用第 2.1 步中描述的 0.02% 麻醉液对鱼进行麻醉，并将鱼置于垂直位置的解剖显微镜下（图 4A）。将鱼放在明亮的表面上，以方便眼花孔穿刺静脉的可视化。</li>
	<li>将玻璃针连接到硅胶毛细血管上（图4B），安装抽屉。用宽尖钳子打破针尖，通过吸气和吹动将针头内部涂上抗凝固液。 注意：建议使用吸盘和长度至少为 50 厘米的硅胶毛细血管，以采取安全措施，避免吸吸时直接接触血液。此外，确保针尖的开口足够大，以便抽取血液。</li>
	<li>将针头引向鱼的斑点区域，瞄准静脉（图5A），用嘴抽血，直到至少四分之一的针头总体积被填充（图5B）。 注意：在从鱼体中取出针头之前，必须停止吸气。</li>
	<li>松开针头，在针尖侧附近放一块胶带。将盖子与一个孔放在收集管上，将针头放在管内，通过孔，针尖放在外侧（图5C）。</li>
	<li>将鱼放入回收水中，并放置至少24小时，然后再将其转移到水族馆系统。 注：要从同一条鱼进行第二次血液取样，请在第一次血液采样一周后对血液进行取样。</li>
	<li>闪光旋转收集的血液1-2秒，在室温下1000克收集管内的血液。</li>
	<li>直接进入下游应用或将血液储存在 -20 °C 下，直到使用。 注： 参考以前的研究性类固醇提取从整个血液48.</li>
</ol>

作者没有什么可透露的。

如以往文献所报道，在其他模型物种中，长期以来一直使用造血术和血液采样来调查与性别类固醇在BPG轴调节中的作用有关的问题。然而，这些技术似乎只适合更大的动物。考虑到常用的远程模型（日本美田）体积小，我们为该物种的去结肠切除术和血液采样提供了详细的协议。
果子切除术鱼类的成活率达到近100%，表明在麦田上应用果子切除术是可行的。同样，血液采样过?...

在脊椎动物中，性类固醇，主要是由性腺动物产生的，通过各种反馈机制1，2，3，4，5在大脑-脑-脑-淋巴（BPG）轴的调节中起着重要的作用。此外，性类固醇影响大脑6，7，8和内分泌细胞，包括脑垂体9，10神经元的增殖和活动，从而在大脑和垂体可塑性的关键作用。 尽管在哺乳动物方面知识相对较好，但由性类固醇调停的BPG轴调节机制在非哺乳动物物种中还远远不为人所知，导致对进化保护原则的理解不充分。仍然有数量有限的研究记录性类固醇在大脑和垂体可塑性的作用， 从而提出了进一步研究性类固醇对各种脊椎动物物种的作用和影响的需要.
在脊椎动物中，远程运动已成为解决许多生物和生理问题的有力示范动物，包括应激反应12、13、生长14、15、营养生理16、17和繁殖2。Teleosts， 其中性别类固醇主要代表雌二 （E2） 和 11- 酮睾酮 （11-KT） 在男性18，19，长期以来一直是可靠的实验模型， 以调查跨物种繁殖的一般原理.Teleosts在下丘脑脑垂体连接20、21和独特的角细胞22中表现出独特性，这有时便于解释调节机制。此外，由于远程实验既适应实验室实验又进行实地实验，因此与其他生物体相比，远程运动具有许多优势。他们相对便宜购买和维持23，24。特别是小型远程模型，如斑马鱼（Danio rerio）和日本美达卡（Oryzias latipes），是具有非常高的活性，生命周期相对较短的物种，能够快速分析基因功能和疾病机制23，从而提供了更大的优势，解决大量的生物和生理问题，考虑到许多发达的协议和遗传工具包可用于这些物种25。
在许多研究中，去除淋巴（淋巴切除术）以及血液采样技术已被用作研究许多生理问题的方法，包括它对26、27、28、鸟类29和两栖动物30种哺乳动物的脊椎动物生殖生理学的影响。虽然腺切除术对生殖生理的影响可以替代模仿性类固醇拮抗剂，如塔莫西芬和氯米芬，药物的作用似乎是不一致的，由于双模效应31，32。长期接触性类固醇拮抗剂可能导致卵巢扩大33，34，这可能会禁用观察其影响的长期目的， 由于不健康的表型.此外， 不可能进行性类固醇拮抗剂治疗后的恢复实验， 以保证某些性类固醇的具体效果.与上述点， 性类固醇对抗剂使用的其他权衡已广泛审查31，32.因此， 结核切除术今天仍然作为一个强大的技术来研究性类固醇的作用.
虽然去角切除术和血液采样技术在更大的物种中相对容易执行， 如欧洲海贝（迪森特拉丘斯拉布拉克斯）35，蓝头瓦塞（塔拉索马双发）36，狗鱼（西利奥尔希努斯犬）37和猫鱼（异种化石和克拉里亚斯浴场）38，39，他们提出了挑战时，应用在小鱼作为梅达卡。例如，使用鱼麻醉输送系统（FADS）40不太可行，而且似乎容易对小鱼造成过度的身体伤害。此外，通常用于大鱼40的腺切除术不适合需要高精度以避免过度伤害的小鱼。最后，由于获得血管的机会有限，而且这些动物的血液量很小，血液采样具有挑战性。因此，在小手术中展示造血术和血液采样的每一步的明确协议非常重要。
该协议演示了腺切除术的分步程序，随后在日本产自东亚的小淡水鱼——日本美中进行血液取样。日本的美田有一个测序的基因组，几个分子和遗传工具可用25，和一个基因性别测定系统，允许调查性差异之前，二次性特征或性腺发育良好41。有趣的是，日本美达卡拥有融合的果子虫，这与许多其他的远程物种42相反。这两种技术加起来总共只需要8分钟，将完成该物种已经存在的视频协议列表，其中包括血管标签43，垂体部分44 和脑神经元45的补丁夹，和原细胞培养46。这些技术将使研究社区调查和更好地了解性类固醇在反馈机制中的作用，以及大脑和垂体可塑性在未来。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Glass capilary</td>
			<td></td>
			<td>GD1</td>
			<td>Glass Capillary with Filament GD-1; Narishige</td>
		</tr>
		<tr>
			<td>Heparin sodium salt</td>
			<td></td>
			<td>H4784-1G</td>
			<td>Sigma-aldrich</td>
		</tr>
		<tr>
			<td>Needle puller</td>
			<td></td>
			<td>P97</td>
			<td>Flaming/Brown Micropipette puller Model P-97; Sutter Instrument</td>
		</tr>
		<tr>
			<td>Nylon thread</td>
			<td></td>
			<td>N45VL</td>
			<td>Polyamide suture, 0.2 metric; Crownjun</td>
		</tr>
		<tr>
			<td>Plastic tube</td>
			<td></td>
			<td>T9661</td>
			<td>Eppendorf Safe-lock microcentifuge tube 1.5 ml, Sigma-aldrich</td>
		</tr>
		<tr>
			<td>Razor blade</td>
			<td></td>
			<td>-</td>
			<td>Astra Superior Platinum Double Edge Razor Blades Green, salonwholesale.com</td>
		</tr>
		<tr>
			<td>Silicone capillary</td>
			<td></td>
			<td>a16090800ux0403</td>
			<td>Uxcell Silicone Tube 1 mm ID x 2 mm OD, amazon.com&#160;</td>
		</tr>
		<tr>
			<td>Tricaine</td>
			<td></td>
			<td>WXBC9102V</td>
			<td>Aldrich chemistry</td>
		</tr>
	</tbody>
</table>

gonadectomy and blood sampling procedures in the small size teleost model japanese medaka (oryzias latipes)

性类固醇，由腺体产生，在大脑和垂体组织可塑性和神经内分泌控制所有脊椎动物通过提供大脑和垂体的反馈发挥关键作用。与哺乳动物相比，Teleost鱼在组织可塑性和生殖策略变化方面具有较高的水平，似乎是研究性类固醇的作用及其作用机制的有用模型。去除性类固醇生产的主要来源使用腺切除术与血液采样测量类固醇水平已经建立和相当可行的大鱼， 是一个强大的技术来调查性类固醇的作用和影响.但是，当在小型远程模型中实施时，这些技术会带来挑战。在这里，我们描述了日本母体的妇腺切除术的分步程序，然后是血液采样。这些协议被证明是非常可行的美达卡表明高存活率， 安全的寿命和表型的鱼， 并在性别类固醇清除方面可重复性.使用这些程序结合使用这种小远程模型的其他优势将大大提高对脊椎动物中性类固醇提供的生殖神经内分泌控制和组织可塑性的反馈机制的理解。

该协议描述了在小尺寸模型远程手术（日本医疗）中执行腺切除术和血液采样的每一步。雌性卵巢切除术（OVX）后鱼类的存活率为100%（10条鱼类中的10条），而94%（18条鱼类中的17条）的雄性在兰花切除术后存活下来。同时，在进行血液采样手术后，所有（38条鱼）都幸存了下来。
沙姆操作的雌性显示卵子（图6A?...

Watch this Scientific Journal Video about Gonadectomy and Blood Sampling Procedures in the Small Size Teleost Model Japanese Medaka Oryzias latipes at JoVE.com

Gonadectomy and Blood Sampling Procedures in the Small Size Teleost Model Japanese Medaka Oryzias latipes

本文描述了一个快速协议去纳切除术和采样血液从小远程鱼，使用日本美达卡（Oryzias拉蒂普斯）作为模型，以调查性类固醇在动物生理中的作用。

小尺寸远程模型日本美达卡（奥里齐亚斯拉蒂普斯）的果纳切除术和血液采样程序

Sex steroids, produced by the gonads, play an essential role in brain and pituitary tissue plasticity and in the neuroendocrine control of reproduction in ...

gonadectomy-blood-sampling-procedures-small-size-teleost-model

Research

JoVE Journal

Biology

Gonadectomy and Blood Sampling Procedures in the Small Size Teleost Model Japanese Medaka (Oryzias latipes)

4.2K 次观看.  Norwegian University of Life Sciences. 本文描述了一个快速协议去纳切除术和采样血液从小远程鱼，使用日本美达卡（Oryzias拉蒂普斯）作为模型，以调查性类固醇在动物生理中的作用。 

视频: Gonadectomy and Blood Sampling Procedures in the Small Size Teleost Model Japanese Medaka Oryzias latipes

Noninvasive In Vivo Small Animal MRI and MRS: Basic Experimental Procedures

Small animal Magnetic Resonance (MR) research has emerged as an important element of modern biomedical research due to its non-invasive nature and the richness of biological information it provides. MR does not require any ionizing radiation and can noninvasively provide higher resolution and better signal-to-noise ratio in comparison to other tomographic or spectroscopic modalities. In this protocol, we will focus on small animal MR imaging and MR spectroscopy (MRI/MRS) to noninvasively acquire relaxation weighted 1H images of mouse and to obtain 31P spectra of mouse muscle. This work does not attempt to cover every aspect of small animal MRI/MRS but rather introduces basic procedures of mouse MRI/MRS experiments. The main goal of this work is to inform researchers of the basic procedures for in vivo MR experiments on small animals. The goal is to provide a better understanding of basic experimental procedures to allow researchers new to the MR field to better plan for non-MR components of their studies so that both MR and non-MR procedures are seamlessly integrated.

Noninvasive In Vivo Small Animal MRI and MRS: Basic Experimental Procedures

This work describes basic procedures of noninvasive small animal MRI and MRS in vivo.

无创在体内小动物的MRI和MRS：基本实验程序

Small animal Magnetic Resonance (MR) research has emerged as an important element of modern biomedical research due to its non-invasive nature and the ...

科研

生物学

Microinjection of Medaka Embryos for use as a Model Genetic Organism

In this video, we demonstrate the technique of microinjection into one-cell stage medaka embryos. Medaka is a small egg-laying freshwater fish that allows both genetic and embryological analyses and is one of the vertebrate model organisms in which genome-wide phenotype-driven mutant screens were carried out 1, as in zebrafish and the mouse. Divergence of functional overlap of related genes between medaka and zebrafish allows identification of novel phenotypes that are unidentifiable in a single species 2, thus medaka and zebrafish are complementary for genetic dissection of vertebrate genome functions.
To take advantage of medaka fish whose embryos are transparent and develop externally, microinjection is an essential technique to inject cell-tracers for labeling cells, mRNAs or anti-sense oligonucleotides for over-expressing and knocking-down genes of interest, and DNAs for making transgenic lines.

Medaka and zebrafish are complementary for genetic dissection of vertebrate genome functions. This protocol highlights the key points for successful microinjection into medaka embryos, an important technique for embryological and genetic analysis using medaka and zebrafish in a laboratory.

作为一个型号的遗传生物鳉胚胎显微注射

In this video, we demonstrate the technique of microinjection into one-cell stage medaka embryos. Medaka is a small egg-laying freshwater fish that allows ...

Dechorionation of Medaka Embryos and Cell Transplantation for the Generation of Chimeras

Medaka is a small egg-laying freshwater fish that allows both genetic and embryological analyses and is one of the three vertebrate model organisms in which genome-wide phenotype-driven mutant screens were carried out 1. Divergence of functional overlap of related genes between medaka and zebrafish allows identification of novel phenotypes that are unidentifiable in a single species 2, thus medaka and zebrafish are complementary for genetic dissection of the vertebrate genome functions. Manipulation of medaka embryos, such as dechorionation, mounting embryos for imaging and cell transplantation, are key procedures to work on both medaka and zebrafish in a laboratory. Cell transplantation examines cell autonomy of medaka mutations. Chimeras are generated by transplanting labeled cells from donor embryos into unlabeled recipient embryos. Donor cells can be transplanted to specific areas of the recipient embryos based on the fate maps 3 so that clones from transplanted cells can be integrated in the tissue of interest during development. Due to the hard chorion and soft embryos, manipulation of medaka embryos is more involved than in zebrafish. In this video, we show detailed procedures to manipulate medaka embryos.

Due to the hard chorion and soft embryos, manipulation of medaka embryos is more involved than in zebrafish. This video shows step-by-step procedures for how to manipulate medaka embryos, including dechorionation, mounting in agarose for imaging and cell transplantation for the production of chimeras. These procedures are essential to use medaka and zebrafish in a laboratory to take full advantage of their complementary features for the genetic dissection of vertebrate genome functions.

鳉胚胎和细胞移植嵌合体生成Dechorionation

Medaka is a small egg-laying freshwater fish that allows both genetic and embryological analyses and is one of the three vertebrate model organisms in ...

Whole-mount Immunohistochemical Analysis for Embryonic Limb Skin Vasculature: a Model System to Study Vascular Branching Morphogenesis in Embryo

Whole-mount immunohistochemical analysis for imaging the entire vasculature is pivotal for understanding the cellular mechanisms of branching morphogenesis. We have developed the limb skin vasculature model to study vascular development in which a pre-existing primitive capillary plexus is reorganized into a hierarchically branched vascular network. Whole-mount confocal microscopy with multiple labelling allows for robust imaging of intact blood vessels as well as their cellular components including endothelial cells, pericytes and smooth muscle cells, using specific fluorescent markers. Advances in this limb skin vasculature model with genetic studies have improved understanding molecular mechanisms of vascular development and patterning. The limb skin vasculature model has been used to study how peripheral nerves provide a spatial template for the differentiation and patterning of arteries. This video article describes a simple and robust protocol to stain intact blood vessels with vascular specific antibodies and fluorescent secondary antibodies, which is applicable for vascularized embryonic organs where we are able to follow the process of vascular development.

We introduce a whole-mount immunohistochemistry and laser scanning confocal microscopy with multiple labelling for analyzing intricate vascular network formation in mouse embryonic limb skin.

全安装胚胎肢体皮肤血管的免疫组织化学分析：一个模型系统，研究在胚胎血管分支形态发生

Whole-mount immunohistochemical analysis for imaging the entire vasculature is pivotal for understanding the cellular mechanisms of branching ...

A Noninvasive Hair Sampling Technique to Obtain High Quality DNA from Elusive Small Mammals

Noninvasive genetic sampling approaches are becoming increasingly important to study wildlife populations. A number of studies have reported using noninvasive sampling techniques to investigate population genetics and demography of wild populations1. This approach has proven to be especially useful when dealing with rare or elusive species2. While a number of these methods have been developed to sample hair, feces and other biological material from carnivores and medium-sized mammals, they have largely remained untested in elusive small mammals. In this video, we present a novel, inexpensive and noninvasive hair snare targeted at an elusive small mammal, the American pika (Ochotona princeps). We describe the general set-up of the hair snare, which consists of strips of packing tape arranged in a web-like fashion and placed along travelling routes in the pikas&#x2019; habitat. We illustrate the efficiency of the snare at collecting a large quantity of hair that can then be collected and brought back to the lab. We then demonstrate the use of the DNA IQ system (Promega) to isolate DNA and showcase the utility of this method to amplify commonly used molecular markers including nuclear microsatellites, amplified fragment length polymorphisms (AFLPs), mitochondrial sequences (800bp) as well as a molecular sexing marker. Overall, we demonstrate the utility of this novel noninvasive hair snare as a sampling technique for wildlife population biologists. We anticipate that this approach will be applicable to a variety of small mammals, opening up areas of investigation within natural populations, while minimizing impact to study organisms.

We present a noninvasive sampling approach to efficiently collect hair samples from elusive small mammals, as shown for the American pika. We demonstrate the utility of this method by extracting DNA from sampled hair and amplifying several types of molecular markers commonly used in studies of wildlife ecology and conservation.

一种无创性的发采样技术，以获得难以捉摸的小型兽类高质量的DNA

Noninvasive genetic sampling approaches are becoming increasingly important to study wildlife populations. A number of studies have reported using ...

Quantifying Single Microvessel Permeability in Isolated Blood-perfused Rat Lung Preparation

The isolated blood-perfused lung preparation is widely used to visualize and define signaling in single microvessels. By coupling this preparation with real time imaging, it becomes feasible to determine permeability changes in individual pulmonary microvessels. Herein we describe steps to isolate rat lungs and perfuse them with autologous blood. Then, we outline steps to infuse fluorophores or agents via a microcatheter into a small lung region. Using these procedures described, we determined permeability increases in rat lung microvessels in response to infusions of bacterial lipopolysaccharide. The data revealed that lipopolysaccharide increased fluid leak across both venular and capillary microvessel segments. Thus, this method makes it possible to compare permeability responses among vascular segments and thus, define any heterogeneity in the response. While commonly used methods to define lung permeability require postprocessing of lung tissue samples, the use of real time imaging obviates this requirement as evident from the present method. Thus, the isolated lung preparation combined with real time imaging offers several advantages over traditional methods to determine lung microvascular permeability, yet is a straightforward method to develop and implement.

The isolated blood-perfused lung preparation makes it feasible to visualize microvessel networks on the lung surface. Here we describe an approach to quantify permeability of single microvessels in isolated lungs using real time fluorescence imaging.

在离体血液灌注大鼠肺备量化单微血管通透性

The isolated blood-perfused lung preparation is widely used to visualize and define signaling in single microvessels. By coupling this preparation with ...

Non-Terminal Blood Sampling Techniques in Guinea Pigs

Guinea pigs possess several biological similarities to humans and are validated experimental animal models1-3. However, the use of guinea pigs currently represents a relatively narrow area of research and descriptive data on specific methodology is correspondingly scarce. The anatomical features of guinea pigs are slightly different from other rodent models, hence modulation of sampling techniques to accommodate for species-specific differences, e.g., compared to mice and rats, are necessary to obtain sufficient and high quality samples. As both long and short term in vivo studies often require repeated blood sampling the choice of technique should be well considered in order to reduce stress and discomfort in the animals but also to ensure survival as well as compliance with requirements of sample size and accessibility. Venous blood samples can be obtained at a number of sites in guinea pigs e.g., the saphenous and jugular veins, each technique containing both advantages and disadvantages4,5. Here, we present four different blood sampling techniques for either conscious or anaesthetized guinea pigs. The procedures are all non-terminal procedures provided that sample volumes and number of samples do not exceed guidelines for blood collection in laboratory animals6. All the described methods have been thoroughly tested and applied for repeated in vivo blood sampling in studies within our research facility.

Though a known model, the guinea pig currently represents a niche in experimental animal sciences and limited data is available on the execution of most procedures. Here we present four different approaches to non-terminal in vivo blood sampling techniques in either conscious or anaesthetized guinea pigs.

豚鼠非终端采血技术

Guinea pigs possess several biological similarities to humans and are validated experimental animal models1-3. However, the use of guinea pigs currently ...

Sampling Blood from the Lateral Tail Vein of the Rat

Blood samples are commonly obtained in many experimental contexts to measure targets of interest, including hormones, immune factors, growth factors, proteins, and glucose, yet the composition of the blood is dynamically regulated and easily perturbed. One factor that can change the blood composition is the stress response triggered by the sampling procedure, which can contribute to variability in the measures of interest. Here we describe a procedure for blood sampling from the lateral tail vein in the rat. This procedure offers significant advantages over other more commonly used techniques. It permits rapid sampling with minimal pain or invasiveness, without anesthesia or analgesia. Additionally, it can be used to obtain large volume samples (upwards of 1 ml in some rats), and it may be used repeatedly across experimental days. By minimizing the stress response and pain resulting from blood sampling, measures can more accurately reflect the true basal state of the animal, with minimal influence from the sampling procedure itself.

Blood samples are useful for assessing biomarkers of physiological states or disease in vivo. Here we describe the methodology to sample blood from the lateral tail vein in the rat. This method provides rapid samples with minimal pain and invasiveness.

从大鼠的侧尾静脉血液取样

Blood samples are commonly obtained in many experimental contexts to measure targets of interest, including hormones, immune factors, growth factors, ...

Noninvasive Monitoring of Lesion Size in a Heterologous Mouse Model of Endometriosis

Here, we describe a protocol for the implementation of a heterologous mouse model in which progression of endometriosis can be assessed in real time through noninvasive monitoring of fluorescence emitted by implanted ectopic human endometrial tissue. For this purpose, biopsies of human endometrium are obtained from donor women ongoing oocyte donation. Human endometrial fragments are cultured in the presence of adenoviruses engineered to express cDNA for the reporter fluorescent protein mCherry. Upon visualization, labeled tissues with an optimal rate of fluorescence after infection are subsequently chosen for the implantation in recipient mice. One week prior to the implantation surgery, recipient mice are oophorectomized, and estradiol pellets are placed subcutaneously to sustain the survival and growth of lesions. On the day of surgery mice are anesthetized, and peritoneal cavity accessed through a small (1.5 cm) incision by the linea-alba. Fluorescently labeled implants are tweezed, briefly soaked in glue and attached to the peritoneal layer. Incisions are sutured, and animals left to recover for a couple of days. Fluorescence emitted by endometriotic implants is usually non-invasively monitored every 3 days for 4 weeks with an in vivo imaging system. Variations in the size of endometriotic implants can be estimated in real time by quantification of the mCherry signal and normalization against the initial time-point showing maximal fluorescence intensity.
Traditional preclinical rodents of models of endometriosis do not allow non-invasive monitoring of lesion in real time but rather allow evaluation of the effects of drugs assayed at the end point. This protocol allows one to track lesions in real time and is more useful to explore the therapeutic potential of drugs in preclinical models of endometriosis. The main limitation of the model thus generated is that non-invasive monitoring is not possible over long periods of time due to the episomal expression of Ad-virus.

Here, we present a protocol for live-imaging of fluorescently labeled human endometrial fragments grafted in mice. The method allows studying the effects of drugs of choice on endometriotic lesion size through monitoring and quantification of fluorescence emitted by the fluorescent reporter on real time

子宫内膜异位症异种小鼠病变大小的无创监测

Here, we describe a protocol for the implementation of a heterologous mouse model in which progression of endometriosis can be assessed in real time ...

Quantification of Circulating Pig-Specific DNA in the Blood of a Xenotransplantation Model

Xenotransplantation is a feasible method to treat organ failure. However, how to effectively monitor the immune rejection of xenotransplantation is a problem for physicians and researchers. This manuscript describes a simple and effective method to monitor immune rejection in pig-to-mouse cell transplantation models and pig-to-monkey artery patch transplantation models. Circulating DNA is a potentially non-invasive biomarker for organ damage. In this study, circulating pig-specific DNA (cpsDNA) was monitored during xenograft rejection by quantitative real-time PCR (qPCR). In this protocol, porcine specific primers were designed, plasmids-containing porcine specific DNA fragments were constructed, and standard curves for quantitation were established. Species-specific primers were then used to quantify cpsDNA by qPCR in pig-to-mouse cell transplantation models and pig-to-monkey artery patch transplantation models. The value of this method suggests that it can be used as a simple, convenient, low cost, and less invasive method to monitor the immune rejection of xenotransplantation.

In this protocol, porcine specific primers were designed, plasmids-containing porcine specific DNA fragments were constructed, and standard curves for quantitation were established. Using species-specific primers, cpsDNA was quantified by qPCR in pig-to-mouse cell transplantation models and pig-to-monkey artery patch transplantation models.

异种移植模型中循环猪特异性DNA的定量

Xenotransplantation is a feasible method to treat organ failure. However, how to effectively monitor the immune rejection of xenotransplantation is a ...

Methods Article

小尺寸远程模型日本美达卡（奥里齐亚斯拉蒂普斯）的果纳切除术和血液采样程序