作者感谢的实验帮助发展中国家从虹膜埃德曼，芭芭拉·冯·博伊斯特，朱莉娅玛丽亚·马丁内斯·戈麦斯小鼠淋巴免疫的方法。还要感谢玛吉阿拉斯和尼古拉Cesarovic您让我们使用他们的手术剧场这个视频制作。

1。小鼠麻醉<ol><li>在缓冲液混合氯胺酮（分离性麻醉剂）和甲苯噻嗪（镇静，镇痛）准备麻醉剂。氯胺酮和甲苯噻嗪最终溶液中的浓度是12.5和2毫克/毫升。 </li><li>使用注射器用25-30号针头通过腹膜内给药注射麻醉​​剂将小鼠。使用0.1 ml/10克的机身重量。 </li><li>为了防止干燥出的角膜应用眼用软膏对小鼠的眼睛。 </li><li>确保鼠标充分捏它的脚或脚趾与镊子麻醉。如果鼠标不与反射的捏反应，继续手术。如果鼠标反应捏有反射或肌肉收缩，等待1-2分钟，然后重复捏测试。如果鼠标仍然反应，请更换鼠标采用了新的动物，并从步骤1.2开始重复步骤。 </li></ol> 2。我的手术鼠标的腹股沟区ncision <ol><li>把鼠标在它的后面。 </li><li>湿1腹股沟区域用消毒乙醇（70％）。为了提供更好的消毒，腹股沟区可被剃光，除去皮毛消毒前用乙醇。 </li><li>取后腿和弯曲髋关节，产生髋关节的一个大约90°的角度。 </li><li>使用弯曲microdissecting钳，取小鼠皮肤在腹股沟区，其中髋关节弯曲和拉湿皮肤略有上升。 </li><li>同时保持肌肤与镊子，通过使用手术剪皮肤切开一个小切口（&lt;5毫米）。 </li><li>将闭合的剪刀的尖端插入切口，并进一步通过打开的剪刀，同时保持它的切口内打开切口。这会导致皮肤撕裂，并应当产生一个直径小于10毫米。注：具有灵活的光导冷光源，可以用来提高知名度。在较老的小鼠，特别是男性，淋巴结的识别可能是困难的，由于在腹股沟区过多的脂肪组织。 </li></ol> 3。淋巴管内注射<ol><li>准备一个注射器（0.5毫升或更少）用28-30ģ皮下注射针，短斜面可能是优选的一长斜面。吸出10μl的疫苗被注射并保证该注射器是自由的空气。 </li><li>本地化与弯钳的帮助和闭合剪刀的尖端处的腹股沟淋巴结。淋巴结会出现灰内越发白的脂肪组织，并且可以通过传入和传出的毛细血管进入和离开淋巴结进行进一步鉴定。 </li><li>受弯钳的分支之间保持它固定的淋巴结。 </li><li>就拿注射器和针头插入朝上的斜面淋巴结;确保整个斜面进入淋巴结。 </li><li>注射了疫苗（10微升）。如果淋巴结点头e的肿胀（吹），注射可以假设成功。如果针头未放置足够深到淋巴结，部分或整个疫苗可能会泄漏出来，而不是进入淋巴结。如果针头插入过深，注射的疫苗将被释放皮下下面的淋巴结。在后一种情况下，重复注射或用新的未处理的小鼠代替鼠标。 </li></ol> 4。通过缝合关闭切口<ol><li>打开无菌手术缝合线的包。 </li><li>抓住的缝合针与针座的针体的远端部分。通过挤压它，直到第一棘轮渔获拧紧持针器。 </li><li>使用镊子抓住皮肤的切口的一侧（取决于外科医生的偏好，齿形或不带齿的钳或皮肤钩可以使用）。插入针约 2-3毫米的切口的边缘上，从皮肤的外部进入。 </li><li>洛斯烯保持与钳子的皮肤和把握切口用钳子的相对侧。插入针约 2-3毫米的切口的边缘上，从皮肤的内侧进入。 </li><li>松开持针器，把握针端与钳。拉带通过皮肤胎面针，而使胎面的方便长度的第一针插入点之外（如果几个小鼠是被处理，留下较短端5-10小鼠可被操作上与一个缝合针/踩）。 </li><li>根据标准协议操刀制作的首选结。一个线圈就足够了，如果切口不大于10mm。对于较大的切口，进行第二次缝合。 </li></ol> 5。运行后的治疗<ol><li>将鼠标放置在笼子里，用纸巾覆盖保暖。当处理多个老鼠，把它们接近对方，以保持温暖。或者，将鼠标在一个温暖的垫子，直到他们才如梦初醒起来。 </li><li>观察小鼠直到他们醒来。 </li><li>每天观察小鼠的问候伤口愈合以及其他临床症状，如伤口感染。伤口通常足以在两天内关闭，并在该过程七天痊愈。 </li></ol>

<ol>
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TMK被命名为专利涉及在人淋巴免疫疗法的发明者。 TMK一直是科学顾问，并已收到差旅费由ImVisioN有限公司，Cytos生物技术，Mannkind公司，并XBiotech美国公司PJ没有任何利益冲突披露。

淋巴内免疫接种和免疫治疗已被证明是合适的两个抗体应答和T细胞应答的刺激。这表现这个视频文章中，接种疫苗的淋巴管内的程序是一个快速简便的方法刺激强烈的免疫反应的小鼠。一个训练有素的外科医生可以在3-4分钟执行程序。这次会议也可两名外科医生，其中一个通常做麻醉和缝合，第二个外科医生在做切口和注射之间共享。该过程的效率，更取决于运营商以外的免疫方法，尤其是当外...

疫苗通常是皮下或肌内注射对免疫应答的刺激。这个程序的成功需要疫苗的有效引流淋巴结其中抗原呈递细胞可以与淋巴细胞相互作用用于生成的T-和B-细胞应答。此外，强度和诱导免疫反应的类型也取决于这种相互作用的密度或频率以及微环境本身，细胞因子，特别是内容。作为注入到外周组织的疫苗只有一小部分到达淋巴结，小鼠和人类通过直接注射该疫苗进入淋巴结的疫苗，该网站被产生的免疫反应，已被执行。在男人中，程序是通过超声波，一个程序也可以用于可视化和诊断在淋巴系统成像剂的给药指导。在小鼠体内，是侵入性检查。在这里，一个小的（5-10毫米）incision被制成在麻醉的动物1的腹股沟区域，淋巴结进行定位和固定钳，和10-20微升的疫苗的一定体积的视觉控制下注入; 10微升用于第一注射和在年轻小鼠用小淋巴结，而20微升可以注入更旧的淋巴结或已催芽的小鼠，其具有较大的淋巴结。切口可闭合使用外科缝合线的单个针迹。通过该方法，将小鼠已接种的质粒DNA 2,3，信使RNA 4，1,3,5,6肽，蛋白质7-10，颗粒11，细菌12以及辅助剂7,13，和免疫大幅改善对所有类型的疫苗反应已​​被观察到。接种淋巴方法特别适合在常规疫苗接种产生的免疫力不足或可用疫苗的量是有限的或版本的情况下Ÿ昂贵。在人类，免疫的淋巴管内的方法已被应用到过敏患者14,15或癌症患者16-21。虽然目前的看法是，在淋巴管内的方法比其它可注射的方法，如肌内和皮下注射更具侵入性的，疼痛感觉不大于静脉穿刺15后更高。预期淋巴内注射疫苗将成为一种替代或补充的预防和治疗尤其是疫苗的其它方法。本文详细阐述了如何淋巴管内接种疫苗的过程中小鼠进行。描述的所有程序，批准了苏黎世和州兽医局，并根据瑞士联邦指导方针以及用于科学目的的动物的保护指令执行。

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			<td height="42" style="height:42px;width:233px;">Ketamine (Ketasol-100)</td>
			<td style="width:135px;">Graeub AG, Switzerland</td>
			<td style="width:119px;"></td>
			<td style="width:167px;">Anesthetics</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:233px;">Xylazine (Rompun)</td>
			<td style="width:135px;">Bayer, Germany</td>
			<td style="width:119px;"></td>
			<td>Anesthetics</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">Viscotears Eye-Gel</td>
			<td style="width:135px;">Novartis, Switzerland</td>
			<td style="width:119px;"></td>
			<td>To keep eyes from drying out during anesthesia.</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:233px;">BD Micro-Fine 0.5 ml</td>
			<td style="width:135px;">BD Medical, France</td>
			<td style="width:119px;"></td>
			<td>29 G Insulin syringes with permanently attached needles</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:233px;">6-0 Dermalon Monofilament nylon</td>
			<td style="width:135px;">Covidien, MA, USA</td>
			<td style="width:119px;"></td>
			<td>For sutures (0.7 metric, 18G, 45 cm, Blue)</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:233px;">Curved forceps, 4.5 inch</td>
			<td style="width:135px;">Polymed, Switzerland</td>
			<td style="width:119px;"></td>
			<td>For incision and holding of lymph node</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:233px;">Straight surgical scissors, 4.5 inch</td>
			<td style="width:135px;">Polymed, Switzerland</td>
			<td style="width:119px;"></td>
			<td>For incision</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:233px;">Needle holder, 5.5 inch</td>
			<td style="width:135px;">Polymed, Switzerland</td>
			<td style="width:119px;"></td>
			<td>To close incision with suture</td>
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intralymphatic immunotherapy and vaccination in mice

疫苗一般是皮下或肌内注射对免疫应答的刺激。这样做的成功需要疫苗的有效引流淋巴结，其中抗原呈递细胞可以与淋巴细胞相互作用的一代想要的免疫反应。的强度和诱导免疫反应的类型也取决于相互作用的密度或频率以及微环境，细胞因子，特别是内容。作为外周注射疫苗仅一分钟级分到达淋巴结，小鼠和人类的疫苗通过直接注射疫苗被执行到腹股沟淋巴结， 即淋巴内注射。在男人中，程序是由超声引导。在小鼠中，小（5-10毫米）切口，在麻醉的动物的腹股沟区域，淋巴结进行定位和固定钳，和10-20微升的疫苗的一定体积的视觉控制下喷射。切口闭合使用手术缝合线单针。小鼠接种的质粒DNA，RNA，肽，蛋白质，颗粒，细菌以及佐剂，并观察到对所有类型的疫苗的免疫应答的强改善。接种淋巴方法特别适合在常规疫苗接种产生的免疫力不足或可用的疫苗量有限的情况下。

在小鼠淋巴内注射的方法，尽管在手术的性质，是直线前进和比较快的。一个训练有素的人能在3-4分钟执行程序。是与一针封闭切口通常在两天内愈合（ 图1） 淋巴内注射疫苗或免疫接种用的mRNA被执行时，质粒DNA，肽，蛋白质，病毒和细菌。 图2示出了免疫与蛋白质磷脂酶A2（PLA2），主要蜜蜂毒液变应原后的抗体产生。 CBA小鼠进行免疫在0天，14，和2...

Watch this Scientific Journal Video about Intralymphatic Immunotherapy and Vaccination in Mice at JoVE.com

Intralymphatic Immunotherapy and Vaccination in Mice

预防性和治疗性疫苗往往不能刺激强烈的免疫反应，由于疫苗的周向引流淋巴结细胞和免疫细胞的参与因而不佳。通过直接注射疫苗到淋巴结的，所谓的淋巴内注射，疫苗的有效性，可以大大地改善和疫苗的剂量可以减少。

淋巴免疫治疗和疫苗接种小鼠

Vaccines are typically injected subcutaneously or intramuscularly for stimulation of immune responses. The success of this requires efficient drainage of ...

intralymphatic-immunotherapy-and-vaccination-in-mice

Research

JoVE Journal

Immunology and Infection

22.9K 次观看.  University Hospital Zurich. 预防性和治疗性疫苗往往不能刺激强烈的免疫反应，由于疫苗的周向引流淋巴结细胞和免疫细胞的参与因而不佳。通过直接注射疫苗到淋巴结的，所谓的淋巴内注射，疫苗的有效性，可以大大地改善和疫苗的剂量可以减少。 