We would like to thank Dr. Marc Liesa, Boston University School of Medicine, helpful discussions; Ms. Karin Murphy, Ms. Heather Gregory, and Mr. John Stone, all from Wake Forest School of Medicine, for helpful technical assistance in the development of this protocol.

注：说明是经医学维克森林学校的机构审查委员会的协议。以书面形式获得知情同意书。所有与会者都健康老年男女的成年人（65-79岁），身体质量指数从23-35。 1.骨骼肌活检<ol><li>如前所述，9后的O / N高速执行清晨的所有活组织切片检查。询问受试者服用阿司匹林，处方药和非处方非甾体抗炎药，或其他化合物可能影响出血，血小板，或瘀伤的一周前的活检来避免。请参与者也从任何剧烈活动为避免前活检至少36小时。 注：肌肉是从股外侧采用与1％利多卡因局部麻醉下经皮针的穿刺活检技术获得。无并发症或o疗法报道从过程的不良事件都发生在我们的临床研究单位。 </li><li>注：所描述的活检过程是改编自该伯格斯特龙10。 <ol><li>简单地说，本地管理1％利多卡因注意不要渗入肌肉。用10分钟的等待时间按照此允许足够的麻木。 </li><li>取活检从肌肉（插入和原点之间的肌肉的中间区域）避免筋膜和myotendonous区的腹中。 <ol><li>利用经皮针（侧切割窗口，内切汽缸吸入可重复使用的辅助装置），并跟随筋膜“啪”的招牌作为指导或阻力。 </li><li>估计的深度与麻醉针，然后再次感觉到它与窄手术刀刀片和通过筋膜进行4-5毫米切口。通过切口推进针，直到它被插入到肌肉。 </li><li>收集与窗口多个样品转向不同的方向。使用60毫升注射器而进退肌肉样品进经皮针两到四倍以不同方向施加连续吸力。中止吸并取出针。 注：每个通（插入和拔出针）花费时间不超过一分钟。 </li><li>有一个助手适用于公司的压力，穿刺部位5分钟建立止血。从吸气管拔掉针头，小心地从窗口桶取出的肌肉样本。 </li></ol></li><li>使第二通如果更多的肌肉，需要通过重复上述过程。 </li><li>使用镊子除去从肌肉样品的可见的血块，称重样品，并立即在含有冰冷的DPBS的管放置。 注：使用这​​种方法的平均产量为150±20毫克。 </li></ol></li></ol> 2.线粒体隔离 &lt;OL&gt; <li>新鲜制备Chappel-佩里（CP）和线粒体测定溶液（MAS）的缓冲器（ 表1）在实验中，或等分的一天，并将其存储在-20℃。制备在二甲基亚砜中的化合物为2.5毫米的浓度和等分并储存在-20℃。使用储存在-20℃下2个月内从作成日的缓冲液和化合物等分试样。 </li><li>删除使用锋利的剪刀和镊子样品可见结缔组织;如果需要的话用解剖显微镜对这一步骤。彻底洗净标本3-4次，用冰冷的DPBS缓冲以除去血液。保持在冰冷的DPBS和过程中的样品，尽快，注意不要超过从活检超过45分钟。采取预防措施，以小心地从肌肉样本中删除任何肌腱或脂肪组织。 </li><li>立即用无菌剪刀印章的肌肉组织成细片，并在500微升暂停含公关1毫升CPIoteinase（枯草菌），在0.2毫克/克组织的浓度。用5分钟在室温下孵育遵循这一点，然后转移到冰上。 </li><li>均质使用自动均质与枯草菌处理的碎组织。请在此过程中冰的样本。均质每个组织样本4次，每次时间为2秒的脉冲，使用自动匀浆在10,000 rpm的速度设置。洗探针，用70％的乙醇，随后通过组织之间的蒸馏水。 </li><li>与CP I（500微升至1毫升）和CPⅡ的2倍体积（1ml至2ml）中的等体积的洗匀浆组织，并在600×g离心，4℃收集在一个离心管中，离心分离机的内容10分钟。通过湿纱布传递上清液，收集滤液弃沉淀，从而消除了大多数非线粒体分数。 </li></ol> 3.洗涤的线粒体<ol><li>从上述步骤离心上清液10，000×g离心，4℃，10分钟。暂停在4ml的CP二缓冲器，并进一步离心沉淀以10,000×g离心，4℃，10分钟。 注：在极少数情况下，薄血颗粒形成如下线粒体沉淀。在这种情况下，通过温和抽吸与CPII缓冲器移除线粒体沉淀。可避免彻底洗去血液在步骤2本步骤。 </li><li>暂停的CP I缓冲2毫升得到的粒料。在这个阶段使用一个小的等分试样，从该悬浮液中蛋白质的估计。重悬的其余样品中的CP I缓冲液和离心如上。暂停在线粒体测定溶液（MAS）的最小量（200微升）的最终的沉淀。 注：蛋白质测定是在这个阶段，因为CP I缓冲区没有BSA，而MAS其中样本将被暂停后确实有BSA在里面。 </li></ol> 4.评估线粒体含量测定蛋白质浓度用BCA蛋白检测试剂盒<p类=“jove_content”&gt;注：使用此浓度来计算用于装载线粒体的量到24孔微孔板进行呼吸计测量，或用于蛋白质印迹实验。考虑到稀释因素（10），用于蛋白质浓度的计算。 5.所描述的罗杰斯，GW 等执行XF检测。12 注：可视化的O 2消耗率（OCR）在皮摩尔O 2 /分钟，或O 2，pH值在数据输出绝对水平。 <ol><li>使用1倍的MAS以制备化合物被注入。添加10倍浓度的化合物的端口的AD，得到的最终浓度如下：端口A，ADP [腺苷5&#39;二磷酸，2mM的50微升]; B端口，寡霉素（2μM，55微升）;端口C，FCCP [羰基氰4-（三氟甲氧基）苯腙，6微米，60微升]和端口D，2μM抗霉素A（65微升）。准备足够的体积为孔的所需数量的化合物。 </li><li>通过滴定测定线粒体的最佳量。例如，负载1.0微克，2.5微克和5.0微克的含基材冰冷1X MAS 50微升体积每孔线粒体。为了尽量减少井之间变化，第一稀释10倍的线粒体在寒冷的1个MAS +基板。接着，提供50微升此悬浮液到每个孔中（除了背景校正孔）。 </li><li>离心板在2000×g离心，20分钟，在4℃。离心后，轻轻地加入450微升1×MAS +琥珀酸盐（10毫摩尔）和鱼藤酮（2μM）（初始条件;见下文），以每孔中。查看在显微镜下的线粒体，以确保均匀的遵守现有井到板转移到XF分析器。 注意：用于将依赖于由任一ETC复合物I或复合体II的呼吸是否被驱动的初始条件。对于复杂II驱动的呼吸，用succinate（10毫米）和鱼藤酮（2μM）作为初始条件。鱼藤酮块复合物I和琥珀提供燃料复合物II（琥珀酸脱氢酶）。来研究呼吸由复合物I驱动，使用丙酮酸和苹果酸盐，各为5毫米或谷氨酸和苹果酸盐的每个的终浓度为10 mM的终浓度为初始条件。后者可以帮助区分之间的三羧酸循环和基板的运输功能障碍。为了研究通过呼吸既复杂我和复杂II驱动，包括丙酮酸和琥珀酸无鱼藤酮或苹果酸作为初始条件。使用棕榈酰肉毒碱作为底物为β氧化。 </li><li>使用呼吸通过编程它如先前描述的12顺序地实时测量的线粒体呼吸。使用设置为表2中提供的呼吸。 注：不同的线粒体呼吸状态的简要说明如下： </li><li&gt;国2 =与基板结合目前的状态;国家3 =在ADP饱和的存在磷酸呼吸;国家4 O =非磷酸化呼吸诱导寡;国家3U =最大的​​解偶联呼吸作用的解偶联剂FCCP刺激;经过复杂的III抑制抗霉素-A的剩余非线粒体呼吸呼吸=。还应当指出的是，OCR测量的组合长度与线粒体浓度可以通过在测量期间消耗氧气影响的数据。 </li></ol> 6.免疫印迹注：由Western印迹确定线粒体标记COX IV和整个组织GAPDH保证线粒体富集的最终样品中<ol><li>分离的分离的线粒体，整个组织提取物由12％十二烷基硫酸钠聚丙烯酰胺凝胶。 </li><li>蛋白质转移到聚偏二氟乙烯（PVDF）膜。</li><li>孵育COX IV抗体（1：20000），其次是辣根过氧化物酶（HRP）缀合的二抗。对于GAPDH的决心，剥离印迹和探针与GAPDH单克隆抗体（1：2000）。 注：如果在ER污染差异是令人关注的，包括ER标志物如ERp72，钙联接蛋白，钙网蛋白或在分析中。 </li></ol>

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The author, George Rogers, is an employee of Seahorse Bioscience that produces the instrument used in this article. Open Access fees were supported by Seahorse Biosciences.

分离线粒体通常用在该检查ETC功能的作用，以及其他线粒体活动，包括基片运输和TCA循环功能研究。用孤立的细胞器呼吸计测定允许直接检查氧化磷酸化的基本过程和等内在特性分离线粒体的相比，全细胞或透化的肌纤维呼吸计仿形具有相对简单的数据解释的优点和没有来自非线粒体过程或改变线粒体质量/生物发生“干预”的。数据标准化是基于线粒体蛋白质含量，从而使简单的交叉对比样本之间的线粒体。分离线粒体的呼吸计谱是当研究的目的是确定潜在的机制和识别特定目标，如ETC成分的优选的方法S /复合物，或线粒体转运机制。 描述的是一种协议，肌肉活检和线粒体功能的孤立小组织样本。这种方法产生因利用自动化均质用户之间可重复的结果与手工操作DOUNCE均质。线粒体的分离可以与肌肉组织的少至20毫克的执行。的，可从该样本大小来获得分离的线粒体的量是足以运行海马板为基础的呼吸测量，同时留下剩余的线粒体为其他实验和储存用于进一步分子分析。可以注意到，该方法可以被转换为XF 96中，即使少量的线粒体的可使用（每孔1-2微克）。 隔离线粒体几个协议依赖于DOUNCE均质初始组织破坏。这种方法的缺点是在实际操作上的初始组织的性质同质化。在均质杵的力量和速度运营商6之间显著不同。这可能会导致在实验对实验变异，以及实验室到实验室变化，并导致难以在实验之间进行比较的数据。这是在人类干预研究特别关注的，当从参与者数据被收集在单独的时间点，通常前和后处理，并有可能在多个位点。我们使用自动均化器进行更一致的方法能产生具有有限的人对人的变化更可再现的结果。制备的速度也使得这种方法适合于处理多个样品在同一时间。通常情况下，最多三个实验可以在一天中进行。 此处所描述的技术的潜在限制因使用分离的细胞器，并使用板为主的格式。例如，皮卡德等 。有妖strated是孤立的线粒体拥有的不同从根本上不同于完整线粒体透肌纤维的功能特性。他们提出，线粒体的分离技术导致改变的生物能的功能，例如相比于透肌纤维伴随着更大的活性氧的产生13显著增加RCR。相比透肌纤维，线粒体的隔离并需要较长的准备时间。而且，蜂窝含量损失减少生理相关性，而这被保持在全细胞和甚至透纤维。与所描述的技术允许利用板系呼吸测量复制每个样品运行。然而，线粒体必须遵守的每个孔的底部。该构造是从它们的正常环境不同，并且可能影响功能特性。此外，应当注意的是，使用该协议线粒体隔离，疗法E可以仍然是从内质网（ER）中的线粒体制剂污染。在ER污染的差异可能会影响线粒体产量和影响结果的决心。 总之，这项研究中提出的数据说明，证实线粒体从使用此过程组织中分离出有功能活性及可用于研究/需要高品质的分离线粒体骨骼肌样品的最小量的应用程序。这种方法的优点是：ⅰ）也能够从大量的骨骼肌的最小隔离线粒体，ⅱ）的方法是快速，iii）与所述基板的技术，有可能在同一时间运行多个样本，和四）有足够的富余组织和孤立的线粒体生物能量检测的样品储存及其他分子生物学研究之后。

线粒体是细胞中的主要能源生产场地和在老化的重要作用，以及各种年龄相关病症如心血管疾病，阿尔茨海默氏病，糖尿病，癌症和肥胖症。分离线粒体的呼吸计谱提供的电子传输链（ETC）的函数直接分析，并已显著我们线粒体生物学的理解和其在健康和疾病中的作用作出了贡献。分离线粒体用于研究等，基板运输，ATP合酶活性，质子泄漏等在这个手稿中描述的方法已被优化，以允许线粒体从人受试者获得骨骼肌组织活检分离的呼吸计分析生物能量学的各个方面。在这个手稿中描述的活检协议已经利用我们的工作人员在过去的12年。我们的团队已经完成了超过700程序，对不同年龄的成年人，高达90岁，和没有任何不良的安全问题的各种慢性疾病的条件。此协议的一个重要方面是，它是专门设计来利用组织的最小量，从而促进其在临床研究中使用。 各种协议已被开发用于分离线粒体。费尔南德斯-Vizarra 等 1,2-描述用于从各种大鼠组织以及培养细胞中分离线粒体的方法。加西亚Cazarin 等 3报道从骨骼肌从大鼠和小鼠中分离线粒体的方法。一种用于从鼠脑分离线粒体方法也有报道通过西亚斯-冈萨雷斯等人 4毛等人的 5报道使用barocycler和/或PCT粉碎机分离线粒体的方法。近日，弗兰科等人 6日报道隔离使用抗高浓缩铀线粒体的方法-TOM22磁珠。 而这些协议产生极好的效果，而在这个手稿中描述的方法的组织尺寸的要求是很高的。例如，Gross 等。5用1.5-1.8克腓肠肌的，并且约2g的肾脏组织。同样，佛朗哥等 6使用500毫克鼠肝组织。根据我们的经验，一般收益率从骨骼肌（ 股外侧 ）穿刺活检预期范围从100-200毫克。在20-50毫克使用此处描述的协议的肌肉组织的评估线粒体功能的能力允许用户每次活组织检查和存储的样本进行多次评估，以便在其他分子生物学实验将来使用。这在临床研究和其他需要勤于采用样本研究的一个关键功能。应当注意的是，先前冷冻线粒体都不好学习耦合呼吸由于OUTEř线粒体膜损伤，细胞色素C活性丧失。我们的方法已被改编及由Chappell的和佩里7公开的方法进行修改。 使用这个手稿中描述的方法，我们最近报道说，线粒体的呼吸速率曲线，从人股外侧隔离直接与体能，测量步速8相关。

<table border="0" cellpadding="0" cellspacing="0" width="593">
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		<tr height="21">
			<td height="21" style="height:21px;width:279px;">Equipment</td>
			<td style="width:201px;"></td>
			<td style="width:115px;"></td>
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		<tr height="21">
			<td height="21" style="height:21px;">Name of the Equipment</td>
			<td>Company</td>
			<td></td>
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			<td height="21" style="height:21px;">Homogenizer Bio-Gen PRO200</td>
			<td>BioExpress</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">Eppendorf Centrifuge 5804 R</td>
			<td>Fisher Scientific</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">Deepwell late Rotor&#160;</td>
			<td>Fisher Scientific</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">6mm University College Hospital Needle</td>
			<td>Cadence</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">60cc syringe</td>
			<td>Fisher Scientific</td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;">96-well plate reader</td>
			<td>Tecan (Genios-basic)</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Seahorse XF 24-3 analyzer</td>
			<td>Seahorse Biosciences, Inc.</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Protein gel system</td>
			<td>Life Technologies (Invitrogen)</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Kodak Gel Logic 112</td>
			<td>Carestream Health, Inc</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Kodak camera assembly</td>
			<td>Carestream Health, Inc</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;"></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Consumables</td>
			<td></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Name&#160;</td>
			<td>Company</td>
			<td>Catalog Number</td>
		</tr>
		<tr height="20">
			<td height="41" rowspan="2" style="height:41px;">XF24 V7 Cell Culture Microplate and XF24 sensor cartridge</td>
			<td rowspan="2">Seahorse Bioscience</td>
			<td>100850-001</td>
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		<tr height="21">
			<td height="21" style="height:21px;">100867-100</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Potassium hydroxide (KOH)</td>
			<td>Sigma-Aldrich Co</td>
			<td>221473</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Hydrochloric acid (HCl)</td>
			<td>Acros</td>
			<td>12421-0010</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Dulbecco&#39;s Phosphate buffered saline&#160;</td>
			<td>Lonza</td>
			<td>17-512F</td>
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		<tr height="21">
			<td height="21" style="height:21px;">Potassium chloride (KCl)</td>
			<td>Fisher Scientific</td>
			<td>P333</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MOPS</td>
			<td>Fisher Scientific</td>
			<td>BP308</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">EDTA</td>
			<td>Fisher Scientific</td>
			<td>BP118</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;">Magnesium sulfate (MgSO4)</td>
			<td>Sigma-Aldrich Co</td>
			<td>M7506</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ATP</td>
			<td>Sigma-Aldrich Co</td>
			<td>A9187</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Fatty acid-free BSA</td>
			<td>Calbiochem</td>
			<td>126575</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Sucrose</td>
			<td>Sigma-Aldrich Co</td>
			<td>S0389</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Bacterial proteinase</td>
			<td>Sigma-Aldrich Co</td>
			<td>P-8038</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">D-Mannitol</td>
			<td>Sigma-Aldrich Co</td>
			<td>M9546</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;">KH2PO4</td>
			<td>Fisher Scientific</td>
			<td>P284</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;">Magnesium chloride (MgCl2)</td>
			<td>Sigma-Aldrich Co</td>
			<td>M9272</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HEPES</td>
			<td>Sigma-Aldrich Co</td>
			<td>H3784</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">EGTA</td>
			<td>Sigma-Aldrich Co</td>
			<td>E3889</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">BCA protein assay kit</td>
			<td>Sigma-Aldrich Co</td>
			<td>PI23227</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Succinic Acid*</td>
			<td>Sigma-Aldrich Co</td>
			<td>S3674</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Pyruvic acid*</td>
			<td>Sigma-Aldrich Co</td>
			<td>P5280</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Malic acid*</td>
			<td>Sigma-Aldrich Co</td>
			<td>2288</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ADP(K+ salt)*</td>
			<td>Sigma-Aldrich Co</td>
			<td>A5285</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">XF Cell mito stress test kit</td>
			<td>Seahorse Biosciences</td>
			<td>101706</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Tween-20</td>
			<td>Santa Cruz Biotechnology, Inc.&#160;</td>
			<td>SC-29113</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NuPAGE 12% Bis-Tris Gel</td>
			<td>Life Technologies (Invitrogen)</td>
			<td>NP0343BOX</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Immobilin Transfer Membranes (0.45um)</td>
			<td>Millipore</td>
			<td>IPVH20200</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">MOPS SDS Running Buffer (20X)-500 ml</td>
			<td>Life Technologies (Invitrogen)</td>
			<td>NP0001</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">NuPAGE Transfer Buffer (20X)-1 liter</td>
			<td>Life Technologies (Invitrogen)</td>
			<td>NP0006-1</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Primary antibodies (mAB to VDAC1/Porin)</td>
			<td>Abcam</td>
			<td>ab14734</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Primary antibodies (mAB to GAPDH)</td>
			<td>Abcam</td>
			<td>ab9484</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Anti-Mouse IgG (Goat), HRP-labeled</td>
			<td>PerkinElmer</td>
			<td>NEF822E001EA</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Anti-Rabbit IgG (Goat), HRP-labeled</td>
			<td>PerkinElmer</td>
			<td>NEF812E001EA</td>
		</tr>
		<tr height="20">
			<td colspan="3" height="20" style="height:20px;">*ADP, succinic acid, pyruvic acid, and malic acid should be adjusted to pH 7.4 with KOH only&#160;</td>
		</tr>
	</tbody>
</table>

preparation and respirometric assessment of mitochondria isolated from skeletal muscle tissue obtained by percutaneous needle biopsy

分离线粒体的呼吸计谱是常用来调查电子传递链的功能。我们描述了使用外通量（XF）分析方法获得人股外侧肌的样品，从骨骼肌组织中极少量隔离线粒体和板基础透气性分析。使用1.0，2.5和5.0微克线粒体获得呼吸速率曲线的比较表明，1.0微克足以测量呼吸和5.0微克提供基于标准误差比较最一致的结果。分离线粒体线粒体标记COX IV和非线粒体组织标记的GAPDH的免疫印迹分析表明，有使用该协议限定非线粒体污染。研究线粒体呼吸测量肌肉组织低至20毫克的能力，使用户能够利用在多个临床研究终点个体活检研究项目。

图1示出了整个协议的详细流程图。 COX IV / GAPDH（ 图2）的蛋白印迹分析轮廓描绘线粒体蛋白质，COX IV，和非线粒体标记的GAPDH的表达。双方COX IV和GAPDH的表达是很明显的，在整个肌肉溶解产物。经过线粒体正在使用这个协议描述的技术分离出来，COX IV频段依然明显，而GAPDH不存在相同的曝光。更长时间的曝光可能会发现一个微弱的GAPDH带。这些印迹表明，分离的线粒体具有最小非线粒体污染。此外，在分离线粒体COX IV的表达是样品之间是一致的。 图3示出了使用1.0微克，2.5微克和5.0微克线粒体由复合物II（琥珀酸和鱼藤酮）从动典型呼吸速率曲线。正如预期的那样，整个OCR增加瓦特第i个较高数额的线粒体。对于该测定计算出的呼吸控制率（RCR）为7.95，表明线粒体制剂是高质量的。此外，国家3U OCR比状态3略高，证实线粒体质量。 为了分析不同量的线粒体时，比较结果的一致性，我们进行了方差分析（方差分析）和计算出的平方（SS），为方差的实际值的使用的总和1.0微克，2.5微克，每5.0线粒体微克加载井（ 图4）。 SS提出的状态2，状态3，状态3U，抗霉素A，和RCR。对于国家2和国家3次测量，单因素方差分析有统计学显著（P &lt;0.01或P &lt;0.05）。同样，方差分析的一种方法是为抗霉素和RCR（P &lt;0.01或P &lt;0.0001统计学显著，分别为无显著差异见过的群体之间的状态3U。钍ESE结果表明，5.0微克，每孔线粒体，得到最低的SS相对于其他的浓度和在24 XF系统使用与我们的人口参与者的最佳量。 图5作为一个导向，以指示多少线粒体蛋白可基于初始肌肉样本大小可以预期的。正如预期的那样有肌肉（毫克）处理的量和最终样品的总线粒体蛋白含量（mg）之间有很强的相关性。 <table border="0" cellpadding="0" cellspacing="0"><tbody><tr><td colspan="2"> Chappel -佩里缓冲I（CPI） </td></tr><tr><td>化学</td><td>浓度</td></tr><tr><td>氯化钾</td><td> 100毫米</td></tr><tr><td> MOPS </td><td> 50毫米</td></tr><tr><td> EDTA </td><td> 1毫米</td></tr><tr><td> 硫酸镁 </td><td> 5毫米</td></tr><tr><td> ATP </td><td> 1毫米</td></tr&gt; <tr><td> pH值</td><td> 7.4 </td></tr><tr><td></td><td></td></tr><tr><td colspan="2"> Chappel -佩里缓冲II（CPII） </td></tr><tr><td>化学</td><td>浓度</td></tr><tr><td>氯化钾</td><td> 100毫米</td></tr><tr><td> MOPS </td><td> 50毫米</td></tr><tr><td> EDTA </td><td> 1毫米</td></tr><tr><td> 硫酸镁 </td><td> 5毫米</td></tr><tr><td> ATP </td><td> 0.2毫米</td></tr><tr><td>脂肪酸免费BSA </td><td> 0.50％ </td></tr><tr><td> pH值</td><td> 7.4 </td></tr><tr><td></td><td></td></tr><tr><td colspan="2"> 线粒体分析解决方案（MAS）（2X） </td></tr><tr><td>化学</td><td>浓度</td></tr><tr><td>蔗糖</td><td> 35毫米</td></tr><tr><td>甘露醇</td><td> 110毫米</td></tr><tr><td> KH 2 PO 4 </td><td> 2.5毫米</td></tr><tr><td> 氯化镁 </td><td> 2.5毫米</td></tr><tr><td> HEPES </td><td> 1.0毫米</td></tr><tr><td> EGTA </td><td> 0.5毫米</td></tr><tr><td>脂肪酸免费BSA </td><td> 0.10％ </td></tr><tr><td> pH值</td><td> 7.4 </td></tr><tr><td></td><td></td></tr><tr><td colspan="2"> 线粒体分析解决方案（MAS）与复合物II的初始条件 </td></tr><tr><td>化学</td><td>浓度</td></tr><tr><td> 1X MAS </td><td></td></tr><tr><td>琥珀酸</td><td> 10毫</td></tr><tr><td>鱼藤酮</td><td> 2μM </td></tr><tr><td> pH值</td><td> 7.4 </td></tr><tr><td></td><td></td></tr><tr><td colspan="2"> 线粒体分析解决方案（MAS）与复杂的I初始条件 </td></tr><tr><td>化学</td><td>浓度</td></tr><tr><td> 1X MAS </td><td></td></tr><tr><td>丙酮酸</td><td> 5毫米</td></tr><tr><td>马拉特</td><td> 5毫米</td></tr><tr><td> pH值</td><td> 7.4 </td></tr><tr><td colspan="2"> *所有的缓冲区，以在去离子水中进行</td></tr></tbody></table> 表1.解决方案和缓冲区的食谱。 <table border="0" cellpadding="0" cellspacing="0" fo:keep-together.within-page="always"><tbody><tr><td colspan="2"> 协议的步骤 </td><td></td></tr><tr><td colspan="3"> StartProtocol </td></tr><tr><td> 命令 </td><td> 时间（min） </td><td> 港 </td></tr><tr><td>校准</td><td> 0.00 </td><td></td></tr><tr><td>等待</td><td> 10.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>等待</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>等待</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 0.50 </td><td></td></tr><tr><td>措施</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>措施</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 0.50 </td><td></td></tr><tr><td>注入</td><td></td><td>一</td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>措施</td><td> 6.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>注入</td><td></td><td> B </td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>措施</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>注入</td><td></td><td> Ç </td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr<html> &gt; <tr><td>措施</td><td> 3.00 </td><td></td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>注入</td><td></td><td> ð </td></tr><tr><td>混合</td><td> 1.00 </td><td></td></tr><tr><td>措施</td><td> 3.00 </td><td></td></tr><tr><td colspan="3"> EndProtocol </td></tr></tbody></table> 表2.混合，测量，混合周期设置为呼吸。 <img alt="图1" src="/files/ftp_upload/52350/52350fig1highres.jpg" /> 图1.流程图整个协议。 <a href="https://www.jove.com/files/ftp_upload/52350/52350fig1large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> /ftp_upload/52350/52350fig2highres.jpg“/&gt; 图2.整个骨骼肌肉组织以及孤立的线粒体代表印迹。整个组织提取物以及分离的线粒体进行免疫与COX IV抗体作为标记线粒体和GAPDH抗体非线粒体控制。没有GAPDH带观察到分离线粒体表示从非线粒体来源很少或没有污染。 <a href="https://www.jove.com/files/ftp_upload/52350/52350fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图3" src="/files/ftp_upload/52350/52350fig3highres.jpg" /> 线粒体图3.代表透气性轮廓人类股外侧隔离三个孤立的浓度的线粒体，5.0微克，2.5微克和1.0微克我们<html>重新使用在该测定。终浓度端口注射后化合物分别为2毫米ADP（A口）; 2μM寡（B口）; 6μMFCCP（C口）; 2μM抗霉素A（D口）。 RCR计算此运行是7.95。 <a href="https://www.jove.com/files/ftp_upload/52350/52350fig3large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图4" src="/files/ftp_upload/52350/52350fig4highres.jpg" /> 图4平方和。求和使用5.0微克，2.5微克和1.0微克线粒体广场为不同的线粒体呼吸状态和RCR的。 <a href="https://www.jove.com/files/ftp_upload/52350/52350fig4large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> ig5highres.jpg“/&gt; 图5.这可以用来作为一个导引来估计，可以基于初始肌肉样品重量回归分析可以预期的线粒体蛋白的量：肌肉（毫克）和总的线粒体蛋白质产量（毫克）的量。正如预期的那样，有肌肉的量和所得到的总的线粒体蛋白质之间的直接正相关关系。

Watch this Scientific Journal Video about Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy at JoVE.com

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

方法活检股外侧 ，准备纯化线粒体和透气性分析描述。使用小肌肉体积使得该技术适用于临床研究应用。

线粒体的制备及透气性评估，从骨骼肌组织通过穿刺活检获得的分离

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining ...

preparation-respirometric-assessment-mitochondria-isolated-from

Research

JoVE Journal

Medicine

19.2K 次观看.  Wake Forest School of Medicine. 方法活检股外侧 ，准备纯化线粒体和透气性分析描述。使用小肌肉体积使得该技术适用于临床研究应用。 

视频: Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

Myo-mechanical Analysis of Isolated Skeletal Muscle

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force generation and fatigability in treated muscle. We describe a detailed approach to evaluating myo-mechanical properties in freshly explanted hindlimb muscle from the mouse. We describe the atraumatic harvest of mouse extensor digitorum longus muscle, mounting the muscle in a muscle strip myograph (Model 820MS; Danish Myo Technology), and the measurement of maximal twitch and tetanic tension, contraction time, and half-relaxation time, using a square pulse stimulator (Model S48; Grass Technologies). Using these measurements, we demonstrate the calculation of specific twitch and tetanic tension normalized to muscle cross-sectional area, the twitch-to-tetanic tension ratio, the force-frequency relationship curve and the low frequency fatigue curve 4. This analysis provides a method for quantitative comparison between therapeutic interventions in mouse models of muscle disease 1,2,3,5, as well as comparison of the effects of genetic modification on muscle function 6,7,8,9.

To assess the in vivo effects of therapeutic interventions for muscle disease, methods are needed to quantitate force generation and fatigability in treated muscle. We detail an approach to evaluating myo-mechanical properties in explanted mouse hindlimb muscle. This analysis provides a robust approach to quantitating the effects of genetic modification on muscle function, as well as comparison of therapies in mouse models of muscle disease.

离体骨骼肌肌的力学分析

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force ...

科研

医学

Skeletal Muscle Gender Dimorphism from Proteomics

Gross contraction in skeletal muscle is primarily determined by a relatively small number of contractile proteins, however this tissue is also remarkably adaptable to environmental factors1 such as hypertrophy by resistance exercise and atrophy by disuse. It thereby exhibits remodeling and adaptations to stressors (heat, ischemia, heavy metals, etc.)2,3. Damage can occur to muscle by a muscle exerting force while lengthening, the so-called eccentric contraction4. The contractile proteins can be damaged in such exertions and need to be repaired, degraded and/or resynthesized; these functions are not part of the contractile proteins, but of other much less abundant proteins in the cell. To determine what subset of proteins is involved in the amelioration of this type of damage, a global proteome must be established prior to exercise5 and then followed subsequent to the exercise to determine the differential protein expression and thereby highlight candidate proteins in the adaptations to damage and its repair. Furthermore, most studies of skeletal muscle have been conducted on the male of the species and hence may not be representative of female muscle. 
In this article we present a method for extracting proteins reproducibly from male and female muscles, and separating them by two-dimensional gel electrophoresis followed by high resolution digital imaging6. This provides a protocol for spots (and subsequently identified proteins) that show a statistically significant (p &lt; 0.05) two-fold increase or decrease, appear or disappear from the control state. These are then excised, digested with trypsin and separated by high-pressure liquid chromatography coupled to a mass spectrometer (LC/MS) for protein identification (LC/MS/MS)5. This methodology (Figure 1) can be used on many tissues with little to no modification (liver, brain, heart etc.).

A straight-forward set of methods to isolate and determine the identity of the most abundant proteins expressed in skeletal muscle. About 800 spots are discerned on a two-dimensional gel from 10 mg muscle; this allows for the determination of gender-specific protein expression. These methods will give equivalent results in most tissues.

骨骼肌蛋白质组学性别异形

Gross contraction in skeletal muscle is primarily determined by a relatively small number of contractile proteins, however this tissue is also remarkably ...

Assessment of Right Ventricular Structure and Function in Mouse Model of Pulmonary Artery Constriction by Transthoracic Echocardiography

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction4. To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential.
A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software.
Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic&#160;and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC.
Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Right ventricle (RV) dysfunction is critical to the pathogenesis of cardiovascular disease, yet limited methodologies are available for its evaluation. Recent advances in ultrasound imaging provide a noninvasive and accurate option for longitudinal RV study. Herein, we detail a step-by-step echocardiographic method using a murine model of RV pressure overload.

右心室结构和功能的肺动脉缩窄经胸超声小鼠模型的评估

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, ...

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&#246;m Technique

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly effective. This video describes the percutaneous biopsy technique using a modified Bergstr&#246;m needle to obtain skeletal muscle tissue samples from the vastus lateralis of human subjects. The Bergstr&#246;m needle consists of an outer cannula with a small opening (&#8216;window&#8217;) at the side of the tip and an inner trocar with a cutting blade at the distal end. Under local anesthesia and aseptic conditions, the needle is advanced into the skeletal muscle through an incision in the skin, subcutaneous tissue, and fascia. Next, suction is applied to the inner trocar, the outer trocar is pulled back, skeletal muscle tissue is drawn into the window of the outer cannula by the suction, and the inner trocar is rapidly closed, thus cutting or clipping the skeletal muscle tissue sample. The needle is rotated 90&#176;&#160;and another cut is made. This process may be repeated three more times. This multiple cutting technique typically produces a sample of 100-200 mg or more in healthy subjects and can be done immediately before, during, and after a bout of exercise or other intervention. Following post-biopsy dressing of the incision site, subjects typically resume their activities of daily living right away and can fully participate in vigorous physical activity within 48-72 hr. Subjects should avoid heavy resistance exercise for 48 hr to reduce the risk of herniation of the muscle through the incision in the fascia.

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&amp;#246;m Technique

The purpose of this video is to present the modified Bergstr&#246;m skeletal muscle biopsy technique on human subjects.

人骨骼肌活检手术使用修改伯格斯坦技术

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly ...

Substernal Thyroid Biopsy Using Endobronchial Ultrasound-guided Transbronchial Needle Aspiration

Substernal thyroid goiter (STG) represents about 5.8% of all mediastinal lesions1. There is a wide variation in the published incidence rates due to the lack of a standardized definition for STG. Biopsy is often required to differentiate benign from malignant lesions. Unlike cervical thyroid, the overlying sternum precludes ultrasound-guided percutaneous fine needle aspiration of STG. Consequently, surgical mediastinoscopy is performed in the majority of cases, causing significant procedure related morbidity and cost to healthcare. Endobronchial Ultrasound-guided Transbronchial Needle Aspiration (EBUS-TBNA) is a frequently used procedure for diagnosis and staging of non-small cell lung cancer (NSCLC). Minimally invasive needle biopsy for lesions adjacent to the airways can be performed under real-time ultrasound guidance using EBUS. Its safety and efficacy is well established with over 90% sensitivity and specificity. The ability to perform EBUS as an outpatient procedure with same-day discharges offers distinct morbidity and financial advantages over surgery. As physicians performing EBUS gained procedural expertise, they have attempted to diversify its role in the diagnosis of non-lymph node thoracic pathologies. We propose here a role for EBUS-TBNA in the diagnosis of substernal thyroid lesions, along with a step-by-step protocol for the procedure.

Substernal thyroid lesions are common, and need to be differentiated from malignancy. Obtaining percutaneous fine needle biopsy is not possible due to its retrosternal location. This article proposes a protocol for biopsy of substernal thyroid lesions using Endobronchial Ultrasound-guided Transbronchial Needle Aspiration (EBUS-TBNA).

胸骨后甲状腺活检使用支气管内超声引导下经支气管针吸

Substernal thyroid goiter (STG) represents about 5.8% of all mediastinal lesions1. There is a wide variation in the published incidence rates due to the ...

Isolation and Immortalization of Patient-derived Cell Lines from Muscle Biopsy for Disease Modeling

The generation of patient-specific cell lines represents an invaluable tool for diagnostic or translational research, and these cells can be collected from skin or muscle biopsy tissue available during the patient&#8217;s diagnostic workup. In this protocol, we describe a technique for live cell isolation from small amounts of muscle or skin tissue for primary cell culture. Additionally, we provide a technique for the immortalization of myogenic cell lines and fibroblast cell lines from primary cells. Once cell lines are immortalized, substantial expansion of patient-derived cells can be achieved. Immortalized cells are amenable to many downstream applications, including drug screening and in vitro correction of the genetic mutation. Altogether, these protocols provide a reliable tool to generate and preserve patient-derived cells for downstream applications.

This protocol describes techniques for live cell isolation and primary culture of myogenic and fibroblast cell lines from muscle or skin tissue. A technique for the immortalization of these cell lines is also described. Altogether, these protocols provide a reliable tool to generate and preserve patient-derived cells for downstream applications.

从肌肉活检患者来源的细胞系疾病建模隔离和永生化

The generation of patient-specific cell lines represents an invaluable tool for diagnostic or translational research, and these cells can be collected ...

Establishment of Human Epithelial Enteroids and Colonoids from Whole Tissue and Biopsy

The epithelium of the gastrointestinal tract is constantly renewed as it turns over. This process is triggered by the proliferation of intestinal stem cells (ISCs) and progeny that progressively migrate and differentiate toward the tip of the villi. These processes, essential for gastrointestinal homeostasis, have been extensively studied using multiple approaches. Ex vivo technologies, especially primary cell cultures have proven to be promising for understanding intestinal epithelial functions. A long-term primary culture system for mouse intestinal crypts has been established to generate 3-dimensional epithelial organoids. These epithelial structures contain crypt- and villus-like domains reminiscent of normal gut epithelium. Commonly, termed &#8220;enteroids&#8221; when derived from small intestine and &#8220;colonoids&#8221; when derived from colon, they are different from organoids that also contain mesenchyme tissue. Additionally, these enteroids/colonoids continuously produce all cell types found normally within the intestinal epithelium. This in vitro organ-like culture system is rapidly becoming the new gold standard for investigation of intestinal stem cell biology and epithelial cell physiology. This technology has been recently transferred to the study of human gut. The establishment of human derived epithelial enteroids and colonoids from small intestine and colon has been possible through the utilization of specific culture media that allow their growth and maintenance over time. Here, we describe a method to establish a small intestinal and colon crypt-derived system from human whole tissue or biopsies. We emphasize the culture modalities that are essential for the successful growth and maintenance of human enteroids and colonoids.

We describe a method to establish human enteroids from small intestinal crypts and colonoids from colon crypts collected from both surgical tissue and biopsies. In this methodological article, we present the culture modalities that are essential for the successful growth and maintenance of human enteroids and colonoids.

从整体组织和活检建立人类上皮Enteroids和Colonoids的

The epithelium of the gastrointestinal tract is constantly renewed as it turns over. This process is triggered by the proliferation of intestinal stem ...

In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses

Optical coherence tomography (OCT) is the optical equivalent of ultrasound imaging, based on the backscattering of near infrared light. OCT provides real time images with a 15 &#181;m axial resolution at an effective tissue penetration of 2-3 mm. Within the OCT images the loss of signal intensity per millimeter of tissue penetration, the attenuation coefficient, is calculated. The attenuation coefficient is a tissue specific property, providing a quantitative parameter for tissue differentiation. 
Until now, renal mass treatment decisions have been made primarily on the basis of MRI and CT imaging characteristics, age and comorbidity. However these parameters and diagnostic methods lack the finesse to truly detect the malignant potential of a renal mass. A successful core biopsy or fine needle aspiration provides objective tumor differentiation with both sensitivity and specificity in the range of 95-100%. However, a non-diagnostic rate of 10-20% overall, and even up to 30% in SRMs, is to be expected, delaying the diagnostic process due to the frequent necessity for additional biopsy procedures. 
We aim to develop OCT into an optical biopsy, providing real-time imaging combined with on-the-spot tumor differentiation. This publication provides a detailed step-by-step approach for percutaneous, needle based, OCT of renal masses.

In Vivo, Percutaneous, Needle Based, Optical Coherence Tomography of Renal Masses

Optical coherence tomography (OCT) is a high resolution imaging technique that allows analysis of tissue specific optical properties providing the means for tissue differentiation. We developed needle based OCT, providing real-time imaging combined with on-the-spot tumor differentiation. This publication describes a method for percutaneous, needle based OCT of renal masses.

在体内 ，经皮，基于针，肾群众的光学相干断层扫描

Optical coherence tomography (OCT) is the optical equivalent of ultrasound imaging, based on the backscattering of near infrared light. OCT provides real ...

Povidone Iodine Rectal Preparation at Time of Prostate Needle Biopsy is a Simple and Reproducible Means to Reduce Risk of Procedural Infection

Single institution and population-based studies highlight that infectious complications following transrectal ultrasound guided prostate needle biopsy (TRUS PNB) are increasing. Such infections are largely attributable to quinolone resistant microorganisms which colonize the rectal vault and are translocated into the bloodstream during the biopsy procedure. A povidone iodine rectal preparation (PIRP) at time of biopsy is a simple, reproducible method to reduce rectal microorganism colony counts and therefore resultant infections following TRUS PNB.
All patients are administered three days of oral antibiotic therapy prior to biopsy. The PIRP technique involves initially positioning the patient in the standard manner for a TRUS PNB. Following digital rectal examination, 15 ml&#160;of a 10% solution of commercially available povidone iodine is mixed with 5 ml&#160;of 1% lidocaine jelly to create slurry. A 4 cm x 4 cm sterile gauze is soaked in this slurry and then inserted into the rectal vault for 2 min after which it is removed. Thereafter, a disposable cotton gynecologic swab is used to paint both the perianal area and the rectal vault to a distance of 3 cm from the anus. The povidone iodine solution is then allowed to dry for 2 - 3 min prior to proceeding with standard transrectal ultrasonography and subsequent biopsy.
This PIRP technique has been in practice at our institution since March of 2012 with an associated reduction of post-biopsy infections from 4.3% to 0.6% (p = 0.02). The principal advantage of this prophylaxis regimen is its simplicity and reproducibility with use of an easily available, inexpensive agent to reduce infections. Furthermore, the technique avoids exposing patients to additional systemic antibiotics with potential further propagation of multi-drug resistant organisms. Usage of PIRP at TRUS PNB, however, is not applicable for patients with iodine or shellfish allergies.

Here, we present a protocol for use at the time of transrectal ultrasound guided prostate needle biopsy (TRUS PNB) that is a simple and cost-effective means to reduce infections following the procedure.

聚维酮碘直肠准备在前列腺穿刺活检的时间是一个简单的和可重复的手段，以减少程序感染的风险

Single institution and population-based studies highlight that infectious complications following transrectal ultrasound guided prostate needle biopsy ...

Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome

Percutaneous muscle biopsy using the Weil-Blakesley conchotome is well established in both clinical and research practice. It is a safe, effective and well tolerated technique. The Weil-Blakesley conchotome has a sharp biting tip with a 4 - 6 mm wide hollow. It is inserted through a 5 - 10 mm skin incision and can be maneuvered for controlled tissue penetration. The tip is opened and closed within the tissue and then rotated through 90 -180&#176; to cut the muscle. The amount of muscle obtained following repeated sampling can vary from 20 mg to 290 mg which can be processed for both histology and molecular studies. The wound needs to be kept dry and vigorous physical activity kept to a minimum for approximately 72 hr although normal levels of activity can restart immediately following the procedure. This procedure is safe and effective when close attention is paid to the selection of subjects, full asepsis and post procedure care. &#160;Both right and left vastus lateralis are suitable for biopsy dependent on participant preference.

Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome

This video demonstrates the technique of percutaneous muscle biopsy of the human vastus lateralis using the Weil-Blakesley conchotome.

人的<em&gt;股外侧肌</em&gt;骨骼肌活检使用韦尔 - Blakesley Conchotome

Percutaneous muscle biopsy using the Weil-Blakesley conchotome is well established in both clinical and research practice. It is a safe, effective and ...