感谢Therwa哈姆扎，约翰·E. TIDWELL，尼娜克洛维斯，和苏珊·史密斯的实验援助和苏珊·史密斯校对。作者还感谢提供临床分离的细菌和约翰·B·巴尼特博士对他的支持和使用的生物安全实验室在西弗吉尼亚大学微生物学，免疫学和细胞生物学系的约翰·托马斯博士。作者承认金融支持接骨和创伤护理基金会和美国国家科学基金会（＃1003907）。显微镜实验和图像分析技术在西弗吉尼亚大学的成像设备，这是支持部分的玛丽巴布伦道夫癌症中心和美国国立卫生研究院授予P20 RR016440。 动物使用的血液提请批准了西弗吉尼亚大学实验动物护理和使用委员会。所有的实验被处决符合所有相关guidelin的ES，法规和监管机构。

1。 PRP的制备和激活 1.1抽血<ol><li>麻醉家兔吸入异氟醚（2％O 2诱导和维护的1％）。 </li><li>绘制2毫升0.129中号柠檬酸三钠（一种抗凝血剂的溶液）放入20毫升注射器。的柠檬酸三钠溶液的制备通过1.897克柠檬酸三钠溶解在50毫升蒸馏H 2 O，并用0.22μm的无菌过滤器过滤。 </li><li>用70％乙醇消毒的兔耳。 </li><li>绘制血液（ 例如 5毫升）通过连接到注射器的蝴蝶针（25 G）从兔耳缘静脉。 </li><li>混血的柠檬酸三钠溶液中轻轻搅拌。血液和柠檬酸三钠溶液的体积比为9:1。 </li></ol> 1.2 PRP制备（ 图1） <ol><li>抗凝固处理的血液转移至50ml塑料离心管中。以10微升的等分试样血以确定基线血小板计数hemocytometry。 </li><li>离心血在300×g离心10分钟，在室温（RT）在离心分离机的摆​​动离开转子。设置加速和制动速度为低（ 图1）。 </li><li>离心分离后，血液被分离成三层。底层是主要的红血细胞，中间层（通常简称为“血沉棕黄层”）组成的浓缩血小板和白细胞，顶端层主要是等离子体，这是血液中的液体成分，和血小板（ 图1）。离心管小心地运送到细胞培养罩;不打扰层。塑料移液器使用1毫升到15毫升塑料管，将所有的血浆，棕黄色的大衣，和2-3毫米厚的红血细胞层。 </li><li>离心传输的示例的第二时间，以3,000 xg离心15分钟，在RT。顶层（上清液），贫血小板血浆（PPP）被视为d被转移到新的管中。 </li><li>获得PRP通过使用PPP，得到2.0×10 6血小板/μl（确定hemocytometry），在其它的血液试样中的血小板浓度调节。 </li></ol> 1.3 PRP激活<ol><li>准备PRP激活溶液用5毫升10％的氯化钙溶解5000 IU牛凝血酶的工作浓度1,000 IU / ml的。 </li><li>添加活化液PRP和PPP，并反复吹打形成PRP和PPP凝胶混合溶液。 PRP或PPP的激活溶液的体积比为1:4。 </li></ol> 2使用kill曲线法的PRP 的体外抗菌试验（ 图2） <ol><li>用无菌接种环，增加数个殖民地的S.黄色葡萄球菌从它的平板培养过夜到5毫升的Mueller Hinton肉汤（MHB）的塑料管中。涡简要然后在37℃下孵育2小时样品下，细菌的介质的光密度，使用分光光度计测定，并调整至光密度等于〜1×10 8 CFU / ml为根据预先确定的标准曲线。 </li><li> 100倍稀释液，使用PBS，得到1×10 6 CFU /毫升的接种物并放置在冰上。 </li><li>设置和无菌，一次性5 ml圆底聚苯乙烯管贴上标签，准备以下的样本组如表1中所示，在每个试管的最终体积为2ml。 </li><li>加入PRP和PPP，或PBS第一聚苯乙烯管中，然后由用于激活的凝血酶溶液（凝胶的形成）。接下来，添加MHB，然后S.黄色葡萄球菌接种物（1×10 6 CFU /毫升），得到的最终浓度为1×10 5 CFU /毫升。 </li><li>孵育在37℃下在150 rpm的搅拌与轨道管。 </li><li>在预先确定的时间点（ 例如，0，1，和2小时），每个试管中混合的解决方案通过重复吸移的（这步骤是非常重要的，因为细菌可能会被困在里面的PRP凝胶）。取10微升的样品，用无菌的0.9％的生理盐水稀释串联，和移液100微升等分的各稀释到胰蛋白酶大豆琼脂（TSA，用5％羊血）CFU计数板。 </li><li>文化的琼脂平板上，在37℃下过夜，然后计算和记录板菌落。标绘数据上logarithimic规模上的x轴和CFU /毫升在y-轴的时间（hr）。 </li></ol>

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作者宣称，他们没有竞争的金融利益。

富血小板血浆已被越来越多地用于临床的应用，由于其促进愈合的特性15-17。在本研究中中，PRP提出了一种新的方法预防感染。 PRP发现对MRSA，MSSA，A组链球菌 ， 淋病奈瑟菌有较强的抗菌性能。 PRP的主要优势在于，相对于传统的抗生素治疗，预防感染包括：（1）目前的抗生素治疗所面临的挑战，包括日 ​​益抗生素的耐药性14,18-20。 PRP可能是一个先进的替代品，因?...

种植体相关感染是一个显着的临床并发症。 金黄色葡萄球菌 （ 金黄色葡萄球菌 ）是孤立从植入物相关感染的最常见的微生物之一，。它是能够生产覆盖的植入物的表面，并可能会导致抗生素耐药性的感染1,2的生物膜。植入相关感染的治疗往往需要长期住院，反复清创术和静脉抗生素治疗时间延长。在抗生素有抗药性的情况下，去除植入物可能是必要的。上升抗生素对细菌的抵抗力也被称为中心疾病控制和预防中心（CDC）为“世界上最紧迫的健康问题之一。”随着时间的推移，如果没有新的有效的抗菌治疗的发展，它是多药耐药病原体与传统的抗生素是无法治愈的。植入相关的预防因此，感染是重要的和新的预防剂或方法是必要的，以防止这类感染。 富血小板血浆（PRP）是一个包含超过30的生长因子，可以帮助骨和骨移植愈合3-5的自体血浓度。已越来越多地在诊所报告，因为它的高浓度的各种生长因子，由血小板释放的PRP中的应用，以提高骨再生和软组织的成熟。 PRP几个特点表明，PRP可能还具有抗菌特性6-9。 PRP中包含了大量的血小板，白细胞高浓度的（其可以具有对细菌和真菌的宿主防御动作），和多个抗菌肽7,8,10。在最近的一大群心脏手术患者的研究中，据透露，术中使用PRP凝胶伤口愈合的显icantly的发病率下降的浅表和深部胸骨感染11。由于这些原因和观察，我们推测 ，PRP，除了其研究的愈合特性，具有抗菌性能。使用PRP，以防止感染的潜在优势，可包括：（ⅰ）PRP是不太可能诱导电阻相比，传统的抗生素治疗。 （ 二 ）PRP也有属性，促进愈合其中可能有一个协同效应对预防感染; PRP的愈合，促进属性可以提供密封，以防止细菌附着从而降低的几率为感染作为病原体和人类的细胞都竞相植入物表面12 ，13。 （ 三 ）PRP是固有的生物相容性，安全，无传染性疾病的风险。 我们的长期目标是使用PRP作为一种新的方法，以防止植入物相关infecti组件。本研究的目的是准备PRP使用两次离心的方法，检查PRP的体外抗菌性能，并描述协议，以评估这种抗菌性能。

<table border="1">
 <tbody>
 <tr>
 <td>Name of Reagent/Material</td>
 <td>Company</td>
 <td>Catalog Number</td>
 <td>Comments</td>
 </tr>
 <tr>
 <td>Bovine thrombin</td>
 <td>King Pharmaceuticals, Inc </td>
 <td>60793-215-05</td>
 <td>Thrombin (bovine origin)</td>
 </tr>
 <tr>
 <td>Calcium chloride</td>
 <td>King Pharmaceuticals, Inc</td>
 <td>60793-215-05</td>
 <td>10% calcium chloride</td>
 </tr>
 <tr>
 <td>Ethanol</td>
 <td>Sigma-Aldrich</td>
 <td>E7023</td>
 <td></td>
 </tr>
 <tr>
 <td>Isoflurane</td>
 <td>Baxter</td>
 <td>1001936060</td>
 <td></td>
 </tr>
 <tr>
 <td>Mueller Hinton broth</td>
 <td>Becton, Dickinson and Company</td>
 <td>275710</td>
 <td></td>
 </tr>
 <tr>
 <td>Phosphate-buffered saline</td>
 <td>Sigma-Aldrich</td>
 <td>D8662</td>
 <td></td>
 </tr>
 <tr>
 <td>Tri-sodium citrate</td>
 <td>Sigma-Aldrich</td>
 <td>W302600</td>
 <td></td>
 </tr>
 <tr>
 <td>Tryptic soy agar</td>
 <td>Fisher Scientific</td>
 <td>R01202</td>
 <td></td>
 </tr>
 <tr>
 <td>Centrifuge</td>
 <td>Kendro Laboratory Products</td>
 <td>750043077</td>
 <td></td>
 </tr>
 <tr>
 <td>Syringe filter</td>
 <td>Millipore</td>
 <td>SLGP033RS</td>
 <td></td>
 </tr>
 </tbody>
</table>

prp as a new approach to prevent infection: preparation and in vitro antimicrobial properties of prp

植入相关感染已成为全球范围内的医疗保健行业越来越具有挑战性由于越来越大的抗药性，抗生素有抗药性的细菌，动物和人类之间传播的，成本高，治疗感染。 在这项研究中，我们公开了一种新的策略，在防止植入物相关感染的基础上的潜在的富血小板血浆（PRP）的抗菌性能，可能是有效的。由于其促进愈合的研究，PRP（生物制品）已经被越来越多地用于临床应用，包括骨科手术，牙周和口腔外科，口腔颌面外科手术，整形外科，运动医学等。 PRP可能是一个先进的替代传统的抗生素治疗，防止植入相关感染。比常规抗生素治疗州市的PRP的使用可能是有利的NCE PRP是不太可能诱导抗生素耐药性和PRP的抗菌和愈合促进性能可能具有协同作用对预防感染。这是众所周知的，病原体和人类细胞竞相种植体表面，和PRP促进愈合的属性可以改善人类的细胞附着，从而减少了感染的几率。此外，PRP是固有的生物相容性，安全，无传染性疾病的风险。 我们的研究中，我们选择了一些临床中常见的骨科感染的细菌菌株和PRP检查是否有对这些细菌的体外抗菌性能。我们已经准备好PRP使用两次离心的做法，让所有的样品中获得相同的血小板浓度。我们已经取得了抗菌剂的研究结果一致，发现PRP具有较强的体外抗菌性能，对细菌，如methi的Cillin的敏感和耐甲氧西林金黄色葡萄球菌，A组链球菌 ， 淋球菌 。因此，使用的PRP可能有潜力以防止感染和昂贵的手术后的植入物相关的感染的治疗，以减少需要。

PRP的可重复使用两次离心的方法（ 图1）的制备。 PRP的发现呈现强（高达100倍减少的CFUs） 在体外的抗菌性能，对耐甲氧西林S.黄色葡萄球菌 （MRSA）（ 图3），这是常见的在世界各地的医院14。同样，，PRP具有较强的抗菌性能，对甲氧西林敏感的S.金黄色葡萄球菌 （MSSA），A组链球菌 ， 淋球菌 。 <p class="jo...

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PRP as a New Approach to Prevent Infection: Preparation and In vitro Antimicrobial Properties of PRP

植入相关感染是一个重要的临床并发症。这项研究描述了一种方法，使用富血小板血浆（PRP），以防止植入物相关感染，给出了协议，准备PRP血小板浓度不变，并报告新发现的抗菌性能的PRP和检查等抗菌性能的相关协议<em&gt;在体外。</em

PRP作为一种新的方法，以防止感染的制备及<em在体外</em&gt;抗菌性能的PRP

Implant-associated infection is becoming more and more challenging to the healthcare industry worldwide due to increasing antibiotic resistance, ...

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PRP as a New Approach to Prevent Infection: Preparation and In vitro Antimicrobial Properties of PRP

21.2K 次观看.  West Virginia University. 植入相关感染是一个重要的临床并发症。这项研究描述了一种方法，使用富血小板血浆（PRP），以防止植入物相关感染，给出了协议，准备PRP血小板浓度不变，并报告新发现的抗菌性能的PRP和检查等抗菌性能的相关协议在体外。