本刊物的资金提供Cerus公司，INTERCEPT血液系统的制造商。

1。全血收集<ol><li>根据当地采血准则的收集全血无偿献血者在450毫升的顶部/底部收集组。 </li><li>的全血被存储为至少2小时，离心分离，分离前的冷却板。 </li></ol> 2。巴菲大衣准备<ol><li>离心的全血，使用硬的自旋分为三层:红血细胞，血沉棕黄层，和血浆分离血液。离心机使用的参数为4,880 RCF为11分钟，在22±2°C。 </li><li>使用自动化的血液分离表达到顶端的的卫星袋和进入底部的卫星袋的红血细胞（RBC）的血浆的收集容器中，留下的血沉棕黄层。目标平均体积和红细胞压积范围为棕黄色大衣约48毫升和37％，分别为。 </li><li>棕黄色的大衣，存放过夜血小板搅拌器在22±2°C。 </li></ol><ol><li>无菌连接7棕黄色大衣和300毫升的SSP +血小板添加剂溶液（PAS）与平行线的列车编组列车的长度，以减少PAS应在顶部的火车。夹线之间的PAS和棕黄色大衣。 </li><li>打开无菌的棕黄色大衣单位之间的连接，并允许排入的棕黄色大衣的最后一个容器。 </li><li>打开焊缝和夹具之间的PAS和棕黄色大衣。允许三分之一的添加剂溶液的冲洗顺序通过每个的血沉棕黄层容器。重复2次以上，每次使用一个一半的剩余PAS。 </li><li>平均棕黄色大衣池体积应为约600毫升。如果目标，满足个人的棕黄色大衣的体积和红细胞压积，血浆比是32 - 47％用于拦截病原体灭活后治理的过程中。 </li><li>丢弃的空SSP +和Buffy外套的容器。 </li><li>为了获得最佳的血小板回收，保持池在搅拌1小时，离心分离前。 </li><li>无菌连接血小板的存储容器，综合性白血球过滤器的棕黄色大衣池。 </li><li>执行一个"软自旋"血沉棕黄层池分开的红血细胞中的血小板悬浮液（462 RCF 9分钟，20秒）。通过去白细胞过滤器的自动血液分离的​​血小板储存容器中表达的血小板悬液。 </li><li>先前描述的处理要求确保的血小板悬浮液符合INTERCEPT处理规格为300毫升- 420毫升体积，2.5-7.0×10 11个血小板的剂量，和≤4×10 6个 / ml的红细胞。 </li></ol> 4。截取处理<ol><li>在年底前集合后第1天（第0天是集合的日子）进行拦截处理。 </li><li>拆开INTERCEPT设置的处理从透明的塑料包装袋的双存储容器。 </li><li>无菌连接管的amotosalen容器的拦截处理组的血小板悬液容器。 </li><li>按照当地的要求与适当的血液产品标识，标记拦截处理组的存储容器。 </li><li>杭血小板和打破第一上amotosalen容器的底部套管，照明容器允许的amotosalen的溶液流入。打破上amotosalen允许血小板流动到照明容器通过amotosalen容器的容器顶端插管。 </li><li>轻轻地混合的血小板和amotosalen混合物，并表示将空气从照明容器到amotosalen容器。 </li><li>表达少量血小板混合物进入管件，填充约4厘米的油管。这可确保在油管和照明容器中的血小板进行病原体inactivati在接受治疗。 </li><li>密封在照明容器和amotosalen容器之间的管子。留下不超过约4厘米的管道，从照明容器延伸。拆下并丢弃空血小板和amotosalen的容器和关闭采样袋的夹子。 </li><li>左边大隔室容器中，并与照明的组织者中的小室的右侧，将到照射器的处理集合。 </li><li>使用手持式条形码的移动设备进入到照射器的捐赠ID，产品代码，和处理集合特定号码。关闭的金属盖和照明的图形界面上提示时，关上抽屉。按"开始"开始的照明。 </li><li>照明灯照明后，删除处理集合。照明器会自动打印处理后的血小板单元（S）的治疗报告。 </li><li>拆开容器的组织者和挂在Platelets和处理集合，打破插管的照明容器的出口处，并允许血小板流入的化合物的吸附装置（CAD）的容器。 </li><li>小心不要弯曲CAD晶片，表达的照明容器使用血浆中提取的CAD容器中的空气。 </li><li>密封的管子的CAD容器的进气口附近。删除和丢弃的空照明容器。 </li><li>将CAD连接的存储容器，容器上的血小板的搅拌器为至少6小时，但不超过16小时。这将导致在≤2μM的浓度残余amotosalen减少。 </li><li> CAD处理后，去除血小板单位的搅拌器。杭血小板。额的套管和允许血小板流入的存储容器。 </li><li>表达到CAD中容器中的空气从所述存储容器。让残留的血小板浓缩液流回进入储存容器的重力。以上的Y型接头和密封管，取出空的CAD容器。 </li><li>重新分配的存储容器中，作为需要之间的容积。量的调整是由重的存储容器。密封管子的存储容器的入口上方几厘米，每个储存容器，这有利于从最终产品中，如5.2中所述获得的无菌样品。 </li></ol> 5。产品抽样<ol><li>对于常规QC测试，最终的储存容器可以被采样一次，通过使用采样袋上的存储容器。要做到这一点，确保血小板的单位是充分混合，然后打开夹子，去眼袋，挤几次。密封管后袋盛满了血小板。血小板样品转移到一个适当的实验室管，并立即进行分析。 </li><li>为了获得样品在不同的时间点，例如在存储过程中的验证研究，无菌连接一个新的取样容器的血小板的收纳容器的管。确保充分混合血小板前转移到取样容器。 </li></ol> 6， 在体外功能评价<ol><li>对于此验证研究， 在体外评价进行了CAD治疗（1天或2天，根据CAD时间）后，再在第4天（或第5天）和第7天。 在体外测量体积，血小板计数，pH值，血气（PO 2，PCO 2），葡萄糖，乳酸和旋转。 </li><li>卷使用添加剂溶液中血小板的比重为1.01克/毫升作为重量确定。 </li><li>对血红蛋白污染视觉评估使用彩色图表比较。 </li><li>旋流目测。 </li><li>请参见"表"其他实验方法的设备。 </li></ol> 7。代表性的成果生产D的过程中开始强强连手剂量棕黄色大衣血小板与个人满足目标规格的棕黄色大衣的体积和红细胞压积生产。因为它是不实际的测量血细胞比容的个人棕黄色大衣在最后的工艺验证，我们开始进行单独的努力，以优化我们的棕黄色大衣，以确保我们能够不断满足目标量和红细胞压积。如表1中所示，我们的优化的血沉棕黄外套相比，毫不逊色，在46±2毫升和37±3％，分别为两个音量和血细胞比容值的目标值。 池，后的体积和血细胞比容的血沉棕黄层池应该是约600毫升和20％，分别为前软自旋离心。如表2中所示，我们的血沉棕黄层池平均615±5毫升，平均血细胞比容19±1％。 在双倍剂量的血小板浓度的软旋转离心结果entrate，将满足输入的要求病原体灭活利用INTERCEPT血液系统DS处理组。 INTERCEPT治疗的主要输入参数包括体积，血小板计数，血浆比，RBC内容。此外，我们的目标是回收相比，≥75％的血小板浓缩物中的血小板的血沉棕黄层池。根据当地的要求，白细胞（WBC）的污染必须&lt;1×10 6 /单位。浓缩血小板在我们的验证，达到了关键参数INTERCEPT治疗以及白细胞的污染和血小板的恢复，如表3所示的目标。 INTERCEPT病原体的失活过程进行双倍剂量的血小板浓缩液使用一个单一的INTERCEPT处理组。处理集合包含两个整体的存储容器允许处理单元完成的路径被分裂成两个单独的治疗性血小板剂量ogen失活过程。欧洲指引规定，75％的单元测试包含≥200×9血小板每治疗dose1的;在瑞典当地的要求，需要有75％的单元测试包含&gt; 240x10 9血小板每剂量。截取后的治疗，平均血小板含量为265±22×10 9组（n = 24）。此外，88％的单位达到或超过240x10 9血小板每治疗量，这是在欧洲的准则和瑞典法规。双倍剂量的棕黄色大衣准备和INTERCEPT处理过程的说明，请参阅图1和图2。 此验证中，我们测量后，的INTERCEPT治疗后分割成单个存储容器（ 即 ）浓缩血小板体外特性的参数进行测量，存储超过7天 ​​。均值和标准差收集血小板剂量，pH值，PO 2，PCO 2，乳酸的生产和葡萄糖消耗。 图3显示了平均起始血小板剂量的双倍剂量的血小板浓缩液INTERCEPT超过7天 ​​的存储处理后，在每个分割产品INTERCEPT治疗前和血小板平均剂量。血小板在储存过程中损失了约9％。这种减少是在储存过程中的常规的血小板。2不同于预期的血小板损失 表4总结了在体外的INTERCEPT血小板治疗后和分裂成单个单元的特点。 每欧洲的要求，血小板的pH值必须保持高于6.4，通过端部的保质期。在处理过程中，血小板浓缩物的pH值略有下降的基础上的血小板浓度，体积，和气体渗透性的血小板STO愤怒容器。 图4示出分割血小板产品的pH值超过7天 ​​的存储。在存储过程中，pH值是稳定和内保持良好的加工要求。 如示于图5A和5B，血小板的O 2的消耗和CO 2生产表明持续呼吸由INTERCEPT血小板的PL2410容器中储存7天的期间， 图6A和6B示出的乳酸和葡萄糖水平超过7天 ​​的存储。血小板葡萄糖消耗和乳酸生产的是彼此一致的，在7天的存储。 5个单位的血糖水平小于1.11毫摩尔/升（用于葡萄糖测定的下限）。 棕黄色涂层的制备及浓缩池验证血小板，符合输入条件，INTERCEPT治疗，具体而言，体积，血小板计数，血浆比，红色blOOD细胞污染。最后的单位符合验证标准的存储空间，包括血小板剂量和pH值超过7天。 <table border="1"><tbody><tr><td> 参数 </td><td> 目标范围 </td><td> 结果平均值±标准差 </td></tr><tr><td>体积（ml） </td><td> 〜48 </td><td> 46±2 </td></tr><tr><td>血细胞比容（％） </td><td> 〜37 </td><td> 37±3 </td></tr><tr><td>保持时间前池</td><td>夜宿于搅拌器</td><td>夜宿于搅拌器</td></tr></tbody></table> 表1。个人捉鬼大衣（N = 19）。 <table border="1"><tbody><tr><td> 参数 </td><td> 目标范围 </td><td> 结果平均值±标准差 </td></tr><tr><td>体积（ml） </td><td> 〜600 </td><td> 615±5 </td></tr><tr><td> Hematocrit（％） </td><td> 〜20 </td><td> 19±1 </td></tr><tr><td>保持时间在离心前</td><td> 1小时搅拌器</td><td> 1小时搅拌器</td></tr></tbody></table> 表2。捉鬼大衣池"（N = 12）。 <table border="1"><tbody><tr><td> 参数 </td><td> 目标 </td><td> 结果平均值±标准差 </td></tr><tr><td>体积（ml） </td><td> 370 - 420 </td><td> 404±8 </td></tr><tr><td>血小板计数（×10 9 /单位） </td><td> 250 - 700 </td><td> 577±62 </td></tr><tr><td>血小板平均回收率（％） </td><td> ≥75 </td><td> 75±4 </td></tr><tr><td>等离子比率（％） </td><td> 32 - 47 </td><td> 38±1 </td></tr><tr><td> RBC（×10 6个 /毫升） </td><td> ≤4 </td><td> 1.2±0.4 </td></tr><tr><td> WBC（×10 6 </s向上&gt; /单位） </td><td> ≤1 </td><td> 0.11±0.1 </td></tr><tr><td>保持时间前INTERCEPT（小时） </td><td> ≤1日结束</td><td> ≤1日结束</td></tr></tbody></table> 表3。双倍剂量的血小板浓缩物（N = 12）。 <table border="1"><tbody><tr><td></td><td> 平均值±标准差 </td><td> 最小 </td><td> 最大 </td></tr><tr><td>体积（ml） </td><td> 199±16 </td><td> 182 </td><td> 236 </td></tr><tr><td>血小板计数（×10 9 /单位） </td><td> 265±22 </td><td> 225 </td><td> 292 </td></tr><tr><td> pH值（22°C） </td><td> 6.9±0.1 </td><td> 6.8 </td><td> 7 </td></tr><tr><td>宝2（千帕） </td><td> 21.4±4.8 </td><td> 12.8 </td><td> 27.3 </td></tr><tr><td> 二氧化碳分压（千帕） </td><td> 4.3及PLusmn 0.4 </td><td> 3.3 </td><td> 4.9 </td></tr><tr><td>乳酸（毫摩尔/升） </td><td> 9.1±1.7 </td><td> 6.8 </td><td> 12.4 </td></tr><tr><td>血糖值（mmol / l）的</td><td> 5.3±0.9 </td><td> 3.7 </td><td> 6.5 </td></tr></tbody></table> 表4。INTERCEPT治疗血小板日1/2（单单元，n = 24）。 <img alt="图1" src="/files/ftp_upload/4414/4414fig1.jpg" /> 图1。生产的双倍剂量的浓缩血小板从池中7棕黄色大衣。 <img alt="图2" src="/files/ftp_upload/4414/4414fig2.jpg" /> 图2。INTERCEPT血液系统双倍剂量的浓缩血小板与病原体灭活。 <img alt="图3" src="/files/ftp_upload/4414/4414fig3.jpg" /> 图3。前平均血小板剂量和INTERCEPT治疗后第7天（截取前N = 12，N = 24后INTERCEPT）。 <img alt="图4" src="/files/ftp_upload/4414/4414fig4.jpg" /> 图4。平均血小板的pH值±SD后的INTERCEPT治疗组（n = 24）。 <img alt="图5A" src="/files/ftp_upload/4414/4414fig5a.jpg" /> 图5A，平均PO 2±SD后的INTERCEPT治疗组（n = 24）。 <img alt="图5B" src="/files/ftp_upload/4414/4414fig5b.jpg" /> 图5B后的INTERCEPT治疗组（n = 24），平均PO 2±SD。 <img alt="图6A" src="/files/ftp_upload/4414/4414fig6a.jpg" /> 图6A，平均乳酸水平±SD后的INTERCEPT治疗组（n = 24）。 <img alt="图6B" src="/files/ftp_upload/4414/4414fig6b.jpg" /> 图6B。平均血糖水平及plusmn; SD截取后的治疗组（n = 24）。

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C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protection+Against+TA-GVHD+Due+to+Platelet+Transfusion+By+Using+Pathogen+Inactivation+with+the+INTERCEPT+Blood+SystemTM+-+Gamma+Irradiation+is+Not+the+Only+Answer.">Protection Against TA-GVHD Due to Platelet Transfusion By Using Pathogen Inactivation with the INTERCEPT Blood SystemTM - Gamma Irradiation is Not the Only Answer.</a> Haematologica. 95 (Extra 1), 230-237 (2010).</li><li>Corash, L., Lin, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Novel+processes+for+inactivation+of+leukocytes+to+prevent+transfusion-associated+graft-verus-host+disease.">Novel processes for inactivation of leukocytes to prevent transfusion-associated graft-verus-host disease.</a> Bone Marrow Transplant. 33, 1-7 (2004).</li></ol>

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捉鬼外套血小板需要几个处理步骤，在收集后的费用，包括人员的时间，耗材，设备和废物，必须考虑到生产的血小板单位的整体成本。提高血小板产量从每一个棕黄色大衣（通过优化的棕黄色大衣体积和红细胞压积）允许生产的棕黄色大衣双倍剂量血小板单位从池中7棕黄色大衣。当被执行时，这个优化血沉棕黄外套的数目需要以产生一个固定数量的血小板剂量可能会减少，从而提高了整体使用率的血沉棕黄外套和使额外的血小板浓缩物（PC）的生产。人员的时间以及耗材和设备的费用，如池组，血小板添加剂溶液，无菌连接晶片，过滤器设置，和离心程序，减少高达50％。除了优化血小板生产和降低相关的成本，氏小号池的方法可以生成一个浓缩血小板（PC）使用的的INTERCEPT双存储集装箱处理组，以满足输入的要求，使我们能够提供更好的保护我们的受血者。 INTERCEPT血液系统的病原体灭活使用amotosalen和紫外线A（UVA）光共价交联DNA和RNA，防止核酸的复制和渲染无法引起疾病的病原体。它可以有效地灭活的病原体，包括病毒，细菌的广谱，寄生虫和污染捐助的白血细胞。4-6的 INTERCEPT提供了一个替代目前的测试范例新出现的病原体，在历史上涉及重大的延迟，而新的测试，并最终实现需要一个重要的金融投资，当测试。7,8它也提供了另一种冗余安全measu水库，如细 ​​菌检测和γ射线。10-12此外，INTERCEPT让我们双倍剂量血小板治疗有利于我们的生产效率，并有助于我们坚持我们的预算。 INTERCEPT和双倍剂量的血沉棕黄层方法可适于各种血液中心的工作流程。作为实施例中，全血集合可以增加至500毫升;集合也可以被存储在22过夜°C分离前。此外，一个备用的血沉棕黄层轮询方法是可能的（ 例如，一个章鱼的方法，而不是一列火车方法）和/或一个自动化装置（ 例如 TACSI）可被用于从其它红细胞第二离心分离和分离的血小板悬浮液。 是必要的，如果增加血小板含量的几种技术是可用的，包括预选捐助者基于对血小板计数的全血，培养过夜，adjus黄贤金，离心设置，或利用池代替7 8棕黄色大衣。 当前可用的容量离心机水桶由于汇集的限制设置，血沉棕黄层池的总体积应是约600毫升。在本协议中所述的离心过程和设置进行了优化，收益率的产品，满足池7棕黄色大衣的的INTERCEPT参数为。离心必须设置被修改，如果修改池中的棕黄色大衣的数量。 在一些国家，血小板最低剂量更高，比瑞典更严格的质量控制要求。因此，我们的结果可能是普遍适用的。在这些国家中，双剂量PC拦截治疗前需要包含大量的血小板，截取后的处理和分割，以满足本地需求。由于最大的血小板含量和治疗体积FOŕINTERCEPT是7×10 11的血小板和420毫升分别，百分比单位超过INTERCEPT处理要求或有被分裂成两个治疗剂量的血小板含量不足，会有所不同，这取决于因素如本地要求血小板剂量，控制准则，血沉棕黄层的优化结果，和生产稳定性。 如果双倍剂量的PC超过INTERCEPT处理的要求，可以手动调整，以满足后续处理。在几次7棕黄色大衣池产生血小板不足截取后的双倍剂量的产品，我们选择不进行拦截处理。另外，可以选择其他血液中心集（ 即大的卷集）单剂量INTERCEPT处理和存储处理的PC，PC作为一个单一的治疗剂量较大，从而对各单位进行病原体灭活。为了确保我们能满足QC要求和病原体灭活的血小板单位可能，我们选择棕黄色大衣的基础上捐助血小板计数，当汇集起来，将导致在5.6x10 11在游泳池血小板的的血小板含量最低的。这将确保足够的INTERCEPT治疗后产生两种治疗剂量的血小板含量。 我们的验证表明，全血衍生棕黄色大衣单位可以成功地汇集和处理INTERCEPT血小板的过程，导致病原体灭活血小板产品，以满足制造（瑞典要求和欧洲的准则）的验收标准和支持的患者需要输注血小板的临床实践准则和标准在瑞典的血小板输注方法。

<table border="1">
	<tbody>
		<tr>
			<td>Material / Equipment</td>
			<td>Company</td>
			<td>Catalogue Number</td>
			<td>Comments</td>
		</tr>
		<tr>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>Whole blood donation, primary separation, and platelet production</td>
		</tr>
		<tr>
			<td>Blood collection pack</td>
			<td>Fenwal</td>
			<td>R6485</td>
			<td>Top/Bottom set</td>
		</tr>
		<tr>
			<td>Automated component extractor</td>
			<td>Fenwal</td>
			<td>Optipress-II</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Blood mixer and balance system</td>
			<td>Baxter</td>
			<td>Easymix V3</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Platelet leukocyte filtration set</td>
			<td>Fenwal</td>
			<td>K4R7042</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Hettich</td>
			<td>Roto Silenta 63 RS</td>
			<td>Version 5.5</td>
		</tr>
		<tr>
			<td>Platelet additive solution - SSP+</td>
			<td>MacoPharma</td>
			<td>SSP2030U</td>
			<td>300 ml</td>
		</tr>
		<tr>
			<td>Sterile tubing welder</td>
			<td>Terumo</td>
			<td>T-SCD</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>INTERCEPT treatment &amp; storage</td>
		</tr>
		<tr>
			<td>INTERCEPT processing set</td>
			<td>Cerus</td>
			<td>INT2503</td>
			<td>Dual Storage (DS) set</td>
		</tr>
		<tr>
			<td>INTERCEPT Illuminator</td>
			<td>Cerus</td>
			<td>INT100</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>PC sample pack</td>
			<td>Fenwal</td>
			<td>FTX 1122</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Incubator</td>
			<td>Helmer</td>
			<td>PC2200/PC3200</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>Agitator</td>
			<td>Helmer</td>
			<td>PF48H/PF96H</td>
			<td>&nbsp;</td>
		</tr>
		<tr>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>&nbsp;</td>
			<td>Evaluation of in vitro Platelet Function</td>
		</tr>
		<tr>
			<td>Blood gas analyzer</td>
			<td>Radiometer</td>
			<td>ABL 735</td>
			<td>Used for pH, blood gases, and lactate measurement</td>
		</tr>
		<tr>
			<td>Chemistry system</td>
			<td>Ortho Clinical Diagnostic</td>
			<td>Vitros 5.1</td>
			<td>Used for glucose measurement</td>
		</tr>
		<tr>
			<td>Hematology analyzer</td>
			<td>Boule Medical AB</td>
			<td>Medonic CA620-Cellguard</td>
			<td>Used for platelet count measurement</td>
		</tr>
		<tr>
			<td>Flow cytometer</td>
			<td>BD</td>
			<td>FACSCanto</td>
			<td>Used for white blood cell measurement</td>
		</tr>
	</tbody>
</table>

preparation and pathogen inactivation of double dose buffy coat platelet products using the intercept blood system

血液中心正面临着许多挑战，其中包括最大化的产量从血液中他们收到的产品捐赠以及确保最高级别的安全输血的病人，包括输血传播疾病的保护。这必须在财政上负责任的态度，最大限度地减少经营开支，包括消耗品，设备，废物和人员成本，其中包括完成。 有几种方法可用于生产浓缩血小板输血。其中最常见的是血沉棕黄层的制备方法，在该方法中，一个单一的治疗血小板单元（≥2.0×10 11每单位或每当地法规血小板）汇集从六个整个献血的血沉棕黄层。生产"双剂量"的全血血小板的过程只是在最近才得到了发展。 这里介绍的是一种新型的制备方法衍生双倍剂量全血血小板浓缩液池7棕黄色的大衣和随后治疗的双重剂量单位与拦截病原体灭活血液系统。 INTERCEPT灭活病毒，细菌，寄生虫和污染的供体白细胞捐献的血液中可能存在的。配对INTERCEPT的双重剂量的棕黄色大衣方法利用INTERCEPT设置的处理与双存储容器（以下简称"DS集"），允许血液中心来对待每双剂量单位在一个单一的病原体灭活处理组，从而最大限度地提高病人的安全同时最大限度地降低成本。血沉棕黄层的双倍剂量方法需要更少的的血沉棕黄外套和时间使用的消耗品，最多至50％（ 例如，池套，过滤器组，血小板添加剂溶液，和无菌连接晶片）单剂量血沉棕黄层的血小板单位相比，制备和处理。其他节省成本，包括减少浪费，更少的设备maintenan策，更低的功率要求，减少人员的时间，和收集成本较低的血液分离技术相比。

Watch this Scientific Journal Video about Preparation and Pathogen Inactivation of Double Dose Buffy Coat Platelet Products using the INTERCEPT Blood System at JoVE.com

Preparation and Pathogen Inactivation of Double Dose Buffy Coat Platelet Products using the INTERCEPT Blood System

本文介绍了厄勒布鲁大学医院的过程中产生双倍剂量棕黄色外套浓缩血小板捐献全血制备和INTERCEPT血液系统的病原体灭活处理。 "<em在体外</em超过7天的存储质量的最终血小板单位进行评估。

巴菲大衣血小板制品双倍剂量使用INTERCEPT血液系统的制备和病原体灭活

Blood centers are faced with many challenges including maximizing production yield from the blood product donations they receive as well as ensuring the ...

preparation-pathogen-inactivation-double-dose-buffy-coat-platelet

Nanyang Technological University

Research

JoVE Journal

Medicine

28.6K 次观看.  Örebro University Hospital. 本文介绍了厄勒布鲁大学医院的过程中产生双倍剂量棕黄色外套浓缩血小板捐献全血制备和INTERCEPT血液系统的病原体灭活处理。 "

视频: Preparation and Pathogen Inactivation of Double Dose Buffy Coat Platelet Products using the INTERCEPT Blood System

Minimal Erythema Dose (MED) Testing

Ultraviolet radiation (UV) therapy is sometimes used as a treatment for various common skin conditions, including psoriasis, acne, and eczema. The dosage of UV light is prescribed according to an individual's skin sensitivity. Thus, to establish the proper dosage of UV light to administer to a patient, the patient is sometimes screened to determine a minimal erythema dose (MED), which is the amount of UV radiation that will produce minimal erythema (sunburn or redness caused by engorgement of capillaries) of an individual's skin within a few hours following exposure. This article describes how to conduct minimal erythema dose (MED) testing. There is currently no easy way to determine an appropriate UV dose for clinical or research purposes without conducting formal MED testing, requiring observation hours after testing, or informal trial and error testing with the risks of under- or over-dosing. However, some alternative methods are discussed.

Minimal Erythema Dose MED Testing

This article describes how to conduct minimal erythema dose (MED) testing in order to determine the lowest dose of ultraviolet radiation that will cause erythema (burning) when administered to an individual.

最小红斑剂量（MED）测试

Ultraviolet radiation (UV) therapy is  sometimes used as a treatment for various common skin conditions, including  psoriasis, acne, and eczema. The ...

科研

医学

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow (ischemia) 1. Middle cerebral artery (MCA) delivers a great majority of blood to the lateral surface of the cortex 2, is the most common site of human stroke 3, and ischemia within its territory can result in extensive dysfunction or death 1,4,5. Survivors of ischemic stroke often suffer loss or disruption of motor capabilities, sensory deficits, and infarct. In an effort to capture these key characteristics of stroke, and thereby develop effective treatment, a great deal of emphasis is placed upon animal models of ischemia in MCA.
Here we present a method of permanently occluding a cortical surface blood vessel. We will present this method using an example of a relevant vessel occlusion that models the most common type, location, and outcome of human stroke, permanent middle cerebral artery occlusion (pMCAO). In this model, we surgically expose MCA in the adult rat and subsequently occlude via double ligature and transection of the vessel. This pMCAO blocks the proximal cortical branch of MCA, causing ischemia in all of MCA cortical territory, a large portion of the cortex. This method of occlusion can also be used to occlude more distal portions of cortical vessels in order to achieve more focal ischemia targeting a smaller region of cortex. The primary disadvantages of pMCAO are that the surgical procedure is somewhat invasive as a small craniotomy is required to access MCA, though this results in minimal tissue damage. The primary advantages of this model, however, are: the site of occlusion is well defined, the degree of blood flow reduction is consistent, functional and neurological impairment occurs rapidly, infarct size is consistent, and the high rate of survival allows for long-term chronic assessment.

Permanent Cerebral Vessel Occlusion via Double Ligature and Transection

We describe a highly reproducible method for the permanent occlusion of a rodent major cerebral blood vessel. This technique can be accomplished with very little peripheral damage, minimal blood loss, a high rate of long-term survival, and consistent infarct volume commensurate with the human clinical population.

永久性脑血管闭塞<em&gt;通过</em&gt;双人结扎横断

Stroke is a leading cause of death, disability, and socioeconomic loss worldwide. The majority of all strokes result from an interruption in blood flow ...

Drug Repurposing Hypothesis Generation Using the "RE:fine Drugs" System

The promise of drug repurposing is that existing drugs may be used for new disease indications in order to curb the high costs and time for approval. The goal of computational methods for drug repurposing is to enable solutions for safer, cheaper and faster drug discovery. Towards this end, we developed a novel method that integrates genetic and clinical phenotype data from large-scale GWAS and PheWAS studies with detailed drug information on the concept of transitive Drug-Gene-Disease triads. We created "RE:fine Drugs," a freely available, interactive dashboard that automates gene, disease and drug-based searches to identify drug repurposing candidates. This web-based tool supports a user-friendly interface that includes an array of advanced search and export options. Results can be prioritized in a variety of ways, including but not limited to, biomedical literature support, strength and statistical significance of GWAS and/or PheWAS associations, disease indications and molecular drug targets. Here we provide a protocol that illustrates the functionalities available in the "RE:fine Drugs" system and explores the different advanced options through a case study.

Here we describe a protocol using the web-based drug repurposing hypothesis generation tool: "RE:fine Drugs." This protocol can be modified to a user's preferences at the level of the query type (gene, drug or disease) and/or the range of available advanced options.

药品再利用产生假设使用"RE:精药品"系统

The promise of drug repurposing is that existing drugs may be used for new disease indications in order to curb the high costs and time for approval. The ...

Turbidimetry on Human Washed Platelets: The Effect of the Pannexin1-inhibitor Brilliant Blue FCF on Collagen-induced Aggregation

Turbidimetry is a laboratory technique that is applied to measure the aggregation of platelets suspended in either plasma (platelet-rich plasma, PRP) or in buffer (washed platelets), by the use of one or a combination of agonists. The use of washed platelets separated from their plasma environment and in the absence of anticoagulants allows for studying intrinsic platelet properties. Among the large panel of agonists, arachidonic acid (AA), adenosine di-phosphate (ADP), thrombin and collagen are the most frequently used. The aggregation response is quantified by measuring the relative optical density (OD) over time of platelet suspension under continuous stirring. Platelets in homogeneous suspension limit the passage of light after the addition of an agonist, platelet shape change occurs producing a small transitory increase in OD. Following this initial activation step, platelet clots form gradually, allowing the passage of light through the suspension as a result of decreased OD. The aggregation process is ultimately expressed as a percentage, compared to the OD of platelet-poor plasma or buffer. Rigorous calibration is thus essential at the beginning of each experiment. As a general rule: calibration to 0% is set by measuring the OD of a non-stimulated platelet suspension while measuring the OD of the suspension medium containing no platelets represents a value of 100%. Platelet aggregation is generally visualized as a real-time aggregation curve. Turbidimetry is one of the most commonly used laboratory techniques for the investigation of platelet function and is considered as the historical gold standard and used for the development of new pharmaceutical agents aimed at inhibiting platelet aggregation. Here, we describe detailed protocols for 1) preparation of human washed platelets and 2) turbidimetric analysis of collagen-induced aggregation of human washed platelets pretreated with the food dye Brilliant Blue FCF that was recently identified as an inhibitor of Pannexin1 (Panx1) channels.

We describe a straightforward method for the isolation of washed platelets from human blood followed by agonist-induced platelet aggregation measurements by turbidimetry. As an example we apply this method for studying the aggregation response of human platelets to collagen after a pre-incubation with the Pannexin1 channel inhibitor Brilliant Blue FCF.

比浊法对人洗涤血小板:所述Pannexin1抑制剂的影响亮蓝FCF对胶原诱导的聚集

Turbidimetry is a laboratory technique that is applied to measure the aggregation of platelets suspended in either plasma (platelet-rich plasma, PRP) or ...

Using Retinal Imaging to Study Dementia

The retina offers a unique &#8220;window&#8221; to study pathophysiological processes of dementia in the brain, as it is an extension of the central nervous system (CNS) and shares prominent similarities with the brain in terms of embryological origin, anatomical features and physiological properties.&#160; The vascular and neuronal structure in the retina can now be visualized easily and non-invasively using retinal imaging techniques, including fundus photography and optical coherence tomography (OCT), and quantified semi-automatically using computer-assisted analysis programs. Studying the associations between vascular and neuronal changes in the retina and dementia could improve our understanding of dementia and, potentially, aid in diagnosis and risk assessment.&#160; This protocol aims to describe a method of quantifying and analyzing retinal vasculature and neuronal structure, which are potentially associated with dementia. This protocol also provides examples of retinal changes in subjects with dementia, and discusses technical issues and current limitations of retinal imaging.

The retina shares prominent similarities with the brain and thus represents a unique window to study vasculature and neuronal structure in the brain non-invasively. This protocol describes a method to study dementia using retinal imaging techniques. This method can potentially aid in diagnosis and risk assessment of dementia.

利用视网膜成像研究痴呆

The retina offers a unique &#8220;window&#8221; to study pathophysiological processes of dementia in the brain, as it is an extension of the central ...

The Pilocarpine Model of Temporal Lobe Epilepsy and EEG Monitoring Using Radiotelemetry System in Mice

Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status epilepticus (SE) is frequently selected to recapitulate spontaneous recurrent seizures (SRS). Here we present a protocol of SE induction by intraperitoneal (i.p.) injection of pilocarpine and monitoring of chronic recurring seizures in live animals using a wireless telemetry video and electroencephalogram (EEG) system. We demonstrated notable behavioral changes that need attention after pilocarpine injection and their correlation with hippocampal neuronal loss at 7 days and 6 weeks post-pilocarpine. We also describe the experimental procedures of electrode implantation for video and EEG recording, and analysis of the frequency and duration of chronic recurrent seizures. Finally, we discuss the possible reasons why the expected results are not achieved in each case. This provides a basic overview of modeling chronic epilepsy in mice and guidelines for troubleshooting. We believe this protocol can serve as a baseline for suitable models of chronic epilepsy and epileptogenesis.

This manuscript describes a method of inducing status epilepticus by systemic pilocarpine injection and of monitoring spontaneous recurrent seizures in live animals using a wireless telemetry video and electroencephalogram system. This protocol can be utilized for studying the pathophysiologic mechanisms of chronic epilepsy, epileptogenesis, and acute seizures.

小鼠颞叶癫痫的 Radiotelemetry 模型及脑电图监测

Temporal lobe epilepsy (TLE) is a common neurological disorder in adulthood. For translational studies of chronic epilepsy, pilocarpine-induced status ...

Routine Screening Method for Microparticles in Platelet Transfusions

Platelet inventory management based on screening microparticle content in platelet concentrates is a new quality improvement initiative for hospital blood banks. Cells fragment off microparticles (MP) when they are stressed. Blood and blood components may contain cellular fragments from a variety of cells, most notably from activated platelets. When performing their roles as innate immune cells and major players in coagulation and hemostasis, platelets change shape and generate microparticles. With dynamic light scattering (DLS)-based microparticle detection, it is possible to differentiate activated (high microparticle) from non-activated (low microparticle) platelets in transfusions, and optimize the use of this scarce blood product. Previous research suggests that providing non-activated platelets for prophylactic use in hematology-oncology patients could reduce their risk of becoming refractory and improve patient care. The goal of this screening method is to routinely differentiate activated from non-activated platelets. The method described here outlines the steps to be performed for routine platelet inventory management in a hospital blood bank: obtaining a sample from a platelet transfusion, loading the sample into the capillary for DLS measurement, performing the DLS test to identify microparticles, and using the reported microparticle content to identify activated platelets.

Platelet inventory management based on screening microparticle content in platelet concentrates is a new quality improvement initiative in hospital blood banks. The goal is to differentiate activated from non-activated platelets to optimize the use of platelets. Providing non-activated platelets to hematology-oncology patients might reduce their high risk to become refractory.

血小板输血中微粒的常规筛选方法

Platelet inventory management based on screening microparticle content in platelet concentrates is a new quality improvement initiative for hospital blood ...

Preparation, Procedures and Evaluation of Platelet-Rich Plasma Injection in the Treatment of Knee Osteoarthritis

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department. Existing non-surgical treatments have a limited effect on the repair of cartilage and on bone regeneration. Platelet-rich plasma (PRP) is an autologous bioactive substance that can repair cartilage injury and accelerate bone regeneration effectively. However, reporting of PRP preparation protocols in clinical studies is highly inconsistent, with the majority of studies providing insufficient information to allow the protocol to be reproduced. We describe a repeatable method of preparing PRP visually, the treatment of KOA using PRP intra-articular injection, and methods of evaluating the outcome. PRP was prepared using manual double centrifugation. The PRP layer was extracted from peripheral blood and used for knee joint cavity injection. Evaluations included assessments of blood platelet concentrations and clinical outcomes. Preparation of PRP by manual centrifugation requires less apparatus and is less costly than plasma filtration or centrifugation using equipment. The centrifugation time of our double centrifugation method was 6 and 5 minutes for the respective centrifugations at forces of 800 and 1400 x g, respectively, to allow for the consistent preparation of standardized PRP. However, a manual method is susceptible to operator error, and PRP batch preparation is not available. Intra-articular injection of PRP proved to be an effective treatment for knee osteoarthritis. The entire treatment procedure took less than 30 minutes, the blood platelet concentration of PRP could be standardized, and treatment was proven to be effective when evaluated by follow-up.

Knee osteoarthritis is frequently seen in the orthopedic department. We introduce in detail the entire knee osteoarthritis treatment process with platelet-rich plasma injection, including preparation, procedures, and evaluation.

富血小板血浆注射液治疗膝关节骨关节炎的制备、程序及评价

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department. Existing non-surgical treatments have a limited ...

An In Vivo Method for Evaluating the Gut-Blood Barrier and Liver Metabolism of Microbiota Products

The gut-blood barrier (GBB) controls the passage of nutrients, bacterial metabolites and drugs from intestinal lumen to the bloodstream. The GBB integrity is disturbed in gastrointestinal, cardiovascular and metabolic diseases, which may result in easier access of biologically active compounds, such as gut bacterial metabolites, to the bloodstream. Thus, the permeability of the GBB may be a marker of both intestinal and extraintestinal diseases. Furthermore, the increased penetration of bacterial metabolites may affect the functioning of the entire organism.
Commonly used methods for studying the GBB permeability are performed ex vivo. The accuracy of those methods is limited, because the functioning of the GBB depends on intestinal blood flow. On the other hand, commonly used in vivo methods may be biased by liver and kidney performance, as those methods are based on evaluation of urine or/and peripheral blood concentrations of exogenous markers. Here, we present a direct measurement of GBB permeability in rats using an in vivo method based on portal blood sampling, which preserves intestinal blood flow and is virtually not affected by the liver and kidney function. 
Polyurethane catheters are inserted into the portal vein and inferior vena cava just above the hepatic veins confluence. Blood is sampled at baseline and after administration of a selected marker into a desired part of the gastrointestinal tract. Here, we present several applications of the method including (1) evaluation of the colon permeability to TMA, a gut bacterial metabolite, (2) evaluation of liver clearance of TMA, and (3) evaluation of a gut-portal blood-liver-peripheral blood pathway of gut bacteria-derived short-chain fatty acids. Furthermore, the protocol may also be used for tracking intestinal absorption and liver metabolism of drugs or for measurements of portal blood pressure.

An In Vivo Method for Evaluating the Gut-Blood Barrier and Liver Metabolism of Microbiota Products

The access of nutrients, microbiota metabolites and medicines to the circulation is controlled by the gut-blood barrier (GBB). We describe a direct method for measuring the GBB permeability in vivo, which, in contrast to commonly used indirect methods, is virtually not affected by liver and kidney functions.

微生物肠道血屏障及肝脏代谢的体内评价方法

The gut-blood barrier (GBB) controls the passage of nutrients, bacterial metabolites and drugs from intestinal lumen to the bloodstream. The GBB integrity ...

Platelet-based Detection of Nitric Oxide in Blood by Measuring VASP Phosphorylation

Platelets are the blood components responsible for proper blood clotting. Their function is highly regulated by various pathways. One of the most potent vasoactive agents, nitric oxide (NO), can also act as a powerful inhibitor of platelet aggregation. Direct NO detection in blood is very challenging due to its high reactivity with cell-free hemoglobin that limits NO half-life to the millisecond range. Currently, NO changes after interventions are only estimated based on measured changes of nitrite and nitrate (members of the nitrate-nitrite-NO metabolic pathway). However precise, these measurements are rather difficult to interpret vis a vis actual NO changes, due to the naturally high baseline nitrite and nitrate levels that are several orders of magnitude higher than expected changes of NO itself. Therefore, the development of direct and simple methods that would allow one to detect NO directly is long overdue. This protocol addresses a potential use of platelets as a highly sensitive NO sensor in blood. It describes initial platelet rich plasma (PRP) and washed platelet preparations and the use of nitrite and deoxygenated red blood cells as NO generators. Phosphorylation of VASP at serine 239 (P-VASPSer239) is used to detect the presence of NO. The fact that VASP protein is highly expressed in platelets and that it is rapidly phosphorylated when NO is present leads to a unique opportunity to use this pathway to directly detect NO presence in blood.

Here, we present a protocol to address the potential use of platelets as a highly sensitive nitric oxide sensor in blood. It describes initial platelet preparation and the use of nitrite and deoxygenated red blood cells as nitric oxide generators.

板基检测血液中一氧化氮的 vasp 磷酸化

Platelets are the blood components responsible for proper blood clotting. Their function is highly regulated by various pathways. One of the most potent ...