这项工作是由美国心脏协会奖13SDG14500046帕金森病，可持续能源教育与研究中心奖（田纳西大学）PD的支持，并由医师的医学教育和研究基金会（田纳西大学）颁发给JM的作者感谢卡罗琳Leplante（耶鲁大学）的协议转换裂殖酵母原生质球;埃里克T. BODER（田纳西大学）为利用他颤抖孵化器，台式离心机，和Western blot分析设备;和中心环境生物技术（Univ.田纳西州）的使用他们的超离心机;半滑舌鳎道姆（科技，奥地利格拉茨大学）​​酵母抗体;妇产科（Univ.田纳西州医学中心）的技术援助部门的人员。

1。从（裂变）酵母细胞中分离脂滴芽殖酵母酿酒酵母从流行的模式生物滴的隔离几乎是相同的以下协议6。在筹备的差异说明。 1。培养酵母细胞<ol><li>准备媒体。在玻璃瓶或培养瓶中结合36克YE5S粉每升DH 2 O的。约2升的介质是必​​要的。高压灭菌介质在121℃下20分钟。允许介质冷却至室温。对于S。酵母与YPD代替YE5S。 </li><li>放置10-20冷却的YE5S媒体毫升到使用无菌技术的250ml培养瓶中。用少量的从琼脂板用无菌木棍或等效的酵母细胞接种的介质。让细胞生长至所需要的光学密度在30℃下测量在光密度波长的使用分光光度计595nm处。 </li><li>将剩余的介质在2.8升瓶。使用不超过1升介质每2.8升的烧瓶中，以确保细胞生长在振荡培养箱中适当混合。湿细胞的最终产量应为约10克至能够获得足够的脂滴的蛋白质和脂质组分析。 </li><li>引入来自步骤1.2将细胞放入2.8升锥形瓶中分别具有1升的制备步骤1.1的介质。 </li><li>生长的细胞所需的密度在30℃下</li><li>确保该细胞有脂滴。保留1毫升细胞。测量样品的光学密度（OD）。在每个细胞的OD乙醇加0.1毫升BODIPY五百〇三分之四百九十三的100mM储备到样品。放置3微升玻璃载玻片和盖玻片之间的样品。荧光显微镜用GFP过滤器（ 图1A）根据可视化。 </li><li>从步骤1.5颗粒的细胞在4＆＃176，C在3,800 xg离心10分钟。取决于离心管的容量分批工作。 </li><li>倒出YE5S媒体和EMM + 600 1mM的缓冲山梨醇洗涤细胞。山梨糖醇渗透提供支持。 </li><li>将细胞转移到无菌的50ml离心管中，沉淀细胞，在4℃以2000×g离心10分钟。除去上清液。称量湿细胞。重悬在2ml的每克（EMM + 600 mM的山梨糖醇）的缓冲单元的单元。 </li></ol> 2。裂殖酵母细胞转化为原生质球和随后的裂解<ol><li>每添加1 5毫克）酵母溶解酶和2）酵母裂解从木霉每克在步骤1.9 ​​称重的湿细胞的酶。对于S。酵母增强这些酶具有的Zymolyase相同量。 </li><li>孵育的细胞在30℃下温和振摇在100rpm下60分钟。 </li><li>检查以确保球状体具有脂质博士oplets。通过重复步骤1.6（ 图1B）。 </li></ol> 3。密度梯度离心<ol><li>通过离心在4℃下沉淀的原生质球在2000×g离心5分钟。 </li><li>小心地取下用塑料移液管的上清液。 </li><li>轻轻重悬沉淀的原生质球用冰冷的10mM的Tris-HCl，600mM的山梨醇和200μMEDTA中。请记住，球状体是很脆弱的。 </li><li>使用相同的设置步骤3.1再次沉淀的原生质球。小心地取出上清液，并用12％的Ficoll塑料移液管，10毫摩尔Tris-HCl和200μMEDTA以2毫升/克细胞的浓度重悬原生质球。 </li><li>转移的原生质球悬浮于玻璃匀浆器匀化轻轻置于冰上20-30笔画，用玻璃匀浆器的宽松杵。 </li><li>转移裂解原生质球到离心管中，并用覆盖相同的VO12％聚蔗糖，10毫摩尔Tris-HCl和200μMEDTA缓冲液的lume明。选择离心管的数量来使用，因此，每个管大约是三分之二满。 </li><li>离心机在4℃下以100,000×g离心1.5小时在SW28转子或等效物。允许转子惯性停止（转子减速= 0）。 </li><li>上方浮动的白色层（ 图2A），其中包含液滴，轻轻转移到使用一个移液管或弯曲的巴斯德吸管一个新的离心管中（次）。添加足够量的12％Ficoll制备，10毫摩尔Tris-HCl和200μMEDTA缓冲液的样品并使其填满大约三分之一的离心管中。 </li><li>添加8％聚蔗糖的等效体积的10mM Tris-HCl中，200μMEDTA缓冲液中的样品（多个）在新的离心管中（次）的顶部。除了这个缓冲后的管应约三分之二满。 </li><li>离心机在4℃下以100,000×g离心1小时，在SW28转子或等效物。允许转子惯性停止（ROTOR减速= 0）。 </li><li>轻轻转上漂浮的白色层（ 图2B），其中将包含液滴，以使用一个移液器或弯曲巴斯德吸管一个新的离心管（S）。添加600mM的山梨糖醇，8％的Ficoll，10毫摩尔Tris-HCl和200μMEDTA缓冲液的样品并使其填满大约三分之一的离心管。 </li><li>加250毫当量体积山梨醇，10毫摩尔Tris-HCl和200μMEDTA缓冲液中的样品（多个）在新的离心管中（次）的顶部。除了这个缓冲后的管应约三分之二满。 </li><li>离心机在4℃下以100,000×g离心1小时，在SW28转子或等效物。允许转子惯性停止（转子减速= 0）。 </li><li>传输时的白色层（ 图2C），这应该只包含脂滴和结合蛋白，透析管路，透析和O / N为10毫米的Tris-HCl和200微米EDTA缓冲液，以消除聚蔗糖。 </li></ol><p类="jove_title"&gt; 2。从人胎盘绒毛细胞中分离脂滴胎盘是从健康妇女与单胎妊娠择期剖宫产分娩发作自然分娩的足月之前收集的。主题写了，因为他们的胎盘的收集知情同意书。胎盘的收集，以及随后的使用，是与来自田纳西州和医学田纳西研究生院的大学的大学在诺克斯维尔机构审查委员会（＃8757B和＃3338，分别）批准执行。 1。分离人胎盘绒毛细胞该协议的第1部分是较早发布的协议由佩特罗夫等人 26的修改准备以下解决方案: <ul><li>氯化钠（1升）:溶解9克氯化钠（M58.4克/摩尔）在DH 2 O，以产生1 L溶液。过滤消毒（0.2;微米膜）。 </li><li> 10倍的Hank平衡盐溶液（10X HBSS）（1升）:4克氯化钾（M74.5克/摩尔）;0.6克KH 2 PO 4（M136.086克/摩尔），80克氯化钠（M58.44克/摩尔）;的Na 2 HPO 4（M141.96克/摩尔）;10克D-葡萄糖（M180.16克/摩尔）。溶解各构成要素在DH 2 O，以产生1 L溶液。过滤消毒（0.2微米膜）。 </li><li>酶消化缓冲液（0.9毫升）:结合70毫升10倍的HBSS与3.3毫升7.5％的Na Biscarbonate，17.5毫升1 M羟乙基，和266.1毫升DH 2 O，过滤消毒（0.2微米膜）。贮存于4℃。 </li><li> DNA酶I:在使用前，加入5毫升无菌0.9％NaCl溶液，以DNA酶的小瓶。置于冰上直至使用或存放于-20°C。 </li></ul><ol><li>胎盘做准备解剖<ol><li>获得人胎盘和过程尽可能靠近输送尽可能的时间。使用密封容器，如冷却器运送胎盘。请务必小心，当手林志玲人体生物材料。 </li><li>在生物安全罩，胎盘转移到无菌高压灭菌的容器中，并用无菌0.9％氯化钠（食盐）溶液胎盘及胎膜仔细洗血。丢弃血性盐水中的1升烧杯中的液体生物废物。 </li><li>将胎盘在无菌区，与光滑表面轴承朝上脐带。有棱，细点剪刀和镊子取出胎膜和脐带。 </li><li>翻转胎盘过来，使产妇表面（粗糙面）朝上。去除上覆的基底板的组织，从表面约3mm。 </li></ol></li><li>解剖胎盘<ol><li>解剖一个子叶的时间避免绒毛膜板。收集绒毛组织置于250ml烧杯中的盐水。 </li><li>通过用钳子漩，使用1升烧杯中的液体废物冲洗组织几次在0.9％NaCl中的单独的烧杯中。 </li><li>在一次传输一个子叶至150毫米的培养皿。握住钳子和刮组织与从培养皿船只刀片。放置刮掉组织成含有0.9％NaCl的单独的烧杯中。重复所有的组织块。 </li><li>转移组织的小部分细胞解离筛，并用0.9％的NaCl冲洗广泛直至洗出液变清晰。重复所有的组织。 </li><li>漂洗刮下组织中的细胞解离筛后，沥干多余的液体和重量的它。 在这一点上，组织可以被存储O / N中的无菌锥形瓶中，在4℃在DMEM中。 </li></ol></li><li>胎盘组织的酶消化<ol><li>制备组织消化混合物:百毫升预热的酶稀释缓冲液，1毫升DNA酶I和20毫升2.5％的胰蛋白酶10倍。 </li><li>分裂组织，转移60克从总采集的组织（通常约250-300克），以500毫升的无菌锥形瓶中。 </li><lI&gt;添加的组织消化混合物含有该组织的锥形烧瓶中，并在摇床上以150rpm 45分钟孵育在37℃。 </li></ol></li><li>收集脱落的细胞<ol><li>第45分钟解离后，在倾斜设置的消化瓶中，直到组织落户。 </li><li>收集上清液，注意不要收集未解离的组织。添加HBSS等量收集到的上清液并转移到无菌的15ml离心管中。对于未解离组织的额外部分，重复从步骤3.2与未解离的组织开始。 </li><li>离心匀浆液在4℃下以1,000×g离心15分钟。 </li><li>对于每个批次，离心后，吸出上清液，而不会干扰沉淀。胎盘绒毛细胞是主要在沉淀的白色部分，上覆的红血细胞。 </li><li>转让绒毛细胞（颗粒的白色部分）到STerile 50ml锥形离心管中，并保持在冰上。 </li><li>从所有三个阶段消化收集细胞后，用无菌的50ml锥形离心管的顶部插入一个100微米尼龙细胞过滤器过滤悬浮液。如果细胞悬浮液的过滤变慢，向上抬起在过滤器上绘制在管内的真空度。 </li><li>离心机在4℃下以1,000×g离心10分钟。小心取出上清液，而不会干扰颗粒。 </li></ol></li></ol> 2。均质胎盘绒毛细胞<ol><li>首先是隔离过程之前，制备含20毫米的Tris-HCl，pH 7.4和1毫米EDTA50毫升低渗裂解中等（HLM）的。添加蛋白酶抑制剂，以HLM只是在使用前，保持介质上的冰。 </li><li>在生物安全罩中，添加冰冷HLM 4倍的细胞沉淀物体积的细胞。轻轻地，彻底地吹打细胞悬浮细胞向上的D-下，用10毫升吸管。 </li><li>在冰上孵育悬浮细胞10分钟。 </li><li>将细胞转移到Dounce匀浆，这应该是在冰上。慢慢地通过应用20-25温柔的笔触与均质的宽松杵同质化的细胞。 </li><li>离心细胞裂解物在4℃下以3,000×g离心10分钟以除去完整的细胞，细胞碎片和细胞核。 </li><li>收集上清液并转移至SW28管（或替代，可以在25,000 XG离心）。离心机在4℃下以25,000 xg离心20分钟以除去线粒体。 </li></ol> 3。通过离心分离脂滴<ol><li>制备50毫升100毫碳酸钠（M 106克/摩尔）缓冲液（pH 11.5）和50毫升60％蔗糖（w / w的）解决方案。添加蛋白酶抑制剂的碳酸钠缓冲液在使用前并保持在冰上。 </li><li>从步骤2.6收集上清液至50ml离心管中，调整到蔗糖和转移到超速离心管中的SW28转子，或等效的底部的20％。覆盖密度调节上清液用约10毫升冰冷的碳酸盐100mM的钠缓冲液（pH 11.5）和0.5-1毫升冰冷的HLM以填充管。 </li><li>离心机在4℃在130,000 xg下使用SW28水平转头45分钟。允许转子惯性停止（转子减速= 0）。 </li><li>收集漂浮的层，调整到10％的蔗糖和转移到一个13.2毫升超速离心管用于SW41Ti转子，或等效的底部。覆盖密度调节上清液用约5毫升冰冷的100mM碳酸钠缓冲液（pH 11.5）和0.5冰冷HLM毫升。 </li><li>离心机在4℃在274,000 xg下使用SW41Ti水平转头60分钟。允许转子惯性停止（转子减速= 0），以尽量减少脂滴层的破坏。 </li><li>仔细收集含顶部浮动层（ 图2D）脂滴用1毫升吸管，然后重复步骤3.4。 </li><li>离心机在4℃在274,000 xg下使用SW41Ti水平转头30分钟。允许转子惯性停止（转子减速= 0）。 </li><li>仔细收集含脂滴用弯曲巴斯德吸管分为三个1.5毫升管中含有100微升HLM顶白色的浮层。 </li></ol> 表征脂滴组分（协议1和2） 在脂滴分数的回收率和纯度可以通过Western Blot检测结合光或电子显微镜进行验证。此外，可以收集和保存确定净化效率后不同离心步骤等分。在蛋白质印迹法，这是比较合适的，比较脂滴分数表示的全细胞裂解液的当量比在T比较，脂滴至其它膜组分的体积他依据总蛋白含量24。

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作者宣称没有竞争的财务权益。

<html> 本协议中的关键步骤 确保培养细胞的生长过程中以与介质和细胞密度是一致的。细胞脂滴是独特的，因为它们相关的蛋白是高度依赖于在其中培养细胞17的环境。因此，其中的细胞生长介质和细胞的密度应密切裂解之前进行监测。 脂滴的蛋白质组合物是酵母细胞的生长阶段的功能。较少的蛋白质将被绑定到液滴在?...

细胞脂滴是具有多种功​​能的细胞活力的细胞器。它们是存储集线器为中性脂质，可被转化成能量，或用于磷脂的合成。液滴发挥中心作用在生理和病理条件，包括动脉粥样硬化，肥胖和相关代谢性疾病，并且还传染病1,2。此外，他们是耐人寻味的来源生物柴油燃料。 对脂滴的细胞作用的许多内容已经从广泛的生物纯化3滴的蛋白质和脂质组分析中得到。这些生物已包括4,5细菌，酵母6-11，12,13植物，线虫14，和苍蝇15,16。由于在人类的代谢性疾病的脂滴的作用的兴趣，液滴也被分离自培养的动物细胞和nimal组织。培养的细胞系已列出的3T3-L1脂肪细胞17，中国仓鼠卵巢（CHO）细胞中的K2 18，人hepatocyes 19,20和上皮细胞系21。动物组织从液滴被分离已包括小鼠骨骼肌22，肝脏23，和乳腺增生病23。如上所述，大多数液滴隔离研究的目的是执行蛋白质组分析的约束因素，并在中性和磷脂脂质组分析。 由于中性脂肪 - 脂滴的最多的部件 - 比大多数其他细胞材料密度较小，传统上一直采用密度梯度离心法进行飞沫隔离。该技术是在这里提出两个的PREPS的核心。以前的技 ​​术6,24相结合，并修改成液滴从培养裂殖酵母细胞的分离的视觉呈现以及从胎盘组织获得的未培养的人类细胞。我们的目标是通过选择两个截然不同的细胞类型作为起始，在液滴分离点，以显示该技术的广泛适用性。这种技术应该是为那些希望以隔离大多数生物滴有用。 方案1描述了从裂殖酵母， 粟酒裂殖酵母 ，它已被用来作为一个模型用于在真核细胞分裂25观察液滴形成脂滴的分离。芽殖酵母酿酒酵母中已被广泛地用来作为模式生物研究脂滴生物学。方案1是适用于两种生物体和在制剂的差异突出显示。 方案2描述了从胎盘绒毛细胞，这是在从人足月胎盘获得的转脂滴的分离。该收集足月胎盘的提供了一个独特的机会，安全和道德得到200-250克现成的人体组织26，其中包含显著数字脂滴的。这是相对于在高等真核生物中，其中液滴从培养的细胞起源最脂滴隔离工作。在这些研究中，脂肪酸，通常加入到培养以促进中性脂质，因而液滴的生长的合成。这是相对于这里的工作，其中脂质小滴在天然条件下在胎盘组织形成。 在脂滴组分的纯度是通过蛋白质印迹分析用细胞器标记的抗体来确定。这两个协议将产生脂滴部分，适合在随后的蛋白质和脂质组分析。

<table><tbody><tr><td>&lt;strong&gt;Protocol 1&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>&lt;strong&gt;1. Growing yeast cells and converting to spheroplasts&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Edinburgh Minimal Media (EMM)</td><td>Sunrise Science Products</td><td>2005</td><td></td></tr><tr><td>Yeast extract with 5 supplements (YE5S)</td><td>Sunrise Science Products</td><td>2011</td><td>YE5S media with 225 mg/ml of each supplement: adeninie, histidine, leucine, lysine, uracil. The equivalent for budding yeast would be YPD.</td></tr><tr><td>YPD powder</td><td>Sunrise Science Products</td><td>1875</td><td>For &lt;i&gt;S. cerevisiae&lt;/i&gt;</td></tr><tr><td>Sorbitol</td><td>Fisher Scientific</td><td>BP439</td><td></td></tr><tr><td>Yeast Lytic Enzyme</td><td>MP Biomedicals</td><td>215352610</td><td></td></tr><tr><td>Lysing Enzymes from Trichoderma harzianum</td><td>Sigma-Aldrich</td><td>L1412</td><td></td></tr><tr><td>Zymolayse-20T</td><td>Sunrise Science Products</td><td>N0766391</td><td>For &lt;i&gt;S. cerevisiae&lt;/i&gt;</td></tr><tr><td>BODIPY 493/503</td><td>Invitrogen</td><td>D-3922</td><td></td></tr><tr><td>Microscope Slides</td><td>Fisher Scientific</td><td>12-544-7</td><td></td></tr><tr><td>Microscope Cover Glass</td><td>Fisher Scientific</td><td>12-542-B</td><td></td></tr><tr><td>Plastic transfer pipette</td><td>Fisher Scientific</td><td>137115AM</td><td></td></tr><tr><td>1 L glass bottle</td><td></td><td></td><td></td></tr><tr><td>250 ml flask</td><td></td><td></td><td></td></tr><tr><td>2.8 L flasks</td><td></td><td></td><td></td></tr><tr><td>&lt;strong&gt;2. Yeast lipid droplet isolation&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Tris-HCl</td><td>Fisher Scientific</td><td>BP153</td><td></td></tr><tr><td>EDTA</td><td>Fisher Scientific</td><td>BP120</td><td></td></tr><tr><td>Ficoll 400</td><td>Fisher Scientific</td><td>BP525</td><td></td></tr><tr><td>12-14k Spectra/Por Dialysis Membrane</td><td>SpectrumLabs</td><td>132680</td><td></td></tr><tr><td>EDTA-free Protease Inhibitor Cocktail Tablets</td><td>Roche Diagnostics</td><td>11873580001</td><td>irritant</td></tr><tr><td>Dounce Homogenizer&amp;nbsp;</td><td>Sigma-Aldrich</td><td>D9938</td><td></td></tr><tr><td>Ultracentrifuge Tubes 25 mm x 89 mm (for SW28)</td><td>Beckman-Coulter</td><td>355642</td><td></td></tr><tr><td>12-14k Spectra/Por Dialysis Membrane</td><td>SpectrumLabs</td><td>132680</td><td></td></tr><tr><td>Temperature-controlled shaker</td><td>New Brunswick Scientific</td><td>C25KC</td><td></td></tr><tr><td>Thermo Sorvall Legend XTR centrifuge</td><td>Thermo-Scientific</td><td>75004521</td><td></td></tr><tr><td>Swinging Bucket Centrifuge Rotor</td><td>Thermo-Scientific</td><td>75003607</td><td></td></tr><tr><td>Fiberlite* F15-6x100y Fixed-Angle Rotor</td><td>Thermo-Scientific</td><td>75003698</td><td></td></tr><tr><td>Ultracentrifuge LB-M</td><td>Beckman-Coulter</td><td></td><td></td></tr><tr><td>SW28 Ultracentrifuge Rotor</td><td>Beckman-Coulter</td><td>342204</td><td></td></tr><tr><td>&lt;strong&gt;Protocol 2&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>&lt;strong&gt;1. Placental villous cells isolation&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Disposable underpads</td><td>Fisher Scientific</td><td>23666062</td><td></td></tr><tr><td>Autoclavable pan (container), 3 L</td><td>Fisher Scientific</td><td>1336110</td><td></td></tr><tr><td>Fine scissors, sharp-sharp, straight</td><td>Fine Science Tools</td><td>1406011</td><td></td></tr><tr><td>London Forceps</td><td>Fine Science Tools</td><td>1108002</td><td></td></tr><tr><td>Dumont #7b Forceps</td><td>Fine Science Tools</td><td>1127020</td><td></td></tr><tr><td>Razor blades</td><td>Fisher Scientific</td><td>S65921</td><td></td></tr><tr><td>Screen cup for CD-1</td><td>Fisher Scientific</td><td>S1145</td><td></td></tr><tr><td>40 mesh screen&amp;nbsp;</td><td>Fisher Scientific</td><td>S0770</td><td></td></tr><tr><td>Fisherbrand cell stainers 100 &amp;mu;m</td><td>Fisher Scientific</td><td>22363549</td><td></td></tr><tr><td>150 mm Petri Dishes</td><td>Fisher Scientific</td><td>NC9054771</td><td></td></tr><tr><td>NaCl</td><td>Fisher Scientific</td><td>S642</td><td></td></tr><tr><td>KCl</td><td>Fisher Scientific</td><td>P333</td><td></td></tr><tr><td>KH&lt;sub&gt;2&lt;/sub&gt;PO&lt;sub&gt;4&lt;/sub&gt;</td><td>Fisher Scientific</td><td>P386</td><td></td></tr><tr><td>Na&lt;sub&gt;2&lt;/sub&gt;HPO&lt;sub&gt;4&lt;/sub&gt;</td><td>Fisher Scientific</td><td>S374</td><td></td></tr><tr><td>D-Glucose</td><td>Fisher Scientific</td><td>D16</td><td></td></tr><tr><td>HEPES</td><td>Fisher Scientific</td><td>BP310</td><td></td></tr><tr><td>2.5% Trypsin 10x</td><td>Invitrogen</td><td>15090046</td><td></td></tr><tr><td>DNase I grade II, from bovine pancreas</td><td>Roche Applied Science</td><td>10104159001</td><td></td></tr><tr><td>Sodium bicarbonate solution</td><td>Sigma-Aldrich</td><td>S8761</td><td></td></tr><tr><td>500 ml Erlenmeyer flasks</td><td></td><td></td><td></td></tr><tr><td>250 ml Beakers</td><td></td><td></td><td></td></tr><tr><td>15 ml Centrifuge tubes</td><td></td><td></td><td></td></tr><tr><td>10 ml Serological pipettes</td><td></td><td></td><td></td></tr><tr><td>50 ml Centrifuge tubes</td><td></td><td></td><td></td></tr><tr><td>DMEM</td><td>Invitrogen</td><td>11965084</td><td></td></tr><tr><td>&lt;strong&gt;2. Lipid droplets isolation from villous placental cells&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Tris-HCl</td><td>Fisher Scientific</td><td>BP153</td><td></td></tr><tr><td>EDTA</td><td>Fisher Scientific</td><td>BP120</td><td></td></tr><tr><td>D-Sucrose</td><td>Fisher Scientific</td><td>BP220</td><td></td></tr><tr><td>Sodium Carbonate&amp;nbsp;</td><td>Fisher Scientific</td><td>BP357</td><td></td></tr><tr><td>EDTA-free protease inhibitor cocktail tablets</td><td>Roche Diagnostics</td><td>11873580001</td><td>irritant</td></tr><tr><td>Dounce homogenizer&amp;nbsp;</td><td>Sigma-Aldrich</td><td>D9938</td><td></td></tr><tr><td>Ultracentrifuge tubes 25 mm x 89 mm (for SW28)</td><td>Beckman-Coulter</td><td>355642</td><td></td></tr><tr><td>Ultra-Clear centrifuge tubes 14 mm x 89 mm (for SW41)</td><td>Beckman-Coulter</td><td>344059</td><td></td></tr><tr><td>Disposable borosilicate glass Pasteur pipets</td><td>Fisher Scientific</td><td>1367820C</td><td></td></tr><tr><td>Biological safety hood&amp;nbsp;</td><td>Thermo-Scientific</td><td></td><td></td></tr><tr><td>Waterbath</td><td>Fisher Scientific</td><td></td><td></td></tr><tr><td>Temperature-controlled shaker</td><td>New Brunswick Scientific</td><td>C25KC</td><td></td></tr><tr><td>Thermo Sorvall Legend XTR centrifuge</td><td>Thermo-Scientific</td><td>75004521</td><td></td></tr><tr><td>Swinging Bucket Centrifuge Rotor</td><td>Thermo-Scientific</td><td>75003607</td><td></td></tr><tr><td>Ultracentrifuge LB-M</td><td>Beckman-Coulter</td><td></td><td></td></tr><tr><td>SW28 Ultracentrifuge Rotor</td><td>Beckman-Coulter</td><td>342204</td><td></td></tr><tr><td>SW41 Ti Ultracentrifuge Rotor</td><td>Beckman-Coulter</td><td>331336</td><td></td></tr><tr><td>&lt;strong&gt;Western blot&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>IRDye 680 Goat Anti-Rabbit IgG</td><td>LI-COR</td><td>926-68071</td><td>dilution 1:15,000</td></tr><tr><td>IRDye 800CW Goat Anti-Mouse IgG</td><td>LI-COR</td><td>926-32210</td><td>dilution 1:5,000</td></tr><tr><td>NuPAGE Novex 12% Bis-Tris gels</td><td>Invitrogen</td><td>NP0341</td><td></td></tr><tr><td>&lt;strong&gt;Primary antibodies for Protocol 1&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Erg6p</td><td>gift from Dr. G. Daum</td><td>Graz University of Technology, Austria</td><td>dilution 1:5,000</td></tr><tr><td>Dpm1p</td><td>Abcam</td><td>ab113686</td><td>4 &amp;mu;g/ml</td></tr><tr><td>Por1p</td><td>gift from Dr. G. Daum</td><td>Graz University of Technology, Austria</td><td>dilution 1:5,000</td></tr><tr><td>Pma1p</td><td>gift from Dr. G. Daum</td><td>Graz University of Technology, Austria</td><td>dilution 1:10,000</td></tr><tr><td>Vma1p (anti-ATP6V1A)</td><td>Abcam</td><td>ab113745</td><td>0.5 &amp;mu;g/ml</td></tr><tr><td>&lt;strong&gt;Primary antibodies for Protocol 2&lt;/strong&gt;</td><td></td><td></td><td></td></tr><tr><td>Perilipin 2 (anti-ADFP)</td><td>Abcam</td><td>ab52355</td><td>2 &amp;mu;g/ml</td></tr><tr><td>Calnexin</td><td>Cell Signaling Technology</td><td>2679</td><td>dilution 1:1,000</td></tr><tr><td>GM130</td><td>Biorbyt</td><td>orb40533</td><td>dilution 1:25</td></tr><tr><td>COX IV</td><td>Cell Signaling Technology</td><td>4850</td><td>dilution 1:1,000</td></tr><tr><td>MEK1</td><td>Biorbyt</td><td>orb38775</td><td>dilution 1:50</td></tr></tbody></table>

isolation of cellular lipid droplets: two purification techniques starting from yeast cells and human placentas

脂滴是可以在大多数真核细胞和某些原核细胞中找到动态的细胞器。在结构上，所述液滴包括由磷脂单分子层所包围的中性脂质的核心。一种在确定液滴的细胞作用最有用的技术一直是蛋白质组学鉴定结合的蛋白，它可以随着液滴被分离的。在这里，两种方法都描述隔离脂滴和他们结合蛋白从两个广泛的真核生物:裂殖酵母和人类胎盘绒毛细胞。虽然这两种技术有差异，主要方法 - 密度梯度离心 - 是由两个准备共享。这显示了所提出的液滴分离技术的广泛适用性。 在第一协议，酵母细胞是将它们的细胞壁消化酶转化成原生质球。由此产生的原生质球，然后GENTLY溶解在宽松的均质机。聚蔗糖加入到裂解物中，以提供密度梯度，将混合物离心分离3次。第一自旋后，脂质微滴被定位于离心管中的白色的浮动层沿与内质网（ER），质膜，并且空泡。随后的两个自旋是用来消除这些其他三个细胞器。其结果是，仅具有小滴和结合蛋白的层。 在第二协议，胎盘绒毛细胞是从人的足月胎盘酶消化，用胰蛋白酶和DNA酶I。将细胞匀浆在一个宽松的匀浆中分离。低速和中速离心步骤用于去除完整的细胞，细胞碎片，细胞核，线粒体和。蔗糖加入到匀浆，以提供密度梯度，将混合物离心，以脂质小滴从其他cellula分开ř分数。 在这两种协议的脂滴的纯度是通过Western印迹分析来证实。从双方的PREPS液滴部分适用于后续的蛋白质和脂质组分析。

如果密度梯度离心和预期一样，浮层应含有脂滴和在整个的高速旋转的进展耗尽其他细胞器。 对于方案1，西方印迹与标记抗体的脂滴（Erg6p），并且已经发现，与在酵母中，ER（Dpm1p），线粒体（Por1p）脂滴的细胞器相互作用进行的，质膜（Pma1p）和液泡（Vma1p）。 每3自旋的浮动层（步骤3.7，3.10，和3.13）收集等量，沉淀用三氯乙酸（15％终浓度），并溶解...

Watch this Scientific Journal Video about Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas at JoVE.com

Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas

两种技术从1分离细胞的脂滴），酵母细胞和2）人类胎盘介绍。这两个程序的核心是密度梯度离心法，在这里可以用肉眼很容易地可视化含有液滴产生的浮层，提取，并通过Western印迹分析纯度量化。

隔离细胞脂滴:两个纯化技术从酵母细胞与人类的胎盘开始

Lipid droplets are dynamic organelles that can be found in most eukaryotic and certain prokaryotic cells. Structurally, the droplets consist of a core of ...

isolation-cellular-lipid-droplets-two-purification-techniques

Universidad de Cartagena UDEC

Research

JoVE Journal

Bioengineering

19.6K 次观看.  University of Tennessee. 两种技术从1分离细胞的脂滴），酵母细胞和2）人类胎盘介绍。这两个程序的核心是密度梯度离心法，在这里可以用肉眼很容易地可视化含有液滴产生的浮层，提取，并通过Western印迹分析纯度量化。 

视频: Isolation of Cellular Lipid Droplets: Two Purification Techniques Starting from Yeast Cells and Human Placentas

Isolation of High-density Lipoproteins for Non-coding Small RNA Quantification

The diversity of small non-coding RNAs (sRNA) is rapidly expanding and their roles in biological processes, including gene regulation, are emerging. Most interestingly, sRNAs are also found outside of cells and are stably present in all biological fluids. As such, extracellular sRNAs represent a novel class of disease biomarkers and are likely involved in cell signaling and intercellular communication networks. To assess their potential as biomarkers, sRNAs can be quantified in plasma, urine, and other fluids. Nevertheless, to fully understand the impact of extracellular sRNAs as endocrine signals, it is important to determine which carriers are transporting and protecting them in biological fluids (e.g., plasma), which cells and tissues contribute to extracellular sRNA pools, and cells and tissues capable of accepting and utilizing extracellular sRNA. To accomplish these goals, it is critical to isolate highly pure populations of extracellular carriers for sRNA profiling and quantification. We have previously demonstrated that lipoproteins, particularly high-density lipoproteins (HDL), transport functional microRNAs (miRNA) between cells and HDL-miRNAs are significantly altered in disease. Here, we detail a new protocol that utilizes tandem HDL isolation with density-gradient ultracentrifugation (DGUC) and fast-protein-liquid chromatography (FPLC) to obtain highly pure HDL for downstream profiling and quantification of all sRNAs, including miRNAs, using both high-throughput sequencing and real-time PCR approaches. This protocol will be a valuable resource for the investigation of sRNAs on HDL.

This protocol describes the isolation and quantification of high-density lipoprotein small RNAs.

高密度脂蛋白的分离的非编码小RNA定量

The diversity of small non-coding RNAs (sRNA) is rapidly expanding and their roles in biological processes, including gene regulation, are emerging. Most ...

科研

生物化学

Lipid Droplet Isolation for Quantitative Mass Spectrometry Analysis

Lipid droplets are vital to the replication of a variety of different pathogens, most prominently the Hepatitis C Virus (HCV), as the putative site of virion morphogenesis. Quantitative lipid droplet proteome analysis can be used to identify proteins that localize to or are displaced from lipid droplets under conditions such as virus infections. Here, we describe a protocol that has been successfully used to characterize the changes in the lipid droplet proteome following infection with HCV. We use Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC) and thus label the complete proteome of one population of cells with &#34;heavy&#34; amino acids to quantitate the proteins by mass spectrometry. For lipid droplet isolation, the two cell populations (i.e.&#160;HCV-infected/&#34;light&#34; amino acids and uninfected control/&#34;heavy&#34; amino acids) are mixed 1:1 and lysed mechanically in hypotonic buffer. After removing the nuclei and cell debris by low speed centrifugation, lipid droplet-associated proteins are enriched by two subsequent ultracentrifugation steps followed by three washing steps in isotonic buffer. The purity of the lipid droplet fractions is analyzed by western blotting with antibodies recognizing different subcellular compartments. Lipid droplet-associated proteins are then separated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Coomassie staining. After tryptic digest, the peptides are quantified by liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS). Using this method, we identified proteins recruited to lipid droplets upon HCV infection that might represent pro- or antiviral host factors. Our method can be applied to a variety of different cells and culture conditions, such as infection with pathogens, environmental stress, or drug treatment.

Lipid droplets are important organelles for the replication of several pathogens, including the Hepatitis C Virus (HCV). We describe a method to isolate lipid droplets for quantitative mass spectrometry of associated proteins; it can be used under a variety of conditions, such as virus infection, environmental stress, or drug treatment.

脂滴隔离定量质谱分析

Lipid droplets are vital to the replication of a variety of different pathogens, most prominently the Hepatitis C Virus (HCV), as the putative site of ...

Quantitative Analysis of the Cellular Lipidome of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

Lipids are structurally diverse amphipathic molecules that are insoluble in water. Lipids are essential contributors to the organization and function of biological membranes, energy storage and production, cellular signaling, vesicular transport of proteins, organelle biogenesis, and regulated cell death. Because the budding yeast Saccharomyces cerevisiae is a unicellular eukaryote amenable to thorough molecular analyses, its use as a model organism helped uncover mechanisms linking lipid metabolism and intracellular transport to complex biological processes within eukaryotic cells. The availability of a versatile analytical method for the robust, sensitive, and accurate quantitative assessment of major classes of lipids within a yeast cell is crucial for getting deep insights into these mechanisms. Here we present a protocol to use liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) for the quantitative analysis of major cellular lipids of S. cerevisiae. The LC-MS/MS method described is versatile and robust. It enables the identification and quantification of numerous species (including different isobaric or isomeric forms) within each of the 10 lipid classes. This method is sensitive and allows identification and quantitation of some lipid species at concentrations as low as 0.2 pmol/&#181;L. The method has been successfully applied to assessing lipidomes of whole yeast cells and their purified organelles. The use of alternative mobile phase additives for electrospray ionization mass spectrometry in this method can increase the efficiency of ionization for some lipid species and can be therefore used to improve their identification and quantitation.

Quantitative Analysis of the Cellular Lipidome of Saccharomyces Cerevisiae Using Liquid Chromatography Coupled with Tandem Mass Spectrometry

We present a protocol using liquid chromatography coupled with tandem mass spectrometry to identify and quantify major cellular lipids in Saccharomyces cerevisiae. The described method for a quantitative assessment of major lipid classes within a yeast cell is versatile, robust, and sensitive.

使用液色谱结合串联质谱法对糖皮质酶细胞脂质的定量分析

Lipids are structurally diverse amphipathic molecules that are insoluble in water. Lipids are essential contributors to the organization and function of ...

Isolation and Processing of Murine White Adipocytes for Transcriptome and Epigenome Analyses

Obesity is a complex disease influenced by genetics, epigenetics, the environment, and their interactions. Mature adipocytes represent the major cell type in white adipose tissue. Understanding how adipocytes function and respond to (epi)genetic and environmental signals is essential for identifying the cause(s) of obesity. RNA and chromatin have previously been isolated from adipocytes using enzymatic digestion. In addition, protocols have been developed for nuclear isolation, where purification is achieved by fluorescence-activated cell sorting (FACS) of adipocyte-specific transgenic reporters. One of the greatest challenges to achieving high yield and quality during such protocols is the substantial amount of lipid contained in adipose tissue. The present protocol describes an optimized procedure for isolating mature adipocytes that leverages heptane to separate lipids from the targets of interest (RNA/chromatin). The resulting RNA has high integrity and generates high-quality RNA-seq results. Likewise, the procedure improves nuclei yield rate and generates reproducible ChIP-seq results across samples. Therefore, the current study provides a reliable and universal murine adipocyte isolation protocol suitable for whole-genome transcriptome and epigenome studies.

The present protocol summarizes a universal method for isolating, purifying, and upstream processing of murine white adipocytes optimized for downstream total RNA sequencing, Nuclei Extraction by SONication (NEXSON), and ChIP-seq.

Obesity is a complex disease influenced by genetics, epigenetics, the environment, and their interactions. Mature adipocytes represent the major cell type ...

遗传学

A Quantitative Assessment of The Yeast Lipidome using Electrospray Ionization Mass Spectrometry

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast Saccharomyces cerevisiae, a genetically and biochemically manipulable unicellular eukaryote with annotated genome and very simple lipidome, is a valuable model for studying biological functions of various lipid species in multicellular eukaryotes2,3,5. S. cerevisiae has 10 major classes of lipids with chain lengths mainly of 16 or 18 carbon atoms and either zero or one degree of unsaturation6,7. Existing methods for lipid identification and quantification - such as high performance liquid chromatography, thin-layer chromatography, fluorescence microscopy, and gas chromatography followed by MS - are well established but have low sensitivity, insufficiently separate various molecular forms of lipids, require lipid derivitization prior to analysis, or can be quite time consuming. Here we present a detailed description of our experimental approach to solve these inherent limitations by using survey-scan ESI/MS for the identification and quantification of the entire complement of lipids in yeast cells. The described method does not require chromatographic separation of complex lipid mixtures recovered from yeast cells, thereby greatly accelerating the process of data acquisition. This method enables lipid identification and quantification at the concentrations as low as g/ml and has been successfully applied to assessing lipidomes of whole yeast cells and their purified organelles. Lipids extraction from whole yeast cells for using this method of lipid analysis takes two to three hours. It takes only five to ten minutes to run each sample of extracted and dried lipids on a Q-TOF mass spectrometer equipped with a nano-electrospray source.

We describe a new quantitative lipidomics method for identifying numerous lipid species in yeast using survey-scan electrospray ionization mass spectrometry (ESI/MS). This method exceeds currently available methods for lipid identification and quantification in the ability to resolve various molecular forms of lipids, sensitivity, and speed.

一个定量评估的酵母使用电喷雾质谱法Lipidome

Lipids are one of the major classes of biomolecules and play important roles membrane dynamics, energy storage, and signalling1-4. The budding yeast ...

生物学

Analysis of Neutral Lipid Synthesis in Saccharomyces cerevisiae by Metabolic Labeling and Thin Layer Chromatography

Neutral lipids (NLs) are a class of hydrophobic, chargeless biomolecules that play key roles in energy and lipid homeostasis. NLs are synthesized de novo&#160;from acetyl-CoA and are primarily present in eukaryotes in the form of triglycerides (TGs) and sterol-esters (SEs). The enzymes responsible for the synthesis of NLs are highly conserved from Saccharomyces cerevisiae (yeast) to humans, making yeast a useful model organism to dissect the function and regulation of NL metabolism enzymes. While much is known about how acetyl-CoA is converted into a diverse set of NL species, mechanisms for regulating NL metabolism enzymes, and how mis-regulation can contribute to cellular pathologies, are still being discovered. Numerous methods for the isolation and characterization of NL species have been developed and used over decades of research; however, a quantitative and simple protocol for the comprehensive characterization of major NL species has not been discussed. Here, a simple and adaptable method to quantify the de novo&#160;synthesis of major NL species in yeast is presented. We apply 14C-acetic acid metabolic labeling coupled with thin layer chromatography to separate and quantify a diverse range of physiologically important NLs. Additionally, this method can be easily applied to study in vivo reaction rates of NL enzymes or degradation of NL species over time.

Analysis of Neutral Lipid Synthesis in Saccharomyces cerevisiae by Metabolic Labeling and Thin Layer Chromatography

Here, a protocol is presented for the metabolic labeling of yeast with 14C-acetic acid, which is coupled with thin layer chromatography for the separation of neutral lipids.

通过代谢标记和薄层色谱分析 糖浆 中中性脂质合成

Neutral lipids (NLs) are a class of hydrophobic, chargeless biomolecules that play key roles in energy and lipid homeostasis. NLs are synthesized de ...

Analysis of Lipid Droplet Content in Fission and Budding Yeasts using Automated Image Processing

Lipid metabolism and its regulation are of interest to both basic and applied life sciences and biotechnology. In this regard, various yeast species are used as models in lipid metabolic research or for industrial lipid production. Lipid droplets are highly dynamic storage bodies and their cellular content represents a convenient readout of the lipid metabolic state. Fluorescence microscopy is a method of choice for quantitative analysis of cellular lipid droplets, as it relies on widely available equipment and allows analysis of individual lipid droplets. Furthermore, microscopic image analysis can be automated, greatly increasing overall analysis throughput. Here, we describe an experimental and analytical workflow for automated detection and quantitative description of individual lipid droplets in three different model yeast species: the fission yeasts Schizosaccharomyces pombe and Schizosaccharomyces japonicus, and the budding yeast Saccharomyces cerevisiae. Lipid droplets are visualized with BODIPY 493/503, and cell-impermeable fluorescent dextran is added to the culture media to help identify cell boundaries. Cells are subjected to 3D epifluorescence microscopy in green and blue channels and the resulting z-stack images are processed automatically by a MATLAB pipeline. The procedure outputs rich quantitative data on cellular lipid droplet content and individual lipid droplet characteristics in a tabular format suitable for downstream analyses in major spreadsheet or statistical packages. We provide example analyses of lipid droplet content under various conditions that affect cellular lipid metabolism.

Here, we present a MATLAB implementation of automated detection and quantitative description of lipid droplets in fluorescence microscopy images of fission and budding yeast cells.

使用自动图像处理分析裂变和萌芽酵母中的脂滴含量

Lipid metabolism and its regulation are of interest to both basic and applied life sciences and biotechnology. In this regard, various yeast species are ...

免疫与感染

Rapid Lipid Droplet Isolation Protocol Using a Well-established Organelle Isolation Kit

Lipid droplets (LDs) are bioactive organelles found within the cytosol of the most eukaryotic and some prokaryotic cells. LDs are composed of neutral lipids encased by a monolayer of phospholipids and proteins. Hepatic LD lipids, such as ceramides, and proteins are implicated in several diseases that cause hepatic steatosis. Although previous methods have been established for LD isolation, they require a time-consuming preparation of reagents and are not designed for the isolation of multiple subcellular compartments. We sought to establish a new protocol to enable the isolation of LDs, endoplasmic reticulum (ER), and lysosomes from a single mouse liver.
Further, all reagents used in the protocol presented here are commercially available and require minimal reagent preparation without sacrificing LD purity. Here we present data comparing this new protocol to a standard sucrose gradient protocol, demonstrating comparable purity, morphology, and yield. Additionally, we can isolate ER and lysosomes using the same sample, providing detailed insight into the formation and intracellular flux of lipids and their associated proteins.

This protocol establishes a novel method of lipid droplet isolation and purification from mouse livers, using a well-established endoplasmic reticulum isolation kit.

利用成熟的细胞隔离试剂盒快速分离脂滴器

Lipid droplets (LDs) are bioactive organelles found within the cytosol of the most eukaryotic and some prokaryotic cells. LDs are composed of neutral ...

Isolation and Characterization of Human Adipocyte-Derived Extracellular Vesicles using Filtration and Ultracentrifugation

Extracellular vesicles (EVs) are lipid enclosed envelopes that carry biologically active material such as proteins, RNA, metabolites and lipids. EVs can modulate the cellular status of other cells locally in tissue microenvironments or through liberation into peripheral blood. Adipocyte-derived EVs are elevated in the peripheral blood and show alterations in their cargo (RNA and protein) during metabolic disturbances, including obesity and diabetes. Adipocyte-derived EVs can regulate the cellular status of neighboring vascular cells, such as endothelial cells and adipose tissue resident macrophages to promote adipose tissue inflammation. Investigating alterations in adipocyte-derived EVs in vivo is complex because EVs derived from peripheral blood are highly heterogenous and contain EVs from other sources, namely platelets, endothelial cells, erythrocytes and muscle. Therefore, the culture of human adipocytes provides a model system for the study of adipocyte derived EVs. Here, we provide a detailed protocol for the extraction of total small EVs from cell culture media of human gluteal and abdominal adipocytes using filtration and ultracentrifugation. We further demonstrate the use of Nanoparticle Tracking Analysis (NTA) for quantification of EV size and concentration and show the presence of EV-protein tumor susceptibility gene 101 (TSG101) in the gluteal and abdominal adipocyte derived-EVs. Isolated EVs from this protocol can be used for downstream analysis, including transmission electron microscopy, proteomics, metabolomics, small RNA-sequencing,&#160;microarrays and can be&#160;utilized in functional in vitro/in vivo studies.

We describe the isolation of human adipocyte-derived extracellular vesicles (EVs) from gluteal and abdominal adipose tissue using filtration and ultracentrifugation. We characterize the isolated adipocyte-derived EVs by determining their size and concentration by Nanoparticle Tracking Analysis and by western blotting for the presence of EV-protein tumor susceptibility gene 101 (TSG101).

Extracellular vesicles (EVs) are lipid enclosed envelopes that carry biologically active material such as proteins, RNA, metabolites and lipids. EVs can ...

Plastoglobule Lipid Droplet Isolation from Plant Leaf Tissue and Cyanobacteria

Plastoglobule lipid droplets are a dynamic sub-compartment of plant chloroplasts and cyanobacteria. Found ubiquitously among photosynthetic species, they are believed to serve a central role in the adaptation and remodeling of the thylakoid membrane under rapidly changing environmental conditions. The capacity to isolate plastoglobules of high purity has greatly facilitated their study through proteomic, lipidomic, and other methodologies. With plastoglobules of high purity and yield, it is possible to investigate their lipid and protein composition, enzymatic activity, and protein topology, among other possible molecular characteristics. This article presents a rapid and effective protocol for the isolation of plastoglobules from chloroplasts of plant leaf tissue and presents methodological variations for the isolation of plastoglobules and related lipid droplet structures from maize leaves, the desiccated leaf tissue of the resurrection plant, Eragrostis nindensis, and the cyanobacterium, Synechocystis sp. PCC 6803. Isolation relies on the low density of these lipid-rich particles, which facilitates their purification by sucrose density flotation. This methodology will prove valuable in the study of plastoglobules from diverse species.

A fast and efficient protocol is presented for the isolation of plastoglobule lipid droplets associated with various photosynthetic organisms. The successful preparation of isolated plastoglobules is a crucial first step that precedes detailed molecular investigations such as proteomic and lipidomic analyses.

从植物叶片组织和蓝藻中分离出塑体球脂滴

Plastoglobule lipid droplets are a dynamic sub-compartment of plant chloroplasts and cyanobacteria. Found ubiquitously among photosynthetic species, they ...

隔离细胞脂滴:两个纯化技术从酵母细胞与人类的胎盘开始

摘要

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此视频中的章节

高密度脂蛋白的分离的非编码小RNA定量

脂滴隔离定量质谱分析

使用液色谱结合串联质谱法对糖皮质酶细胞脂质的定量分析

Isolation and Processing of Murine White Adipocytes for Transcriptome and Epigenome Analyses

一个定量评估的酵母使用电喷雾质谱法Lipidome

通过代谢标记和薄层色谱分析糖浆中中性脂质合成

使用自动图像处理分析裂变和萌芽酵母中的脂滴含量

利用成熟的细胞隔离试剂盒快速分离脂滴器

Isolation and Characterization of Human Adipocyte-Derived Extracellular Vesicles using Filtration and Ultracentrifugation

从植物叶片组织和蓝藻中分离出塑体球脂滴

隔离细胞脂滴:两个纯化技术从酵母细胞与人类的胎盘开始

摘要

探索更多视频

此视频中的章节

高密度脂蛋白的分离的非编码小RNA定量

脂滴隔离定量质谱分析

使用液色谱结合串联质谱法对糖皮质酶细胞脂质的定量分析

Isolation and Processing of Murine White Adipocytes for Transcriptome and Epigenome Analyses

一个定量评估的酵母使用电喷雾质谱法Lipidome

通过代谢标记和薄层色谱分析 糖浆 中中性脂质合成

使用自动图像处理分析裂变和萌芽酵母中的脂滴含量

利用成熟的细胞隔离试剂盒快速分离脂滴器

Isolation and Characterization of Human Adipocyte-Derived Extracellular Vesicles using Filtration and Ultracentrifugation

从植物叶片组织和蓝藻中分离出塑体球脂滴

通过代谢标记和薄层色谱分析糖浆中中性脂质合成