Establishment of a simplified dichotomic size-exclusion chromatography for isolating extracellular vesicles toward clinical applications.
Establishment of a simplified dichotomic size-exclusion chromatography for isolating extracellular vesicles toward clinical applications.
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建立简化的二分尺寸排阻色谱法用于分离细胞外囊泡以供临床应用
DOI：
10.1002/jev2.12145
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发表时间：
2021-09
影响因子：
16
通讯作者：
Cui Y
中科院分区：
文献类型：
--
作者：
Guo J;Wu C;Lin X;Zhou J;Zhang J;Zheng W;Wang T;Cui Y
尺寸 - 排斥色谱（SEC）是一种从复杂样品中分离出细胞外蔬菜（EV）的方法，可以有效地去除高丰富的蛋白质，而经常需要使用多样化的柱子设置，我们旨在在此研究中使用多样化的列。人血清（HS），CL-6B和CL-4B在常规的SEC中表现出较窄的EV和蛋白质峰，较高的分辨率和EV纯度，将其床量增加到20 mL，而Cl-6b和Cl-4b列的分辨率可以显着改善，而CL-6B列则具有较高的峰值级别。进一步建立了一种简化的二分法SEC方法，该方法仅需要两个大量的理由才能在洗脱体中获得1和蛋白质中的EV。这两种方法在去除电动汽车的不同共隔离污染物蛋白方面具有不同的能力，二分法SEC和超速离心可以将EV与人血浆中的EV隔离开来，具有可比的纯度。
Size‐exclusion chromatography (SEC) is a widely adopted method for the isolation of extracellular vesicles (EVs) from complex samples. SEC can efficiently remove high‐abundant proteins, while often requires multiple fractionation operation using diversified column settings. In this study, we aim to establish a simplified SEC method to acquire high quality EVs. In comparison of all three cross‐linked Sepharose resins with the sample types of FBS and human serum (HS), CL‐6B and CL‐4B showed superior performance in regular SEC to CL‐2B in terms of significantly narrower EV and protein peaks, higher resolutions and EV purity. By increasing their bed volumes to 20 ml, the resolutions of CL‐6B and CL‐4B columns could be significantly improved, while the CL‐6B column had the best performance with higher particle yields and tighter EV peaks. With the CL‐6B 20 ml column, we further established a simplified dichotomic SEC method that only requires two bulk elutions to acquire EVs in the Eluate 1 and proteins in the Eluate 2. We further justified that such CL‐6B columns were reusable for at least 10 consecutive times, and the dichotomic SEC was applicable to EV isolations from HS and FBS‐free supernatants of fluorescently labelled and unlabelled SW620 cells. The proteomics analysis implicated that although the two methods had dissimilar abilities in removing different co‐isolating contaminant proteins from EVs, the dichotomic SEC and ultracentrifugation could isolate EVs from human plasma with comparable purity. This dichotomic SEC has its intriguing potential to be used for EV preparation toward clinical testing and/or basic research.
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影响因子：
16
作者：
Mateescu B;Kowal EJ;van Balkom BW;Bartel S;Bhattacharyya SN;Buzás EI;Buck AH;de Candia P;Chow FW;Das S;Driedonks TA;Fernández-Messina L;Haderk F;Hill AF;Jones JC;Van Keuren-Jensen KR;Lai CP;Lässer C;Liegro ID;Lunavat TR;Lorenowicz MJ;Maas SL;Mäger I;Mittelbrunn M;Momma S;Mukherjee K;Nawaz M;Pegtel DM;Pfaffl MW;Schiffelers RM;Tahara H;Théry C;Tosar JP;Wauben MH;Witwer KW;Nolte-'t Hoen EN
通讯作者：
Nolte-'t Hoen EN
DOI：
10.1007/s00018-018-2773-4
发表时间：
2018-08
期刊：
Cellular and molecular life sciences : CMLS
影响因子：
--
作者：
Karimi N;Cvjetkovic A;Jang SC;Crescitelli R;Hosseinpour Feizi MA;Nieuwland R;Lötvall J;Lässer C
通讯作者：
Lässer C
影响因子：
64.5
作者：
Boelens MC;Wu TJ;Nabet BY;Xu B;Qiu Y;Yoon T;Azzam DJ;Twyman-Saint Victor C;Wiemann BZ;Ishwaran H;Ter Brugge PJ;Jonkers J;Slingerland J;Minn AJ
通讯作者：
Minn AJ
DOI：
10.1073/pnas.1521230113
发表时间：
2016-02-23
影响因子：
11.1
作者：
Kowal, Joanna;Arras, Guillaume;Thery, Clotilde
通讯作者：
Thery, Clotilde
影响因子：
16
作者：
Crescitelli R;Lässer C;Szabó TG;Kittel A;Eldh M;Dianzani I;Buzás EI;Lötvall J
通讯作者：
Lötvall J