Name: sample preparation and imaging of exosomes by transmission electron microscopy
Uploaded: 2018-01-04T13:00:00.000+00:00
Duration: 11 min 15 s
Description: Watch this Scientific Journal Video about Sample Preparation and Imaging of Exosomes by Transmission Electron Microscopy at JoVE.com

Bu araştırma Bio tarafından desteklenen &#38; tıbbi teknoloji geliştirme programı Ulusal Araştırma Vakfı (NMG) &#38; Kore hükümeti tarafından finanse (MSIP &#38; MOHW) (No. 2016M3A9B6904244). Biz de eksozom arıtma Medicinal Bioconvergence Araştırma Merkezi üyeleri teşekkür ederiz.

1. blok hazırlık, kesit, boyama ve ya görüntüleme
<ol>
	<li>Santrifüjü 100.000 x g 1,5 saat23de tarafından gelen Kültür süpernatant HCT116 hücre exosomes cips. Kültür süpernatant kaldırmak ve dikkatle 1 mL 0.1 M sodyum cacodylate çözüm (pH 7,0) 1 h 4 ° C'de % 2.5 oxazolidin ile arıtılmış eksozom Pelet düzeltmek 0.1 M sodyum cacodylate için 160 mL distile su (DW) 4,28 g Kakodilik asit geçiyoruz. PH 7.4 0,1 M HCl o zaman ilâ 200 mL distile su ile yapmak için ayarlayın.</li>
	<li>Sabitleştirici kaldırmak ve granül 1 mL 0.1 M sodyum cacodylate arabelleği ile oda sıcaklığında durulayın. 10 dk süren her değişiklikle üç kez tekrarlayın.</li>
	<li>Sonrası 4 ° C'de % 2 Osmiyum tetroxide 1 h için 1 mL örnekleriyle düzeltmek</li>
	<li>Sabitleştirici kaldırmak ve durulama üç kez 0.1 M sodyum cacodylate ile tampon her 10 min.</li>
	<li>Kademeli aseton serisi ile 10 min için kuluçkaya (% 50, % 60, % 70, %80, % 90'ı, % 95, % 100, sırasıyla) shaker üzerinde.</li>
	<li>Aseton kaldırmak ve 3:1 aseton: düşük viskozite gömme karışımı 30 dk için çözüm kuluçkaya. Ya Pelet tüp var.</li>
	<li>Orta kaldırmak ve 1:1 aseton: düşük viskozite gömme karışımı orta ekleyin, sonra 30 dk için kuluçkaya.</li>
	<li>Orta kaldırmak ve 1:3 aseton: düşük viskozite gömme karışımı orta ekleyin, sonra 30 dk için kuluçkaya.</li>
	<li>Orta kaldırın ve % 100 karışımı katıştırma viskozitesi düşük ve gecede oda sıcaklığında kuluçkaya ekleyin.</li>
	<li>Örnek saf düşük viskozite 65 ° C'de 24 h için gömme kalıp ve fırında kullanarak karışımı katıştırma katıştırılmış</li>
	<li>Bir ultra-microtome üzerinden 60 nm kalınlı¤ında bölümleri hazırlayın.</li>
	<li>% 2 uranyl asetat için 20 dk ile çift-leke ve sitrat 10 min için yol. Reynolds götürmek eriyik için fazla 50 mL distile su ile 1.33 g kurşun nitrat ve sodyum sitrat 1.76 g ekleyin.</li>
	<li>Kılavuz 80, transmisyon elektron mikroskobu altında gözlemlemek kV.</li>
	<li>"Al" düğmesini tıklatın ve ardından "Dosya" ve "80, elektron mikroskobu altında CCD kamera sisteminde farklı kaydet" seçeneğini tıklatın kV. Otomatik ayarları pozlama süresi için izleyin.</li>
</ol>
2. IMMUNO-boyama bölümü ve görüntüleme (şekil 1)
<ol>
	<li>60 nm ultrathin bölümleri bir ultra-microtome kullanarak kesme ve nikel ızgara üzerinde toplamak.</li>
	<li>0,02 M glycine için ücretsiz aldehit gruplarını gidermek 10 dk 50 µL damla kılavuzlarda kuluçkaya.</li>
	<li>DW 100 μL içinde üç kez her 1s içinde % 1 BSA içeren PBS için oda sıcaklığında 10 dk. Incubate için durulayın.</li>
	<li>50-100 µL damla KRS antikor24 (1: %100 0,1 BSA içeren PBS içinde) anti kılavuzlarda kuluçkaya 1 h (gerekirse, bu adım 4 ° C'de gecede yapılmalıdır.)</li>
	<li>10 dk % 0,1 BSA içeren PBS, Izgaralar beş ayrı damla (50 µL) ile yıkayın.</li>
	<li>Izgaralar 2nd antikor için 1 h (Anti-tavşan IgG % 0,1 BSA içeren PBS 10 nm altın parçacık (1: 100) Birleşik) damla aktarın.</li>
	<li>Yıkama Izgaralar beş damla % 0,1 BSA içeren PBS (50 µL) her 10 dk ayırın.</li>
	<li>Çift-%2 uranyl asetat için karanlık koşullarda 20 dk ve Reynolds kurşun sitrat 10 dk, sırasıyla leke.</li>
	<li>"Al" düğmesini tıklatın ve ardından "Dosya" ve "80, TEM altında CCD kamera sisteminde farklı kaydet" seçeneğini tıklatın kV. Pozlama süresi otomatik ayarları izledi.</li>
</ol>
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/56482/56482fig1.jpg"> 
Şekil 1: sürecin numune hazırlama ve immunostaining. (A) Çift Kişilik sabit eksozom susuz ve düşük viskozite karışımı reçine katıştırma sızmış. (B) katıştırma reçine. (C) elmas bıçak ile Ultra ince bölümü. IMMUNO-altın etiketleme için (D) kurulum. Izgaralar ultrathin bölümleri ile ıslak kağıt havlu ile yerleştirilen bir parafilm üzerinde sıvı damlacıkları olarak konur. Her bölüm 50-100 µL antikor damlacıkları ile inkübe sonra arabellek ile yıkanır. (E) uranyl asetat ile boyama çift ve sitrat yol. Kapağı heavy metal boyama için alüminyum folyo ile kaplı. (F) çözüm boyama filtre kağıdı tarafından kaldırılır. <a href="//ecsource.jove.com/files/ftp_upload/56482/56482fig1large.jpg" target="_blank">Bu rakam daha büyük bir versiyonunu görüntülemek için buraya tıklayınız.</a>
3. negatif boyama
<ol>
	<li>Parlaklık-ince formvar/karbon film kaplı 200 mesh bakır EM Izgaralar için 1dk kızdırma deşarj tarafından terhis edildi.</li>
	<li>% 2 ile 1 mL saf exosomes saptamak Paraformaldehyde (PFA) 5 min için. Dikkat: Paraformaldehyde dumanı zehirlidir. Tüm iş havalandırılmış duman mahallede yapılmalıdır.</li>
	<li>5-7 µL eksozom süspansiyon çözüm ızgara üzerinde yük ve 1 dk. için kuluçkaya. Ya konsantrasyonu çok yüksek ise, konsantrasyon ile 1/2 - 1/5 oranında seyreltin.</li>
	<li>Hemen filtre uygulanan % 1'uranyl asetat (UA) çözüm EM ızgara yüzeyi şırınga tarafından ~ 20 damla ile leke.</li>
	<li>Aşırı UA çözüm çizgilerini kılavuz kenar filtre kağıdı ile irtibata geçerek kaldırın.</li>
	<li>Hızlı kılavuz bir damla su ile durulayın. Bu adımı aşırı boyama çözüm kaldırır.</li>
	<li>Izgarayı cımbızla tutup masaya koyun ve kılavuz altında oda sıcaklığında 10 dakika kurumaya kısmen kültür yemeği ile kapsar.</li>
	<li>Kılavuz tarafından TEM 80, gelecekteki gözlem için bir EM kılavuz kutusunda saklamak kV.</li>
</ol>
4. bütün dağ Immunostaining için
<ol>
	<li>Parlaklık-deşarj ince formvar/karbon film kaplı 200 mesh bakır EM Izgaralar için 30 s.</li>
	<li>% 2 ile 1 mL saf exosomes saptamak Paraformaldehyde (PFA) 5 min için. Dikkat: Paraformaldehyde dumanı zehirlidir. Tüm iş havalandırılmış duman mahallede yapılmalıdır.</li>
	<li>5-7 µL eksozom çözüm kılavuz üzerinde sabit yük ve 5 dk. için durulama ile 100 µL PBS, üç kez her 10 min için kuluçkaya.</li>
	<li>Izgaralar 0,05 M glycine için ücretsiz aldehit gruplarını gidermek 10 dk 50 µL ile tedavi.</li>
	<li>Izgaralar engelleme arabellek (%1 BSA içeren PBS) atmak için 30 dk aktarın.</li>
	<li>Izgaralar ile 50-100 µL PD-L1 antikor (1: %100 0,1 BSA içeren PBS içinde) anti 1 h (gerekirse, bu adım 4 ° C'de gecede yapılmalıdır) kuluçkaya.</li>
	<li>10 dk % 0,1 BSA içeren PBS, kılavuz beş ayrı damla (50 µL) ile yıkayın.</li>
	<li>Kılavuz için 1 h 2nd antikor bir damla aktarın.</li></ol>9-11, 1: %100 0,1 BSA içeren PBS içinde seyreltilmiş nm altın parçacık için anti-fare IgG Birleşik.
	<li>10 dk % 0,1 BSA içeren PBS, kılavuz beş ayrı damla (50 µL) ile yıkayın. İki ayrı damla (50 µL) kılavuzla DW ile yıkayın. 3.4 3,8 için açıklandığı gibi % 2 uranyl asetat ile negatif boyama gerçekleştirmek.</li>
	<li>Kılavuz tarafından TEM 80, gelecekteki gözlem için bir EM kılavuz kutusunda saklamak kV.</li>

<ol>
	<li>Friend, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Observations+on+cell+lines+derived+from+a+patient+with+Hodgkin's+disease.">Observations on cell lines derived from a patient with Hodgkin's disease.</a> Cancer Res. 38 (8), 2581-2591 (1978).</li><li>Dvorak, H. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tumor+shedding+and+coagulation.">Tumor shedding and coagulation.</a> Science. 212 (4497), 923-924 (1981).</li><li>Zaborowski, M. P., Balaj, L., Breakefield, X. O., Lai, C. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+Vesicles:+Composition,+Biological+Relevance,+and+Methods+of+Study.">Extracellular Vesicles: Composition, Biological Relevance, and Methods of Study.</a> Bioscience. 65 (8), 783-797 (2015).</li><li>Yanez-Mo, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biological+properties+of+extracellular+vesicles+and+their+physiological+functions.">Biological properties of extracellular vesicles and their physiological functions.</a> J Extracell Vesicles. 4, 27066 (2015).</li><li>Wu, Y., Deng, W., Klinke, D. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes:+improved+methods+to+characterize+their+morphology,+RNA+content,+and+surface+protein+biomarkers.">Exosomes: improved methods to characterize their morphology, RNA content, and surface protein biomarkers.</a> Analyst. 140 (19), 6631-6642 (2015).</li><li>Gyorgy, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+vesicles,+current+state-of-the-art:+emerging+role+of+extracellular+vesicles.">Membrane vesicles, current state-of-the-art: emerging role of extracellular vesicles.</a> Cell Mol Life Sci. 68 (16), 2667-2688 (2011).</li><li>Barile, L., Vassalli, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes:+Therapy+delivery+tools+and+biomarkers+of+diseases.">Exosomes: Therapy delivery tools and biomarkers of diseases.</a> Pharmacol Ther. , (2017).</li><li>Vlassov, A. V., Magdaleno, S., Setterquist, R., Conrad, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes:+current+knowledge+of+their+composition,+biological+functions,+and+diagnostic+and+therapeutic+potentials.">Exosomes: current knowledge of their composition, biological functions, and diagnostic and therapeutic potentials.</a> Biochim Biophys Acta. 1820 (7), 940-948 (2012).</li><li>Villarroya-Beltri, C., Baixauli, F., Gutierrez-Vazquez, C., Sanchez-Madrid, F., Mittelbrunn, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sorting+it+out:+regulation+of+exosome+loading.">Sorting it out: regulation of exosome loading.</a> Semin Cancer Biol. 28, 3-13 (2014).</li><li>Thery, C., Ostrowski, M., Segura, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Membrane+vesicles+as+conveyors+of+immune+responses.">Membrane vesicles as conveyors of immune responses.</a> Nat Rev Immunol. 9 (8), 581-593 (2009).</li><li>Camussi, G., Deregibus, M. C., Bruno, S., Cantaluppi, V., Biancone, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes/microvesicles+as+a+mechanism+of+cell-to-cell+communication.">Exosomes/microvesicles as a mechanism of cell-to-cell communication.</a> Kidney Int. 78 (9), 838-848 (2010).</li><li>Zaharie, F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosome-Carried+microRNA-375+Inhibits+Cell+Progression+and+Dissemination+via+Bcl-2+Blocking+in+Colon+Cancer.">Exosome-Carried microRNA-375 Inhibits Cell Progression and Dissemination via Bcl-2 Blocking in Colon Cancer.</a> J Gastrointestin Liver Dis. 24 (4), 435-443 (2015).</li><li>Fuhrmann, G., Neuer, A. L., Herrmann, I. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+vesicles+-+a+promising+avenue+for+the+detection+and+treatment+of+infectious+diseases?.">Extracellular vesicles - a promising avenue for the detection and treatment of infectious diseases?.</a> Eur J Pharm Biopharm. , (2017).</li><li>Lotvall, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Minimal+experimental+requirements+for+definition+of+extracellular+vesicles+and+their+functions:+a+position+statement+from+the+International+Society+for+Extracellular+Vesicles.">Minimal experimental requirements for definition of extracellular vesicles and their functions: a position statement from the International Society for Extracellular Vesicles.</a> J Extracell Vesicles. 3, 26913 (2014).</li><li>Liu, X., Wang, H. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single+particle+electron+microscopy+reconstruction+of+the+exosome+complex+using+the+random+conical+tilt+method.">Single particle electron microscopy reconstruction of the exosome complex using the random conical tilt method.</a> J Vis Exp. (49), (2011).</li><li>Wang, J., Yao, Y., Wu, J., Li, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+and+analysis+of+exosomes+secreted+from+macrophages+extracted+by+different+methods.">Identification and analysis of exosomes secreted from macrophages extracted by different methods.</a> Int J Clin Exp Pathol. 8 (6), 6135-6142 (2015).</li><li>Thery, C., Amigorena, S., Raposo, G., Clayton, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Isolation+and+characterization+of+exosomes+from+cell+culture+supernatants+and+biological+fluids.">Isolation and characterization of exosomes from cell culture supernatants and biological fluids.</a> Curr Protoc Cell Biol. , 22 (2006).</li><li>Mazzeo, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosome+secretion+by+eosinophils:+A+possible+role+in+asthma+pathogenesis.">Exosome secretion by eosinophils: A possible role in asthma pathogenesis.</a> J Allergy Clin Immunol. 135 (6), 1603-1613 (2015).</li><li>Sokolova, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterisation+of+exosomes+derived+from+human+cells+by+nanoparticle+tracking+analysis+and+scanning+electron+microscopy.">Characterisation of exosomes derived from human cells by nanoparticle tracking analysis and scanning electron microscopy.</a> Colloids Surf B Biointerfaces. 87 (1), 146-150 (2011).</li><li>Sharma, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structural-mechanical+characterization+of+nanoparticle+exosomes+in+human+saliva,+using+correlative+AFM,+FESEM,+and+force+spectroscopy.">Structural-mechanical characterization of nanoparticle exosomes in human saliva, using correlative AFM, FESEM, and force spectroscopy.</a> ACS Nano. 4 (4), 1921-1926 (2010).</li><li>Raposo, G., Stoorvogel, W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Extracellular+vesicles:+exosomes,+microvesicles,+and+friends.">Extracellular vesicles: exosomes, microvesicles, and friends.</a> J Cell Biol. 200 (4), 373-383 (2013).</li><li>Bosch, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Trehalose+prevents+aggregation+of+exosomes+and+cryodamage.">Trehalose prevents aggregation of exosomes and cryodamage.</a> Sci Rep. 6, 36162 (2016).</li><li>Lobb, R. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimized+exosome+isolation+protocol+for+cell+culture+supernatant+and+human+plasma.">Optimized exosome isolation protocol for cell culture supernatant and human plasma.</a> J Extracell Vesicles. 4, 27031 (2015).</li><li>Kim, S. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Caspase-8+controls+the+secretion+of+inflammatory+lysyl-tRNA+synthetase+in+exosomes+from+cancer+cells.">Caspase-8 controls the secretion of inflammatory lysyl-tRNA synthetase in exosomes from cancer cells.</a> J Cell Biol. , (2017).</li><li>Conde-Vancells, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+and+comprehensive+proteome+profiling+of+exosomes+secreted+by+hepatocytes.">Characterization and comprehensive proteome profiling of exosomes secreted by hepatocytes.</a> J Proteome Res. 7 (12), 5157-5166 (2008).</li><li>Kadiu, I., Narayanasamy, P., Dash, P. K., Zhang, W., Gendelman, H. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biochemical+and+biologic+characterization+of+exosomes+and+microvesicles+as+facilitators+of+HIV-1+infection+in+macrophages.">Biochemical and biologic characterization of exosomes and microvesicles as facilitators of HIV-1 infection in macrophages.</a> J Immunol. 189 (2), 744-754 (2012).</li><li>Gong, J., Korner, R., Gaitanos, L., Klein, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosomes+mediate+cell+contact-independent+ephrin-Eph+signaling+during+axon+guidance.">Exosomes mediate cell contact-independent ephrin-Eph signaling during axon guidance.</a> J Cell Biol. 214 (1), 35-44 (2016).</li><li>Melo, S. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glypican-1+identifies+cancer+exosomes+and+detects+early+pancreatic+cancer.">Glypican-1 identifies cancer exosomes and detects early pancreatic cancer.</a> Nature. 523 (7559), 177-182 (2015).</li><li>Valadi, H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exosome-mediated+transfer+of+mRNAs+and+microRNAs+is+a+novel+mechanism+of+genetic+exchange+between+cells.">Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells.</a> Nat Cell Biol. 9 (6), 654-659 (2007).</li><li>van Niel, G., Heyman, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+epithelial+cell+cytoskeleton+and+intracellular+trafficking.+II.+Intestinal+epithelial+cell+exosomes:+perspectives+on+their+structure+and+function.">The epithelial cell cytoskeleton and intracellular trafficking. II. Intestinal epithelial cell exosomes: perspectives on their structure and function.</a> Am J Physiol Gastrointest Liver Physiol. 283 (2), G251-G255 (2002).</li></ol>

Yazarlar ifşa gerek yok.

Bu makalede ayrıntılı eksozom'ın yapısı gözlemleyerek ve onun belirli proteinlerin etiketleme için bir protokol sunar. Negatif boyama eksozom17görüntüleme için en iyi yöntem olarak kabul edilmiştir. Bu geleneksel teknik exosomes Kupası şeklindeki yapı göstermiştir. Ancak, bu fincan şekli bir kurutma işlemi nedeniyle oluşan yapay doku biçimidir. Cryo-TEM sonuçları exosomes sulu çözüm25,26yılında tamamen kürese...

Ekstrasellüler veziküller (EVs) hücreleri tarafından salgılanan lipid bilayer veziküller vardır ve büyüklükleri 30 ve 300 nm arasında değişmektedir. EVs veziküller Hodgkin Hastaligi1hastalarda kanıt ile 1978 yılında ilk bildirildi. Bu veziküller 40-120 nanometre boyutunda bulunduğu rapor edilmiştir. 1980 yılında, bu EVs pıhtılaşma sistemi ve tromboz, bir damar kanseri hastalar2içinde bir kan pıhtısı oluşumu katılmaktadırlar bildirildi. 30 yıl sonra EVs tümör işgali, bağışıklık kaçış ve anjiogenezi3tanıtmak için önemli bir faktör olarak bildirilmiştir. Buna ek olarak, EVs fonksiyonları alanlarda hücresel etkileşimlerin iltihabı, bağışıklık bozukluğu, nörolojik hastalık ve kanser4düzenleyiciler olarak incelenmiştir. EVs protein, mRNA ve mikroRNA5gibi belirli biomolecules içerdiğinden, teşhis ve tedavi uygulama onların potansiyel analiz6,7oldu. EVs exosomes, prostasomes, oncosomes, dexosomes, microparticles, promininosomes, argosomes ve hücresel kökeni ve biyolojik fonksiyon3bağlı eksozom benzeri veziküller dahil olmak üzere alt gruplar oluşan bir kategori vardır. Buna ek olarak, onların Biyogenez üzerinde bağlı olarak, bu EVs microvesicles (döken microvesicles, 100-1000 nm) ve exosomes (30-300 nm)8,9bölünmüş olabilir. Bunlar arasında ya bir hücre iletişim bağışıklık yanıtı10, kanser11,12ve bulaşıcı hastalık13olarak bildirilmiştir.
Exosomes olarak bir biyomarker erken tanı için ilgi hızla artıyor ve arıtma ve exosomes karakterizasyonu moleküler görüntüleme teknikleri ile eşlik etmelidir. Farklı boyutlarda ve türleri exosomes, kendi kökeni ve işlev14, bağlı olarak, Morfoloji mikroskobu teknikleri gibi elektron mikroskobu yüksek çözünürlüklü tarafından ayrılır. Çoğu exosomes negatif lekeli transmisyon elektron mikroskobu (TEM)15,16,17tarafından görüntülenmiştir ve bu sonuçlar immunolabeling bütün Dağı vezikül içinde belirli bir protein tarafından teyit edildi 18. birkaç araştırma grubu aracılığıyla tarama elektron mikroskobu, Atomik kuvvet mikroskobu19,20ve Cryo-TEM21,22yapısı bildirdin. Ancak, bu teknikler eksozom yapısı eğitim için yararlı olsa da, ya içinde bulunan spesifik proteinlerin konumunu gözlemlemek için yetersiz bunlar. Bu nedenle, belirli protein etiketleme ile exosomes görüntüleme için bir protokol tanıttı. Biz blok hazırlanması, ultrathin kesit ve immunostaining protein'ın detaylı eksozom konumda ascertaining için uygulanır. Bu negatif boyama ve geleneksel eksozom karakterizasyonu için kullanılan bütün Dağı immunostaining ile karşılaştırıldı.

<table><tbody><tr><td>Glutaraldehyde</td><td>EMS</td><td>16200</td><td></td></tr><tr><td>Sodium cacodylate&amp;nbsp;</td><td>EMS</td><td>12300</td><td></td></tr><tr><td>Osmium tetroxide 1 g crystal</td><td>EMS</td><td>19100</td><td>Use only in fume hood</td></tr><tr><td>acetone</td><td>JUNSEI</td><td>1B2031</td><td></td></tr><tr><td>Spurr medium</td><td>TED PELLA</td><td>18108</td><td></td></tr><tr><td>Ultra-microtome</td><td>Leica</td><td>UCT</td><td></td></tr><tr><td>Uranyl acetate</td><td>EMS</td><td>22400</td><td>Hazardous chemical</td></tr><tr><td>Lead citrate</td><td>EMS</td><td>17900</td><td></td></tr><tr><td>Transmission Electron Microscopy</td><td>Hitachi</td><td>H7600</td><td></td></tr><tr><td>nickel grid</td><td>EMS</td><td>G200-Ni</td><td></td></tr><tr><td>Copper grid</td><td>EMS</td><td>G200-Cu</td><td></td></tr><tr><td>glycine</td><td>SIGMA</td><td>022K5404</td><td></td></tr><tr><td>Phosphate buffer saline</td><td>SIGMA</td><td>P4417</td><td></td></tr><tr><td>Bovine serum albumin</td><td>Aurion</td><td>25557</td><td></td></tr><tr><td>1st Antibody</td><td></td><td></td><td>purification in manually&amp;nbsp;</td></tr><tr><td>Purified anti-human CD274 (B7-H1, PD-L1) Antibody</td><td>BioLegend</td><td>329710</td><td>Whole mount Immumogold</td></tr><tr><td>Purified Mouse IgG2b, &amp;kappa; Isotype Ctrl</td><td>BioLegend</td><td>401202</td><td>Whole mount Immumogold (Control)</td></tr><tr><td>2nd Anti-mouse igG conjugated 9-11 nm gold particle</td><td>sigma</td><td>G7652</td><td></td></tr><tr><td>Transmission Electron Microscopy</td><td>JEOL</td><td>JEM2200FS</td><td></td></tr><tr><td>Glow discharger</td><td>JEOL</td><td>JFC1100E</td><td></td></tr><tr><td>Carbon grid</td><td>EMS</td><td>121119</td><td></td></tr><tr><td>Paraformaldehyde</td><td>EMS</td><td>19210</td><td></td></tr><tr><td>Formvar carbon coated Copper Grid (200 meh)</td><td>EMS</td><td>FCF200-CU</td><td></td></tr><tr><td>Formvar carbon coated&amp;nbsp; Nickel Grid (200 mesh)</td><td>EMS</td><td>FCF200-NI</td><td></td></tr></tbody></table>

sample preparation and imaging of exosomes by transmission electron microscopy

Exosomes hücre dışı veziküller nano boyutlu vücut sıvıları tarafından salgılanan vardır ve onları salgılar hücre özellikleri temsil etmek için bilinir. İçeriği ve salgılanan veziküller morfolojisi hücre davranış ya da fizyolojik durum, örneğin hücre büyümesi, geçiş, bölünme ve ölüm yansıtır. Exosomes rolü son derece boyutuna bağlı olabilir ve exosomes boyutu 30 ila 300'e değişir nm. En çok kullanılan eksozom görüntüleme için diğer sonuçlar Cryo-transmisyon elektron mikroskobu, elektron mikroskobu tarama ve Atomik kuvvet mikroskobu dayalı iken negatif boyama yöntemidir. Tipik eksozom'ın Morfoloji negatif boyama ile değerlendirildi, bir fincan şekli olmakla birlikte, daha fazla ayrıntılı bilgi henüz net değildir. Bir eksozom yapısal çalışma ile iyi karakterize gerekli özellikle de sağlık ve ilaç alanlar var. Bu nedenle, işlev bağımlı Morfoloji eksozom detaylı yapısında belirli bir protein etiketleme gibi elektron mikroskobu teknikleri tarafından doğrulanması gerekir. Ayrıntılı yapısı gözlemlemek için ultrathin kesitli görüntülerini ve exosomes negatif lekeli görüntüleri karşılaştırıldı. Bu protokol için iletim elektron mikroskopi exosomes negatif boyama, Bütün Dağı IMMUNO-boyama, blok hazırlık, ince bölüm ve IMMUNO-altın etiketleme gibi görüntüleme için öneririz.

Şu anda, exosomes transmisyon elektron mikroskobu tarafından boyutu ve şekli kategoride sınıflandırılır. Şekil 2 negatif lekeli eksozom ve bağışıklık etiketli eksozom tüm bağlama durumu gösterir. Şekil 3 kesitli eksozom ve immün etiketli exosomes ince kesit sonra gösterir. IMMUNO-altın belirli proteinlerin antikorları kullanarak boyama olumlu bir eksozom tanımlamak ve proteinler arasında eksozom türleri s...

Watch this Scientific Journal Video about Sample Preparation and Imaging of Exosomes by Transmission Electron Microscopy at JoVE.com

Sample Preparation and Imaging of Exosomes by Transmission Electron Microscopy

Numune hazırlama ve transmisyon elektron mikroskobu tarafından Exosomes görüntüleme

Bu iletişim kuralı gerekli transmisyon elektron mikroskobu negatif boyama, ayrıntılı yapısı ve IMMUNO-altın spesifik proteinlerin konumlarda exosomes belirlemek için etiketleme için ultrathin kesit dahil olmak üzere için çeşitli teknikler açıklanır.

Exosomes are nano-sized extracellular vesicles secreted by body fluids and are known to represent the characteristics of cells that secrete them. The ...

sample-preparation-imaging-exosomes-transmission-electron

Universidad de Cartagena UDEC

Research

JoVE Journal

Biology

38.8K Görüntüleme.  Asan Medical Center. Bu iletişim kuralı gerekli transmisyon elektron mikroskobu negatif boyama, ayrıntılı yapısı ve IMMUNO-altın spesifik proteinlerin konumlarda exosomes belirlemek için etiketleme için ultrathin kesit dahil olmak üzere için çeşitli teknikler açıklanır.

Video: Sample Preparation and Imaging of Exosomes by Transmission Electron Microscopy

Imaging of Extracellular Vesicles by Atomic Force Microscopy

Exosomes and other extracellular vesicles (EVs) are molecular complexes consisting of a lipid membrane vesicle, its surface decoration by membrane proteins and other molecules, and diverse luminal content inherited from a parent cell, which includes RNAs, proteins, and DNAs. The characterization of the hydrodynamic sizes of EVs, which depends on the size of the vesicle and its coronal layer formed by surface decorations, has become routine. For exosomes, the smallest of EVs, the relative difference between the hydrodynamic and vesicles sizes is significant. The characterization of vesicles sizes by the cryogenic transmission electron microscopy (cryo-TEM) imaging, a gold standard technique, remains a challenge due to the cost of the instrument, the expertise required to perform the sample preparation, imaging and data analysis, and a small number of particles often observed in images. A widely available and accessible alternative is the atomic force microscopy (AFM), which can produce versatile data on three-dimensional geometry, size, and other biophysical properties of extracellular vesicles. The developed protocol guides the users in utilizing this analytical tool and outlines the workflow for the analysis of EVs by the AFM, which includes the sample preparation for imaging EVs in hydrated or desiccated form, the electrostatic immobilization of vesicles on a substrate, data acquisition, its analysis, and interpretation. The representative results demonstrate that the fixation of EVs on the modified mica surface is predictable, customizable, and allows the user to obtain sizing results for a large number of vesicles. The vesicle sizing based on the AFM data was found to be consistent with the cryo-TEM imaging.&#160;

A step-by-step procedure is described for label-free immobilization of exosomes and extracellular vesicles from liquid samples and their imaging by atomic force microscopy (AFM). The AFM images are used to estimate the size of the vesicles in the solution and characterize other biophysical properties.&#160;

Atomik Kuvvet Mikroskobu ile Hücre Dışı Veziküllerin Görüntülenmesi

Exosomes and other extracellular vesicles (EVs) are molecular complexes consisting of a lipid membrane vesicle, its surface decoration by membrane ...

Biyokimya

Single Particle Electron Microscopy Reconstruction of the Exosome Complex Using the Random Conical Tilt Method

Single particle electron microscopy (EM) reconstruction has recently become a popular tool to get the three-dimensional (3D) structure of large macromolecular complexes. Compared to X-ray crystallography, it has some unique advantages. First, single particle EM reconstruction does not need to crystallize the protein sample, which is the bottleneck in X-ray crystallography, especially for large macromolecular complexes. Secondly, it does not need large amounts of protein samples. Compared with milligrams of proteins necessary for crystallization, single particle EM reconstruction only needs several micro-liters of protein solution at nano-molar concentrations, using the negative staining EM method. However, despite a few macromolecular assemblies with high symmetry, single particle EM is limited at relatively low resolution (lower than 1 nm resolution) for many specimens especially those without symmetry. This technique is also limited by the size of the molecules under study, i.e. 100 kDa for negatively stained specimens and 300 kDa for frozen-hydrated specimens in general.
For a new sample of unknown structure, we generally use a heavy metal solution to embed the molecules by negative staining. The specimen is then examined in a transmission electron microscope to take two-dimensional (2D) micrographs of the molecules. Ideally, the protein molecules have a homogeneous 3D structure but exhibit different orientations in the micrographs. These micrographs are digitized and processed in computers as "single particles". Using two-dimensional alignment and classification techniques, homogenous molecules in the same views are clustered into classes. Their averages enhance the signal of the molecule's 2D shapes. After we assign the particles with the proper relative orientation (Euler angles), we will be able to reconstruct the 2D particle images into a 3D virtual volume.
In single particle 3D reconstruction, an essential step is to correctly assign the proper orientation of each single particle. There are several methods to assign the view for each particle, including the angular reconstitution1 and random conical tilt (RCT) method2. In this protocol, we describe our practice in getting the 3D reconstruction of yeast exosome complex using negative staining EM and RCT. It should be noted that our protocol of electron microscopy and image processing follows the basic principle of RCT but is not the only way to perform the method. We first describe how to embed the protein sample into a layer of Uranyl-Formate with a thickness comparable to the protein size, using a holey carbon grid covered with a layer of continuous thin carbon film. Then the specimen is inserted into a transmission electron microscope to collect untilted (0-degree) and tilted (55-degree) pairs of micrographs that will be used later for processing and obtaining an initial 3D model of the yeast exosome. To this end, we perform RCT and then refine the initial 3D model by using the projection matching refinement method3.

This article describes a standard method to get a three-dimensional (3D) reconstruction of biological macromolecules using negative staining electron microscopy (EM). In this protocol, we explain how to get the 3D structure of the Saccharomyces cerevisiae exosome complex at medium resolution using the random conical tilt reconstruction method (RCT).

Rastgele Konik Tilt Yöntemiyle Exosome Kompleksi Tek Parçacık Elektron Mikroskobu İmar

Single particle electron microscopy (EM) reconstruction has recently become a popular tool to get the three-dimensional (3D) structure of large ...

Biyoloji

Using Nanoplasmon-Enhanced Scattering and Low-Magnification Microscope Imaging to Quantify Tumor-Derived Exosomes

Infected or malignant cells frequently secrete more exosomes, leading to elevated levels of disease-associated exosomes in the circulation. These exosomes have the potential to serve as biomarkers for disease diagnosis and to monitor disease progression and treatment response. However, most exosome analysis procedures require exosome isolation and purification steps, which are usually time-consuming and labor-intensive, and thus of limited utility in clinical settings. This report describes a rapid procedure to analyze specific biomarkers on the outer membrane of exosomes without requiring separate isolation and purification steps. In this method, exosomes are captured on the surface of a slide by exosome-specific antibodies and then hybridized with nanoparticle-conjugated antibody probes specific to a disease. After hybridization, the abundance of the target exosome population is determined by analyzing low-magnification dark-field microscope (LMDFM) images of the bound nanoparticles. This approach can be easily adopted for research and clinical use to analyze membrane-associated exosome biomarkers linked to disease.

Clinical translation of exosome-derived biomarkers for diseased and malignant cells is hindered by the lack of rapid and accurate quantification methods. This report describes the use of low-magnification dark-field microscope images to quantify specific exosome subtypes in small volume serum or plasma samples.

Tümör türevi Exosomes ölçmek için Nanoplasmon-gelişmiş saçılma ve düşük büyütme mikroskop görüntüleme kullanma

Infected or malignant cells frequently secrete more exosomes, leading to elevated levels of disease-associated exosomes in the circulation. These exosomes ...

Biyomühendislik

Isolation and Profiling of MicroRNA-containing Exosomes from Human Bile

Exosome research in the last three years has greatly extended the scope towards identification and characterization of biomarkers and their therapeutic uses.
Exosomes have recently been shown to contain microRNAs (miRs). MiRs themselves have arisen as valuable biomarkers for diagnostic purposes. As specimen collection in clinics and hospitals is quite variable, miRNA isolation from whole bile varies substantially. To achieve robust, accurate and reproducible miRNA profiles from collected bile samples in a simple manner required the development of a high-quality protocol to isolate and characterize exosomes from bile. The method requires several centrifugations and a filtration step with a final ultracentrifugation step to pellet the isolated exosomes. Electron microscopy, Western blots, flow cytometry and multi-parameter nanoparticle optical analysis, where available, are crucial characterization steps to validate the quality of the exosomes. For the isolation of miRNA from these exosomes, spiking the lysate with a non-specific, synthetic miRNA from a species like Caenorhabditis elegans, i.e., Cel-miR-39, is important for normalization of RNA extraction efficiency. The isolation of exosome from bile fluid following this method allows the successful miRNA profiling from bile samples stored for several years at -80 &#176;C.

Bile fluid is a valuable source of extracellular vesicles/exosomes that contain potentially important biomarkers. This protocol represents a robust method to isolate exosomes from human bile for further analyses including miRNA profiling.

İzolasyon ve profil MicroRNA içeren İnsan Bile dan ekzozomlar

Exosome research in the last three years has greatly extended the scope towards identification and characterization of biomarkers and their therapeutic ...

Tıp

Isolation and Characterization of RNA-Containing Exosomes

The field of exosome research is rapidly expanding, with a dramatic increase in publications in recent years. These small vesicles (30-100 nm) of endocytic origin were first proposed to function as a way for reticulocytes to eradicate the transferrin receptor while maturing into erythrocytes1, and were later named exosomes. Exosomes are formed by inward budding of late endosomes, producing multivesicular bodies (MVBs), and are released into the environment by fusion of the MVBs with the plasma membrane2. Since the first discovery of exosomes, a wide range of cells have been shown to release these vesicles. Exosomes have also been detected in several biological fluids, including plasma, nasal lavage fluid, saliva and breast milk3-6. Furthermore, it has been demonstrated that the content and function of exosomes depends on the originating cell and the conditions under which they are produced. A variety of functions have been demonstrated for exosomes, such as induction of tolerance against allergen7,8, eradication of established tumors in mice9, inhibition and activation of natural killer cells10-12, promotion of differentiation into T regulatory cells13, stimulation of T cell proliferation14 and induction of T cell apoptosis15. Year 2007 we demonstrated that exosomes released from mast cells contain messenger RNA (mRNA) and microRNA (miRNA), and that the RNA can be shuttled from one cell to another via exosomes. In the recipient cells, the mRNA shuttled by exosomes was shown to be translated into protein, suggesting a regulatory function of the transferred RNA16. Further, we have also shown that exosomes derived from cells grown under oxidative stress can induce tolerance against further stress in recipient cells and thus suggest a biological function of the exosomal shuttle RNA17. Cell culture media and biological fluids contain a mixture of vesicles and shed fragments. A high quality isolation method for exosomes, followed by characterization and identification of the exosomes and their content, is therefore crucial to distinguish exosomes from other vesicles and particles. Here, we present a method for the isolation of exosomes from both cell culture medium and body fluids. This isolation method is based on repeated centrifugation and filtration steps, followed by a final ultracentrifugation step in which the exosomes are pelleted. Important methods to identify the exosomes and characterize the exosomal morphology and protein content are highlighted, including electron microscopy, flow cytometry and Western blot. The purification of the total exosomal RNA is based on spin column chromatography and the exosomal RNA yield and size distribution is analyzed using a Bioanalyzer.

This paper demonstrates methods for the isolation, purification and detection of exosomes, as well as techniques for analysis of their molecular content. These methods are adaptable for exosome isolation from both cell culture media and biological fluids, and can beyond analysis of molecular content also be useful in functional studies.

RNA İçeren Exosomes İzolasyonu ve Karakterizasyonu

The field of exosome research is rapidly expanding, with a dramatic increase in publications in recent years. These small vesicles (30-100 nm) of ...

An Innovative Method for Exosome Quantification and Size Measurement

Although the biological importance of exosomes has recently gained an increasing amount of scientific and clinical attention, much is still unknown about their complex pathways, their bioavailability and their diverse functions in health and disease. Current work focuses on the presence and the behavior of exosomes (in vitro as well as in vivo) in the context of different human disorders, especially in the fields of oncology, gynecology and cardiology.
Unfortunately, neither a consensus regarding a gold standard for exosome isolation exists, nor is there an agreement on such a method for their quantitative analysis. As there are many methods for the purification of exosomes and also many possibilities for their quantitative and qualitative analysis, it is difficult to determine a combination of methods for the ideal approach. 
Here, we demonstrate nanoparticle tracking analysis (NTA), a semi-automated method for the characterization of exosomes after isolation from human plasma by ultracentrifugation. The presented results show that this approach for isolation, as well as the determination of the average number and size of exosomes, delivers reproducible and valid data, as confirmed by other methods, such as scanning electron microscopy (SEM).

A method for isolation of exosomes from whole blood and further analysis by nanoparticle tracking using a semi-automatic instrument is presented in this article. The presented technology provides an extremely sensitive method for visualizing and analyzing particles in liquid suspension.

Exosome Kantifikasyon ve Boyut Ölçüm Yenilikçi Yöntem

Although the biological importance of exosomes has recently gained an increasing amount of scientific and clinical attention, much is still unknown about ...

Biyomedikal Uygulamalarda Eksozomların Sürekli Verilmesi için Mikroiğne Yamaları

Fabrication and Characterization of Microneedle Patches for Loading and Delivery of Exosomes

Exosomes, as emerging &#34;next-generation&#34; biotherapeutics and drug delivery vectors, hold immense potential in diverse biomedical fields, ranging from drug delivery and regenerative medicine to disease diagnosis and tumor immunotherapy. However, the rapid clearance by traditional bolus injection and poor stability of exosomes restrict their clinical application. Microneedles serve as a solution that prolongs the residence time of exosomes at the administration site, thereby maintaining the drug concentration and facilitating sustained therapeutic effects. In addition, microneedles also possess the ability to maintain the stability of bioactive substances. Therefore, we introduce a microneedle patch for loading and delivering exosomes and share the methods, including isolation of exosomes, fabrication, and characterization of exosome-loaded microneedle patches. The microneedle patches were fabricated using trehalose and hyaluronic acid as the tip materials and polyvinylpyrrolidone as the backing material through a two-step casting method. The microneedles demonstrated robust mechanical strength, with tips able to withstand 2 N. Pig skin was used to simulate human skin, and the tips of microneedles completely melted within 60 s after skin puncture. The exosomes released from the microneedles exhibited morphology, particle size, marker proteins, and biological functions comparable to those of fresh exosomes, enabling dendritic cells uptake and promoting their maturation.

Yazar Spotlight: Gelişmiş Eksozom İletimi ve Stabilitesi için Yenilikçi Mikroiğne Tabanlı Stratejiler

Exosomes possess significant clinical potential, but their practical application is limited due to easy in vivo clearance and poor stability. Microneedles present a solution by enabling localized delivery by puncturing physiological barriers and dry-state preservation, thereby addressing the limitations of exosome administration and expanding their clinical utility.

Eksozomların yüklenmesi ve teslimi için mikroiğne yamalarının üretimi ve karakterizasyonu

Exosomes, as emerging &#34;next-generation&#34; biotherapeutics and drug delivery vectors, hold immense potential in diverse biomedical fields, ranging ...

Large-Scale Preparation of Synovial Fluid Mesenchymal Stem Cell-Derived Exosomes by 3D Bioreactor Culture

Exosomes secreted by mesenchymal stem cells (MSCs) have been suggested as promising candidates for cartilage injuries and osteoarthritis treatment. Exosomes for clinical application require large-scale production. To this aim, human synovial fluid MSCs (hSF-MSCs) were grown on microcarrier beads, and then cultured in a dynamic three-dimension (3D) culture system. Through utilizing 3D dynamic culture, this protocol successfully obtained large-scale exosomes from SF-MSC culture supernatants. Exosomes were harvested by ultracentrifugation and verified by a transmission electron microscope, nanoparticle transmission assay, and western blotting. Also, the microbiological safety of exosomes was detected. Results of exosome detection suggest that this approach can produce a large number of good manufacturing practices (GMP)-grade exosomes. These exosomes could be utilized in exosome biology research and clinical osteoarthritis treatment.

Here, we present a protocol to produce a large number of GMP-grade exosomes from synovial fluid mesenchymal stem cells using a 3D bioreactor.

Sinovyal Sıvı Mezenkimal Kök Hücre Kaynaklı Eksozomların 3D Biyoreaktör Kültürü ile Büyük Ölçekli Hazırlanması

Exosomes secreted by mesenchymal stem cells (MSCs) have been suggested as promising candidates for cartilage injuries and osteoarthritis treatment. ...

Numune hazırlama ve transmisyon elektron mikroskobu tarafından Exosomes görüntüleme

Özet

Daha Fazla Video Keşfet

Bu videodaki bölümler

Atomik Kuvvet Mikroskobu ile Hücre Dışı Veziküllerin Görüntülenmesi

Rastgele Konik Tilt Yöntemiyle Exosome Kompleksi Tek Parçacık Elektron Mikroskobu İmar

Tümör türevi Exosomes ölçmek için Nanoplasmon-gelişmiş saçılma ve düşük büyütme mikroskop görüntüleme kullanma

İzolasyon ve profil MicroRNA içeren İnsan Bile dan ekzozomlar

RNA İçeren Exosomes İzolasyonu ve Karakterizasyonu

Exosome Kantifikasyon ve Boyut Ölçüm Yenilikçi Yöntem

Eksozomların yüklenmesi ve teslimi için mikroiğne yamalarının üretimi ve karakterizasyonu

Eksozomların yüklenmesi ve teslimi için mikroiğne yamalarının üretimi ve karakterizasyonu

Eksozomların yüklenmesi ve teslimi için mikroiğne yamalarının üretimi ve karakterizasyonu

Sinovyal Sıvı Mezenkimal Kök Hücre Kaynaklı Eksozomların 3D Biyoreaktör Kültürü ile Büyük Ölçekli Hazırlanması