A size-exclusion-based approach for purifying extracellular vesicles from human plasma
Open accessAbstract
Extracellular vesicles (EVs) are released into blood from multiple organs and carry molecular cargo that facilitates inter-organ communication and an integrated response to physiological and pathological stimuli. Interrogation of the protein cargo of EVs is currently limited by the absence of optimal and reproducible approaches for purifying plasma EVs that are suitable for downstream proteomic analyses. We describe a size-exclusion chromatography (SEC)-based method to purify EVs from platelet-poor plasma (PPP) for proteomics profiling via high-resolution mass spectrometry (SEC-MS). The SEC-MS method identifies more proteins with higher precision than several conventional EV isolation approaches. We apply the SEC-MS method to identify the unique proteomic signatures of EVs released from platelets, adipocytes, muscle cells, and hepatocytes, with the goal of identifying tissue-specific EV markers. Furthermore, we apply the SEC-MS approach to evaluate the effects of a single bout of exercise on EV proteomic cargo in human plasma.
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A 2021 optimized size-exclusion chromatography method for proteomic analysis of plasma-derived extracellular vesicles from platelet-poor plasma has greater precision than conventional techniques and shows a distinct exosome protein cargo profile after acute exercise in humans.
“A 2021 paper presented an optimized size-exclusion chromatography method for proteomic analysis of plasma-derived extracellular vesicles from platelet-poor plasma; this technique has greater precision than conventional extracellular vesicle techniques and demonstrates a distinct exosome protein cargo profile after acute exercise in humans37.”
From Exerkines in health, resilience and disease (2022) · cites this paper as [37]
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Tissue-based map of the human proteome
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Shedding light on the cell biology of extracellular vesicles
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MethodInfluentialBiogenesis, Secretion, and Intercellular Interactions of Exosomes and Other Extracellular Vesicles
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“It has been shown that exercise-induced muscle tissue damage attracts many circulating cells, including those involved in immunological reactions that remove degraded proteins and stimulate protein synthesis (Pedersen and Hoffman-Goetz, 2000).”
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Keerthikumar, S., Chisanga, D., Ariyaratne, D., Saffar, H. A., Anand, S., Zhao, K., Samuel, M., Pathan, M., Jois, M., Chilamkurti, N., et al. (2015) · Journal of Molecular Biology · doi:10.1016/j.jmb.2015.09.019 · Cited by 1,447 · cited 1× · see citing works
“Toward this goal, we established a reference elution range for EVs by using the ExoCarta top 100 (ECT100), a list of the most commonly observed proteins in exosomes (Keerthikumar et al., 2016).”
MethodSingle‐step isolation of extracellular vesicles by size‐exclusion chromatography
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Extracellular Vesicles in Cancer: Exosomes, Microvesicles and the Emerging Role of Large Oncosomes
Minciacchi, V. R., Freeman, M., & Vizio, D. D. (2015) · Seminars in Cell and Developmental Biology · doi:10.1016/j.semcdb.2015.02.010 · Cited by 900 · cited 1× · see citing works
“EVs are membranous structures released from most cells that serve as vehicles to exchange protein, nucleic acid, and lipid cargo (Korutla et al., 2019; Minciacchi et al., 2015; van Niel et al., 2018) between distant organs.”
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“This is suggestive of suboptimal EV recovery with UTC, which was noted by others (Livshits et al., 2015).”
BackgroundExtracellular Vesicles Provide a Means for Tissue Crosstalk during Exercise
Whitham, M., Parker, B. L., Friedrichsen, M. H., Hingst, J. R., Hjorth, M., Hughes, W. E., Egan, C. L., Cron, L., Watt, K. I., Kuchel, R. P., et al. (2018) · Cell Metabolism · doi:10.1016/j.cmet.2017.12.001 · Cited by 689 · cited 4× · see citing works
“We identified 3,409 proteins in this experiment by using a short 1-h MS method, substantially shorter than UTC-MS methods described previously (Whitham et al., 2018).”
BackgroundMethodInfluentialDetailed analysis of the plasma extracellular vesicle proteome after separation from lipoproteins
Karimi, N., Cvjetkovic, A., Jang, S. C., Crescitelli, R., Feizi, M. A. H., Nieuwland, R., Lötvall, J. O., & Lässer, C. (2018) · Cellular and Molecular Life Sciences · doi:10.1007/s00018-018-2773-4 · Cited by 591 · cited 2× · see citing works
“This separation enables improved proteomic characterization of EVs (Jeannin et al., 2018; Karimi et al., 2018), largely by increasing isolation yield and reducing plasma protein contamination (Stranska et al.”
BackgroundTabb, D. L., Vega-Montoto, L., Rudnick, P. A., Variyath, A. M., Ham, A.-J. L., Bunk, D. M., Kilpatrick, L. E., Billheimer, D. D., Blackman, R. K., Cardasis, H. L., et al. (2010) · Journal of Proteome Research · doi:10.1021/pr9006365 · Cited by 551 · cited 1× · see citing works
“54; Table S1), which is within the acceptable limits of typical label-free proteomics experiments (Al Shweiki et al., 2017; Shen et al., 2018; Tabb et al., 2010).”
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“Sixteen additional individuals performed a single bout of unilateral knee extensor resistance exercise consisting of three sets of ten repetitions at 70% of their 1-repetitionmaximum (1RM) (Robinson et al., 2017).”
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“Although this step is strongly recommended and widely recognized by the EV research community, several recent studies examining the plasma EV proteome have utilized frozen serum or plasma without platelet depletion (Chen et al., 2017; Jeannin et al., 2018; Whitham et al., 2018; Zhang et al., 2018), which facilitates a…”
BackgroundProteomic Analysis of Exosomes Isolated from Human Malignant Pleural Effusions
Bard, M. P., Hegmans, J. P., Hemmes, A., Luider, T. M., Willemsen, R., Severijnen, L.-A. A., van Meerbeeck, J. P., Burgers, S. A., Hoogsteden, H. C., & Lambrecht, B. N. (2004) · American Journal of Respiratory Cell and Molecular Biology · doi:10.1165/rcmb.2003-0238oc · Cited by 416 · cited 1× · see citing works
“For example, proteomic characterization of plasma-derived EVs isolated with differential ultracentrifugation (UTC) has been confounded by the co-isolation of an overwhelming number of plasma protein contaminants (Bard et al., 2004; György et al., 2011).”
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“For example, proteomic characterization of plasma-derived EVs isolated with differential ultracentrifugation (UTC) has been confounded by the co-isolation of an overwhelming number of plasma protein contaminants (Bard et al., 2004; György et al., 2011).”
BackgroundLacroix, R., Judicone, C., PONCELET, P., Robert, S., ARNAUD, L., Sampol, J., & Dignat‐George, F. (2011) · Journal of Thrombosis and Haemostasis · doi:10.1111/j.1538-7836.2011.04610.x · Cited by 401 · cited 2× · see citing works
“We also demonstrate the importance of following the rigorous pre-analytical guidelines established by the ISEV and ISTH to limit ex vivo generation of platelet EVs (Artoni et al., 2012; Ayers et al., 2011; Burnouf et al., 2014; LaCroix et al., 2012).”
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“, 2018), largely by increasing isolation yield and reducing plasma protein contamination (Stranska et al., 2018).”
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Physical exercise induces rapid release of small extracellular vesicles into the circulation
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“The effects of exercise on EV release into circulation has been investigated by others (Frühbeis et al., 2015; Whitham et al., 2018), and we considered acute exercise as an appropriate physiological intervention to determine the ability of the approach to detect the EV proteome changes.”
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“Platelet EVs were prepared as previously described (Prokopi et al., 2009).”
MethodSasaki, T., Brakebusch, C., Engel, J., & Timpl, R. (1998) · The EMBO Journal · doi:10.1093/emboj/17.6.1606 · Cited by 245 · cited 2× · see citing works
“These proteins also have high molecular weights and high plasma concentrations (A2M, FN1) or the ability to form multi-molecular complexes (THBS-1 and LGALS3BP) (Sasaki et al., 1998; Tan et al., 2006).”
BackgroundCorso, G., Mäger, I., Lee, Y., Görgens, A., Bultema, J. J., Giebel, B., Wood, M. J. A., Nordin, J. Z., & Andaloussi, S. E. (2017) · Scientific Reports · doi:10.1038/s41598-017-10646-x · Cited by 245 · cited 1× · see citing works
“Sizeexclusion chromatography (SEC)-based EV isolation was reported to rapidly isolate relatively pure EVs from plasma (Böing et al., 2014; Corso et al., 2017; Kreimer and Ivanov, 2017) and was shown to produce exosomes that are suitable for downstream proteomics analysis (Jeannin et al.”
BackgroundExosomes as new players in metabolic organ cross‐talk
Guay, C., & Regazzi, R. (2017) · Diabetes Obesity and Metabolism · doi:10.1111/dom.13027 · Cited by 244 · cited 1× · see citing works
“Integrated physiology relies on proper communication between different organs, and interest in extracellular vesicles (EVs) has grown rapidly in recent years, because of their role in inter-organ communication (Colombo et al., 2014; Guay and Regazzi, 2017; van Niel et al., 2018).”
BackgroundThe Human Erythrocyte Proteome
Kakhniashvili, D. G., Bulla, L. A., & Goodman, S. R. (2004) · Molecular & Cellular Proteomics · doi:10.1074/mcp.m300132-mcp200 · Cited by 220 · cited 1× · see citing works
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“We also demonstrate the importance of following the rigorous pre-analytical guidelines established by the ISEV and ISTH to limit ex vivo generation of platelet EVs (Artoni et al., 2012; Ayers et al., 2011; Burnouf et al., 2014; LaCroix et al., 2012).”
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“We also demonstrate the importance of following the rigorous pre-analytical guidelines established by the ISEV and ISTH to limit ex vivo generation of platelet EVs (Artoni et al., 2012; Ayers et al., 2011; Burnouf et al., 2014; LaCroix et al., 2012).”
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“A particularly problematic issue is the several-fold increase in plasma EV concentration caused by platelets upon cooling/ freezing (Artoni et al., 2012; György et al., 2012).”
BackgroundZhang, H., Liu, J., Qu, D. C., Wang, L., Wong, C. M., Lau, C., Huang, Y., Wang, Y. F., Huang, H., Xia, Y. Y., et al. (2018) · Proceedings of the National Academy of Sciences · doi:10.1073/pnas.1721521115 · Cited by 138 · cited 1× · see citing works
“community, several recent studies examining the plasma EV proteome have utilized frozen serum or plasma without platelet depletion (Chen et al., 2017; Jeannin et al., 2018; Whitham et al., 2018; Zhang et al., 2018), which facilitates analysis of samples from previous wellcharacterized studies.”
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“These proteins also have high molecular weights and high plasma concentrations (A2M, FN1) or the ability to form multi-molecular complexes (THBS-1 and LGALS3BP) (Sasaki et al., 1998; Tan et al., 2006).”
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“54; Table S1), which is within the acceptable limits of typical label-free proteomics experiments (Al Shweiki et al., 2017; Shen et al., 2018; Tabb et al., 2010).”
BackgroundProteomic Profiling of Serum-Derived Exosomes from Ethnically Diverse Prostate Cancer Patients
Turay, D., Khan, S., Osterman, C. J. D., Curtis, M. P., Khaira, B., Neidigh, J. W., Mirshahidi, S., Casiano, C. A., & Wall, N. R. (2015) · Cancer Investigation · doi:10.3109/07357907.2015.1081921 · Cited by 71 · cited 1× · see citing works
“, 2019), polymer precipitation (Turay et al., 2016), and immunoprecipitation (Ghosh et al.”
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“This separation enables improved proteomic characterization of EVs (Jeannin et al., 2018; Karimi et al., 2018), largely by increasing isolation yield and reducing plasma protein contamination (Stranska et al.”
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“Sizeexclusion chromatography (SEC)-based EV isolation was reported to rapidly isolate relatively pure EVs from plasma (Böing et al., 2014; Corso et al., 2017; Kreimer and Ivanov, 2017) and was shown to produce exosomes that are suitable for downstream proteomics analysis (Jeannin et al.”
BackgroundProteomic Analysis of Extracellular Vesicles for Cancer Diagnostics
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“EVs are membranous structures released from most cells that serve as vehicles to exchange protein, nucleic acid, and lipid cargo (Korutla et al., 2019; Minciacchi et al., 2015; van Niel et al., 2018) between distant organs.”
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MethodResidual platelets are the main determinants of microparticles count in frozen-thawed plasma
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“We also demonstrate the importance of following the rigorous pre-analytical guidelines established by the ISEV and ISTH to limit ex vivo generation of platelet EVs (Artoni et al., 2012; Ayers et al., 2011; Burnouf et al., 2014; LaCroix et al., 2012).”
BackgroundMethodInfluential
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Extracellular vesicles as tools and targets in therapy for diseases
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“Despite UC being a commonly used EV isolation method, several studies have reported poor reproducibility (Tian et al., 2020; Vanderboom et al., 2021).”
MethodSelf-citationBiofluid specific protein coronas affect lipid nanoparticle behavior in vitro
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“Purification of human plasma exosomes for proteomics: optimization and application to detect changes in response to exercise 40,41 Size exclusion chromatography (SEC) 29 Large particles elute faster in the chromatography since they are less retained by matrix.”
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“were low g isolated followed by size exclusion chromatography (SEC) likely avoiding physical damage or aggregation of those EVs [214, 221, 222].”
MethodChong, M. C., Shah, A. D., Schittenhelm, R. B., Silva, A., James, P. F., Wu, S. S. X., & Howitt, J. A. (2024) · Acta Physiologica · doi:10.1111/apha.14095 · Cited by 28 · see citing works
“22,23 In addition, other factors that may affect the protein composition of circulating EVs after exercise such as age and aerobic fitness have not been examined.”
BackgroundExercise, healthy ageing, and the potential role of small extracellular vesicles
McIlvenna, L. C., & Whitham, M. (2022) · The Journal of Physiology · doi:10.1113/jp282468 · Cited by 27 · see citing works
“(2021) were analysed and cross-referenced with the CellAge Database using Perseus (Tyanova et al., 2016).”
BackgroundMethodInfluentialMcIlvenna, L. C., Parker, H., Seabright, A. P., Sale, B., Anghileri, G., Weaver, S. R., Lucas, S. J. E., & Whitham, M. (2023) · The Journal of Physiology · doi:10.1113/jp284047 · Cited by 27 · see citing works
“Furthermore, we and others (Vanderboom et al., 2021; Whitham et al., 2018) have carried out unbiased, high coverage, shotgun proteomic analyses of crude EV isolates from platelet-free plasma taken from exercising human participants.”
BackgroundInfluentialMass spectrometry‐based proteomics approaches to interrogate skeletal muscle adaptations to exercise
Cervone, D. T., Moreno‐Justicia, R., Quesada, J. P., & Deshmukh, A. S. (2023) · Scandinavian Journal of Medicine and Science in Sports · doi:10.1111/sms.14334 · Cited by 26 · see citing works
Llorente, A., Brokāne, A., Mlynska, A., Puurand, M., Sagini, K., Folkmane, S., Hjorth, M., Martín-Gracia, B., Romero, S., Skorinkina, D., et al. (2024) · Journal of Extracellular Vesicles · doi:10.1002/jev2.12500 · Cited by 25 · see citing works
“…after exercise compared to before exercise are involved in signal transduction, the immune response, metabolism and oxidative stress (Apostolopoulou et al., 2021; Chong et al., 2024; Giloteaux et al., 2024; Kobayashi et al., 2021; Lisi et al., 2023; Vanderboom et al., 2021; Whitham et al., 2018).”
BackgroundCatitti, G., Bellis, D. D., Vespa, S., Simeone, P., Canonico, B., & Lanuti, P. (2022) · International Journal of Molecular Sciences · doi:10.3390/ijms232214098 · Cited by 26 · see citing works
Comprehensive review of MS‐based studies on N‐glycoproteome and N‐glycome of extracellular vesicles
Li, Y., Wang, J., Chen, W., Lu, H., & Zhang, Y. (2023) · PROTEOMICS · doi:10.1002/pmic.202300065 · Cited by 25 · see citing works
Engineered extracellular vesicles: an emerging nanomedicine therapeutic platform
Tang, J., Li, D., Wang, R., Li, S., Xing, Y., & Yu, F. (2025) · Chemical Communications · doi:10.1039/d4cc06501h · Cited by 24 · see citing works
Extracellular Vesicles in Pathophysiology: A Prudent Target That Requires Careful Consideration
Shahi, S. K., Kang, T., & Fonseka, P. (2024) · Cells · doi:10.3390/cells13090754 · Cited by 23 · see citing works
“As mentioned earlier, the biomolecular cargo of EVs is largely determined by the cells of origin [59–62].”
BackgroundMohamed, A. H., Abaza, T., Youssef, Y. A., Rady, M., Fahmy, S. A., Kamel, R., Hamdi, N., Efthimiadou, E. Κ., Braoudaki, M., & Youness, R. A. (2025) · Nanoscale Advances · doi:10.1039/d4na00393d · Cited by 23 · see citing works
Torres, A., Bernardo, L., Sánchez, C., Morato, E., Solana, J. C., & Carrillo, E. (2024) · Biological Procedures Online · doi:10.1186/s12575-024-00243-4 · Cited by 22 · see citing works
“However, this method has not yet been tested on long-term stored biobank plasma samples [39, 40].”
MethodA standardized method for plasma extracellular vesicle isolation and size distribution analysis
Diehl, J. N., Ray, A., Collins, L. B., Peterson, A., Alexander, K. C., Boutros, J. G., Ikonomidis, J. S., & Akerman, A. W. (2023) · PLoS ONE · doi:10.1371/journal.pone.0284875 · Cited by 22 · see citing works
Newman, L. A., Useckaite, Z., & Rowland, A. (2022) · Journal of Extracellular Biology · doi:10.1002/jex2.56 · Cited by 21 · see citing works
“…has been reported to isolate a greater ratio of vesicle to serum proteins, compared to precipitation reagents or conventional techniques such as ultracentrifugation, and can reproducibly isolate vesicles containing characteristic EV markers (Monguió-Tortajada et al., 2019; Vanderboom et al., 2021).”
InfluentialReymond, S., Gruaz, L., & Sanchez, J. (2023) · Analytical and Bioanalytical Chemistry · doi:10.1007/s00216-023-04684-w · Cited by 21 · see citing works
“Of note, Vanderboom and colleagues [25] developed a SECMS approach, which identified almost 1300 proteins in EVs derived from 2 mL of plasma and more than 2700 proteins when participants performed aerobic exercise.”
BackgroundInfluentialLattmann, E., Raess, L., Tognetti, M., Gómez, J. M. M., Lapaire, V., Bruderer, R., Reiter, L., Feng, Y., Steinmetz, L. M., & Levesque, M. P. (2024) · Cellular and Molecular Life Sciences · doi:10.1007/s00018-024-05137-y · Cited by 21 · see citing works
“This aligns with recent findings where SEC of platelet-poor plasma outperformed other EV isolation methods in the detection of EV markers [64].”
ResultMlynska, A., Dobrovolskienė, N., Žilionytė, K., Lukaseviciute, G., Sagini, K., Martin-Gracia, B., Romero, S., Llorente, A., Linē, A., Butkute, A., et al. (2025) · Journal of sport and health science/Journal of Sport and Health Science · doi:10.1016/j.jshs.2025.101041 · Cited by 20 · see citing works
Extracellular vesicles contribute to the beneficial effects of exercise training in APP/PS1 mice
Fuller, O. K., McLennan, E. D., Egan, C. L., Perera, N., Terry, L. V., Pyun, J., de Mendonca, M., Telles, G. D., Smeuninx, B., Burrows, E. L., et al. (2025) · iScience · doi:10.1016/j.isci.2025.111752 · Cited by 17 · see citing works
“54,84 These findings underscore the complex roleofEVsinmediatingthesystemiceffectsofexercise, suggestingthatthedifferential cargoofEVsfromphysicalactivity could influence metabolic and neurodegenerative processes.”
György, B., Pálóczi, K., Balbisi, M., Turiák, L., Drahos, L., Visnovitz, T., Koltai, E., & Radak, Z. O. (2024) · Current Issues in Molecular Biology · doi:10.3390/cimb46050264 · Cited by 16 · see citing works
Pataky, M. W., Heppelmann, C. J. H., Sevits, K. J., Asokan, A. K., Kumar, A. P., Klaus, K. A., Dasari, S., Kunz, H., Strub, M. D., Robinson, M. M., et al. (2025) · Nature Communications · doi:10.1038/s41467-025-60049-0 · Cited by 16 · see citing works
Self-citationStar, A. T., Hewitt, M., Badhwar, A., Ding, W., Tremblay, T., Hill, J. J., Willmore, W. G., Sandhu, J. K., & Haqqani, A. S. (2025) · Proteomes · doi:10.3390/proteomes13030045 · Cited by 15 · see citing works
“These findings align with other reports that SEC-based isolation, especially using Izon qEV columns, yields deeper proteomic coverage and cleaner EV preparations than precipitation or basic centrifugation methods [1,3].”
InfluentialModern Strategies of Drug Therapy: Multi-Target Drug Delivery, Bioimaging, Diagnostics
Fedorova, O. V., Ovchinnikova, I. G., Rusinov, G. L., Avdeeva, V. V., Zhdanov, А. P., Zhizhin, K. Y., Kuznetsov, N. T., Zakharova, L. Y., Kuznetsova, D. A., Razuvayeva, Y. S., et al. (2025) · Russian Journal of General Chemistry · doi:10.1134/s1070363225606726 · Cited by 15 · see citing works
Ramachandran, R. A., Lemoff, A. S., & Robertson, D. M. (2023) · Infection and Immunity · doi:10.1128/iai.00036-23 · Cited by 15 · see citing works
“For mass spectrometric analysis, SEC coupled with high-resolution mass spectrometry of human plasma EVs improved EV protein identification and precision compared to UC (55).”
MethodAibaidula, A., Fain, C. E., Garcia, L. M. C., Wier, A., Bouchal, S. M., Bauman, M. M. J., Jung, M. K., Sarkaria, J. N., Johnson, A. J., & Parney, I. F. (2023) · Neuro-Oncology Advances · doi:10.1093/noajnl/vdad082 · Cited by 14 · see citing works
Mastrototaro, L., & Roden, M. (2023) · PROTEOMICS · doi:10.1002/pmic.202300078 · Cited by 12 · see citing works
“Arecentstudydemonstrates that SEC improves proteomic characterization of plasma-derived EV and identifies more EV marker proteins compared to UC [147].”
BackgroundResultInfluentialExtracellular vesicle biomarkers in ocular fluids associated with ophthalmic diseases
Shiju, T. M., & Yuan, A. (2024) · Experimental Eye Research · doi:10.1016/j.exer.2024.109831 · Cited by 12 · see citing works
Jaimes, M. A., Liao, C., Chen, M. M., Czosseck, A., Lee, T., Chou, Y., Chen, Y., Lin, S., Lai, J. J., & Lundy, D. J. (2023) · Journal of Extracellular Biology · doi:10.1002/jex2.86 · Cited by 12 · see citing works
“Regarding circulating EVs specifically, a 2021 study also found that both resistance training and aerobic exercise altered EV cargo in human blood plasma (Vanderboom et al., 2021).”
BackgroundExtracellular vesicles in atherosclerosis cardiovascular disease: emerging roles and mechanisms
Deng, H., Qiu, W., Zhang, Y., & Hua, J. (2025) · Frontiers in Cardiovascular Medicine · doi:10.3389/fcvm.2025.1611557 · Cited by 11 · see citing works
Emerging Frontiers in acute kidney injury: The role of extracellular vesicles
Li, S., Zhou, L., Huang, Y., & Tang, S. (2025) · Bioactive Materials · doi:10.1016/j.bioactmat.2025.02.018 · Cited by 11 · see citing works
“…into four categories: a. Size/density-based centrifugation methods, such as ultracentrifugation (UC) [172], size exclusion chromatography (SEC) [173], and density gradient ultracentrifugation [174]; b. Filtration-based separations, including ultrafiltration (UF) [175] and Exodisc [176]; c.…”
Brendborg, N., & Febbraio, M. A. (2025) · The Journal of Physiology · doi:10.1113/jp286747 · Cited by 10 · see citing works
Multiomics analysis to evaluate the enrichment of extracellular vesicles from human plasma
Su, H., Fowler, C. J., Masters, C. L., Barnham, K. J., Reid, G. E., & Vella, L. J. (2025) · Journal of Lipid Research · doi:10.1016/j.jlr.2025.100877 · Cited by 9 · see citing works
“Some studies have previously reported the proteome (14, 16, 18, 20, 22, 24, 39 – 42) and lipidome of EVs in blood (17, 19, 21).”
Guerricchio, L., Barile, L., & Bollini, S. (2024) · International Journal of Molecular Sciences · doi:10.3390/ijms25116187 · Cited by 8 · see citing works
Filter-Aided Extracellular Vesicle Enrichment (FAEVEr) for Proteomics
Pauwels, J., de Steene, T. V., Velde, J. V. D., Muyer, F. D., Pauw, D. D., Baeke, F., Eyckerman, S., & Gevaert, K. (2025) · Molecular & Cellular Proteomics · doi:10.1016/j.mcpro.2025.100907 · Cited by 8 · see citing works
“…pro fi ling, with the most prominent strategies being based on size (ultracentrifugation (UC) and size-exclusion chromatography (SEC)) (34, 35), density (density-gradient UC) (36), precipitation (37 – 39), EV membrane interaction (EVTrap) (9, 40), or capture of molecular markers present…”
Leme, A. F. P., Yokoo, S., Normando, A. G. C., Ormonde, J. V. S., Domingues, R. R., Cruz, F. F., Silva, P. L., Souza, B. S. D. F., dos Santos, C. C., Castro-Faria-Neto, H. C., et al. (2024) · Cytotherapy · doi:10.1016/j.jcyt.2024.02.001 · Cited by 7 · see citing works
Hu, A., Zhang, J., Zhang, L., Wang, Z., Dai, J., Lin, L., Yan, G., Shen, F., & Shen, H. (2024) · Analytical Chemistry · doi:10.1021/acs.analchem.4c03006 · Cited by 7 · see citing works
Elsworthy, R. J., Dunleavy, C., Whitham, M., & Aldred, S. (2022) · Aging and Health Research · doi:10.1016/j.ahr.2022.100093 · Cited by 7 · see citing works
Beckner, M. E., Karl, J. P., Armstrong, N. J., Carrigan, C. T., Wilson, M. A., Pirnstill, H. D., Karis, A. J., Pasiakos, S. M., McClung, J. P., Barringer, N., et al. (2025) · Physiological Genomics · doi:10.1152/physiolgenomics.00155.2025 · Cited by 6 · see citing works
Wei, M., Wang, D., Xu, W., Zang, X., & Jiang, J. (2026) · International Journal of Nanomedicine · doi:10.2147/ijn.s583756 · Cited by 5 · see citing works
Ráez-Meseguer, C., Amengual-Tugores, A. M., Forteza‐Genestra, M. A., Orvay, F., Gomila, R. M., Martorell-Crespí, G., Calvo, J., Gayà, A., Monjo, M., & Ramis, J. M. (2025) · Journal of Proteome Research · doi:10.1021/acs.jproteome.5c00089 · Cited by 4 · see citing works
Vanderboom, P. M., Chawla, Y., Dasari, S., Kapoor, I., Kumar, S. K., Nair, K. S., & Gonsalves, W. I. (2024) · European Journal Of Haematology · doi:10.1111/ejh.14246 · Cited by 4 · see citing works
“6 Briefly, frozen platelet-poor plasma was thawed on ice and centrifuged at 2000 g for 10 min to pellet debris.”
Self-citationSagini, K., Selvarajah, M., Romero, S., Bassols‐Citores, E., Martín-Gracia, B., Ramírez‐Garrastacho, M., Rise, F., Endzeliņš, E., Sadovska, L., Skorinkina, D., et al. (2025) · The FASEB Journal · doi:10.1096/fj.202500678r · Cited by 4 · see citing works
“Plasma EVs were enriched by SEC, which removes soluble proteins and the smallest lipoproteins [24, 41–43].”
InfluentialAbbasi, A., Hansen, N. F., Palade, J., Paredes, D. M., Meechoovet, B., Keuren‐Jensen, K. R. V., Pirrotte, P., & Stringer, W. W. (2026) · Scientific Reports · doi:10.1038/s41598-025-23760-y · Cited by 3 · see citing works
Peris-Torres, C., & Rodrı́guez-Manzaneque, J. C. (2026) · FEBS Journal · doi:10.1111/febs.70433 · Cited by 3 · see citing works
“Supporting this, EV-associated ADAMTS proteoglycanases have been identified in diverse biological sources, including primary myotube-derived vesicles [25] and biological fluids such as urine and maternal plasma (Table 1), indicating physiological roles beyond the cell of origin.”
Exercise-primed extracellular vesicles improve cell-matrix adhesion and chondrocyte healthPreprint
Iijima, H., Wang, K., D’Amico, E., Tang, W., Rogers, R. J., Jakicic, J. M., & Ambrosio, F. (2023) · Research Square · doi:10.21203/rs.3.rs-2958821/v1 · Cited by 3 · see citing works
Bunn, K. E., Bressman, Z. J., Nation, G. K., Wu, P., Amin, T., Lehman, C., Lin, A., Gray, C. J., Laidlaw, T. M., Cahill, K. N., et al. (2025) · American Journal of Respiratory Cell and Molecular Biology · doi:10.1165/rcmb.2024-0414oc · Cited by 2 · see citing works
“Cell – derived EVs Are Present in Nasal Lining Fluid EVs in bio fl uids carry cargos that re fl ect cells within and adjacent to that bio fl uid (28, 29).”
Fliflet, A. M., Spradlin, R. A., Tan, Y., Vijayan, A. N., Choi, S. J., Kao, W., Aksamitiene, E., Nelappana, M. B., Ding, S., Huang, K., et al. (2026) · The FASEB Journal · doi:10.1096/fj.202505096rr · Cited by 2 · see citing works
Tacrolimus regulates extracellular vesicle secretion from T cells via autophagy-lysosomal pathway
Chen, C., Hung, T., Huang, Y.-J., Hung, H.-S., Hu, C., & Lee, P. (2024) · Biomedicine & Pharmacotherapy · doi:10.1016/j.biopha.2024.117765 · Cited by 2 · see citing works
You, Y., Zhang, Z., Cortes, S., Nguyen, S., Vadakattu, P., Melvin, B. C., Mann, S. D. J., Ray, N. B., Bregendahl, M. E., McLean, P. J., et al. (2025) · Cell Communication and Signaling · doi:10.1186/s12964-025-02263-3 · Cited by 2 · see citing works
“Previous studies reported that SEC-based isolation minimally alters EV characteristics with less plasma protein contamination compared to other conventional isolation approaches such as ultracentrifugation and precipitation [24, 25].”
Vallejo, M. C., Sarkar, S., Elliott, E. C., Henry, H. R., Huang, F., Payne, S. H., Ramanadham, S., Sims, E. K., Metz, T., Mirmira, R. G., et al. (2022) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/2022.09.28.509996 · Cited by 2 · see citing works
Exosome-Delivered Factors and their Influence on Angiogenic Processes in Bone Healing
Emami, A., & Oskouie, I. M. (2026) · Regenerative Engineering and Translational Medicine · doi:10.1007/s40883-025-00535-5 · Cited by 1 · see citing works
Davies, C., Mittal, R., Li, C. Y., Marwede, H., Bergman, J. E., Hilton, N., Mittal, J., Bhattacharya, S. K., & Eshraghi, A. A. (2022) · Journal of Clinical Medicine · doi:10.3390/jcm11144072 · Cited by 1 · see citing works
“Among the techniques used to identify and characterize host pathways, tandem mass spectrometry (MS/MS or MS) has recently become a powerful tool [26,27].”
BackgroundVanderboom, P. M., Chawla, Y., Dasari, S., Kapoor, I., Kumar, S. K., N, K. S., & Gonsalves, W. I. (2023) · Research Square · doi:10.21203/rs.3.rs-3673796/v1
Self-citationMulti-omics characterization of highly enriched human plasma extracellular vesiclesPreprint
Su, H., Fowler, C. J., Masters, C. L., Barnham, K. J., Reid, G. E., & Vella, L. J. (2024) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/2024.03.25.586695
“Multiple studies have previously reported the proteomes [27, 36, 40-47] and lipidomes of blood EV preparations [48-50] However, the large majority of isolation methods used in these studies would have co-isolated contaminates.”
Isolation of functional supramolecular attack particles (SMAPs)Preprint
Jainarayanan, A., Chandrasekar, V., Almeida, V. P., Widdess, M. A., Staton, C. C., Bernand, A., Pfannenstill, V., Flannery, S. E., Watson, H., Wicki, A., et al. (2025) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.1101/2025.11.10.687675
Identifying Subcellular Structure Components in Escherichia Coli by Crosslinking and SEC‐MS
Victor, R. A., Lipinski, A. A., Langlais, P. R., & Schwartz, J. C. (2026) · PROTEOMICS · doi:10.1002/pmic.70105
“SEC-MS is an important method in the fields of proteomics, structural biology, and biopharmaceutical development [ 27, 28 ].”
Extracellular vesicle proteomics and metabolomics: analytical frameworks and translational insights
Akchiche, Y. F., Ousmaal, M. E. F., & Boumehira, A. Z. (2025) · Bioanalysis · doi:10.1080/17576180.2026.2617928
Gupta, D., Sevits, K. J., Klaus, K. A., LOONA, S. S., PARMAR, V., Kim, Y., Heppelmann, C. J. H., Leija, R. G., Kunz, H., Lucien, F., et al. (2026) · medRxiv · doi:10.64898/2026.05.14.26353181 · Cited by 1 · see citing works
Self-citationde Souza, T. D. F. G., Klassen, N., Obi, P. O., Özerkliğ, B., Tiede, T., Srivastava, A., Pascoe, C. D., Marin, S. E., Dhingra, S., Greenberg, C. R., et al. (2026) · bioRxiv (Cold Spring Harbor Laboratory) · doi:10.64898/2026.06.12.731704
Methods for the Investigation of Protein–Ligands Interactions
Iacobucci, I., Cozzolino, F., Cipollone, I., & Monti, M. (2026) · Advances in experimental medicine and biology · doi:10.1007/978-3-032-21638-0_8
Extracellular vesicles as mediators of exercise effects in cancer
Aires, I., Moreira‐Gonçalves, D., D’Agostino, V. G., Ferreira, R. M., & Vitorino, R. M. P. (2026) · Molecular Medicine · doi:10.1186/s10020-026-01577-3
“…prioritizes EVs carrying myogenic miRs (miR-1, miR-133 and miR-206) and regulatory proteins targeting mTOR, PI3K/AKT, TGF-β and IL6 signaling axes, supporting muscle hypertrophy, tissue repair and immune surveillance in recipient cells (Vanderboom et al. 2021, Conkright et al. 2024, Gonzalez 2016).”
Jo, E., Yang, H., Lee, S., Seo, C. W., Jung, W., Shin, Y.-H., Park, H., Lee, Y.-S., Lee, E.-Y., Song, H.-Y., et al. (2026) · Journal of Proteome Research · doi:10.1021/acs.jproteome.6c00305
EV‐LYSG101: Extracellular Vesicles Loaded With Lysin LYSG101 for Potent Anti‐Staphylococcal Activity
Hoelzinger, D. B., Celaya, A. K. D., Myers, C. E., Dyal, N. P., Saling, C. F., Schuch, R., & Suh, G. A. (2026) · Journal of Extracellular Biology · doi:10.1002/jex2.70174
Exercise enhances aged chondrocyte health through microRNA-29-enriched extracellular vesicles
Iijima, H., Dave, K. M., Wang, K., Khay, A. M., D'Amico, E., Tang, W.-Y., Rogers, R. J., Jakicic, J. M., & Ambrosio, F. (2026) · Nature Aging · doi:10.1038/s43587-026-01225-9
Maqbool, H., Ullah, N., Fazil, I., Khan, M., Khan, M. A., Chaudhary, H. J., Zheng, J., & Naz, I. (2026) · World Journal of Microbiology and Biotechnology · doi:10.1007/s11274-026-05301-5
“…(SEC) can reduce the mechanical stresses associated with repeated pelleting and is readily amenable to standardized workflows however, its resolving power may be limited when vesicles and similarly sized non-vesicular particles co-elute (Vanderboom et al. 2021; Monguió-Tortajada et al. 2019 ).”
- Authors
- Vanderboom, P. M., Dasari, S., Ruegsegger, G. N., Pataky, M. W., Lucien, F., Heppelmann, C. J., Lanza, I. R., & Nair, K. S.
- Affiliations
- Mayo Clinic (US)
- Corresponding author
- Nair, K. Sreekumaran
- ORCID
- Venue
- Cell Reports Methods
- Publisher
- Elsevier BV
- Published
- 2021-07
- Volume
- 1
- Issue
- 3
- Pages
- 100055
- ISSN
- 2667-2375 (ISSN-L)
- PMC
- PMC8336930
- OpenAlex
- W3184543356
- Cited by
- 75
- References
- 44
- Citations by year
- 2026: 14 · 2025: 20 · 2024: 19 · 2023: 12 · 2022: 10
- FWCI
- 3.22
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- Cell Reports Methods (publishedVersion, cc-by-nc-nd)
- PubMed Central (publishedVersion, cc-by-nc-nd)
- PubMed (publishedVersion)
- DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
- Funders
- Mayo Clinic (T32-AG-49672, U24-DK-100469)
- National Center for Advancing Translational Sciences
- Georgia Clinical and Translational Science Alliance
- National Institutes of Health (R01 AG062859, UL1 TR002377)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Extracellular vesicles in disease
- Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Caveolin-1 and cellular processes
- Keywords
- extracellular vesicles, size exclusion chromatography, platelet-poor plasma, EV proteome, inter-organ communication
- MeSH terms
- Extracellular Vesicles, Chromatography, Gel, Humans, Proteins, Mass Spectrometry, Proteomics
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