Display options
Cite
Buschmann D, Kirchner B, Hermann S, et al. Evaluation of serum extracellular vesicle isolation methods for profiling miRNAs by next-generation sequencing. J Extracell Vesicles. 2018;7(1):1481321doi: 10.1080/20013078.2018.1481321.
Buschmann, D., Kirchner, B., Hermann, S., Märte, M., Wurmser, C., Brandes, F., Kotschote, S., Bonin, M., Steinlein, O. K., Pfaffl, M. W., Schelling, G., & Reithmair, M. (2018). Evaluation of serum extracellular vesicle isolation methods for profiling miRNAs by next-generation sequencing. Journal of extracellular vesicles, 7(1), 1481321. https://doi.org/10.1080/20013078.2018.1481321
Buschmann, Dominik, et al. "Evaluation of serum extracellular vesicle isolation methods for profiling miRNAs by next-generation sequencing." Journal of extracellular vesicles vol. 7,1 (2018): 1481321. doi: https://doi.org/10.1080/20013078.2018.1481321
Buschmann D, Kirchner B, Hermann S, Märte M, Wurmser C, Brandes F, Kotschote S, Bonin M, Steinlein OK, Pfaffl MW, Schelling G, Reithmair M. Evaluation of serum extracellular vesicle isolation methods for profiling miRNAs by next-generation sequencing. J Extracell Vesicles. 2018 Jun 04;7(1):1481321. doi: 10.1080/20013078.2018.1481321. eCollection 2018. PMID: 29887978; PMCID: PMC5990937.
Copy Download .nbib Format: NLM
Share it on

J Extracell Vesicles. 2018 Jun 04;7(1):1481321. doi: 10.1080/20013078.2018.1481321. eCollection 2018.

Evaluation of serum extracellular vesicle isolation methods for profiling miRNAs by next-generation sequencing.

Journal of extracellular vesicles

Dominik Buschmann, Benedikt Kirchner, Stefanie Hermann, Melanie Märte, Christine Wurmser, Florian Brandes, Stefan Kotschote, Michael Bonin, Ortrud K Steinlein, Michael W Pfaffl, Gustav Schelling, Marlene Reithmair

Affiliations

  1. Institute of Human Genetics, University Hospital, LMU Munich, Munich, Germany.
  2. Division of Animal Physiology and Immunology, TUM School of Life Sciences Weihenstephan, Technical University of Munich, Freising, Germany.
  3. Dr. von Hauner Children's Hospital, LMU Munich,  Munich, Germany.
  4. Department of Anesthesiology, University Hospital, LMU Munich, Munich, Germany.
  5. Chair of Animal Breeding, TUM School of Life Sciences Weihenstephan, Technical University of Munich, Freising, Germany.
  6. IMGM Laboratories GmbH, Planegg, Germany.

PMID: 29887978 PMCID: PMC5990937 DOI: 10.1080/20013078.2018.1481321

Abstract

Extracellular vesicles (EVs) are intercellular communicators with key functions in physiological and pathological processes and have recently garnered interest because of their diagnostic and therapeutic potential. The past decade has brought about the development and commercialization of a wide array of methods to isolate EVs from serum. Which subpopulations of EVs are captured strongly depends on the isolation method, which in turn determines how suitable resulting samples are for various downstream applications. To help clinicians and scientists choose the most appropriate approach for their experiments, isolation methods need to be comparatively characterized. Few attempts have been made to comprehensively analyse vesicular microRNAs (miRNAs) in patient biofluids for biomarker studies. To address this discrepancy, we set out to benchmark the performance of several isolation principles for serum EVs in healthy individuals and critically ill patients. Here, we compared five different methods of EV isolation in combination with two RNA extraction methods regarding their suitability for biomarker discovery-focused miRNA sequencing as well as biological characteristics of captured vesicles. Our findings reveal striking method-specific differences in both the properties of isolated vesicles and the ability of associated miRNAs to serve in biomarker research. While isolation by precipitation and membrane affinity was highly suitable for miRNA-based biomarker discovery, methods based on size-exclusion chromatography failed to separate patients from healthy volunteers. Isolated vesicles differed in size, quantity, purity and composition, indicating that each method captured distinctive populations of EVs as well as additional contaminants. Even though the focus of this work was on transcriptomic profiling of EV-miRNAs, our insights also apply to additional areas of research. We provide guidance for navigating the multitude of EV isolation methods available today and help researchers and clinicians make an informed choice about which strategy to use for experiments involving critically ill patients.

Keywords: Extracellular vesicle; biomarker; exosome isolation; miRNA; next-generation sequencing; precipitation; sepsis; small RNA sequencing; ultracentrifugation

References

  1. Nanomedicine. 2017 Apr;13(3):1011-1020 - PubMed
  2. J Immunol Methods. 2016 Feb;429:39-49 - PubMed
  3. J Extracell Vesicles. 2017 Nov 15;6(1):1396823 - PubMed
  4. Sci Rep. 2016 Jun 20;6:28006 - PubMed
  5. Clin Biochem. 2014 Jan;47(1-2):135-8 - PubMed
  6. Inflamm Res. 2014 Sep;63(9):741-56 - PubMed
  7. Mol Immunol. 2012 Apr;50(4):278-86 - PubMed
  8. J Extracell Vesicles. 2017 Mar 15;6(1):1294340 - PubMed
  9. BMC Genomics. 2017 Jan 7;18(1):50 - PubMed
  10. J Immunol Methods. 2014 Sep;411:55-65 - PubMed
  11. Clin Chem Lab Med. 2014 Jun;52(6):927-33 - PubMed
  12. J Extracell Vesicles. 2014 Feb 04;3:null - PubMed
  13. Int J Infect Dis. 2015 Nov;40:135-41 - PubMed
  14. Oncotarget. 2017 Jul 18;8(29):47317-47329 - PubMed
  15. G3 (Bethesda). 2017 Jan 5;7(1):31-39 - PubMed
  16. Sci Rep. 2017 May 2;7(1):1342 - PubMed
  17. J Histochem Cytochem. 2015 Mar;63(3):181-9 - PubMed
  18. J Extracell Vesicles. 2017 Jun 20;6(1):1329476 - PubMed
  19. Proc Natl Acad Sci U S A. 2016 Feb 23;113(8):E968-77 - PubMed
  20. PLoS One. 2015 Aug 28;10(8):e0136133 - PubMed
  21. PLoS One. 2017 Jan 6;12 (1):e0164644 - PubMed
  22. J Extracell Vesicles. 2016 Oct 31;5:32945 - PubMed
  23. Bioinformatics. 2015 Nov 15;31(22):3718-20 - PubMed
  24. PLoS One. 2009 Oct 12;4(10):e7405 - PubMed
  25. PLoS One. 2015 Dec 21;10(12):e0145686 - PubMed
  26. Methods Mol Biol. 2017;1545:55-70 - PubMed
  27. Proc Natl Acad Sci U S A. 2011 Mar 22;108(12):5003-8 - PubMed
  28. Methods. 2015 Oct 1;87:46-58 - PubMed
  29. Microbiologyopen. 2015 Jan 21;:null - PubMed
  30. Mol Biosyst. 2016 Apr 26;12 (5):1407-19 - PubMed
  31. Microb Pathog. 2017 Mar;104:161-163 - PubMed
  32. J Extracell Vesicles. 2015 Dec 23;4:29509 - PubMed
  33. Nucleic Acids Res. 2014 Jan;42(Database issue):D68-73 - PubMed
  34. Langmuir. 2012 Jun 19;28(24):9131-9 - PubMed
  35. J Transl Med. 2018 Jan 9;16(1):1 - PubMed
  36. Sci Rep. 2016 Apr 18;6:24316 - PubMed
  37. J Chronic Dis. 1987;40(5):373-83 - PubMed
  38. Front Neurosci. 2017 May 22;11:278 - PubMed
  39. Nucleic Acids Res. 2015 Jan;43(Database issue):D123-9 - PubMed
  40. Genomics. 2011 Aug;98(2):152-3 - PubMed
  41. J Extracell Vesicles. 2013 Jan 10;2:null - PubMed
  42. Sci Rep. 2016 Jan 20;6:19529 - PubMed
  43. BMC Genomics. 2013 May 10;14:319 - PubMed
  44. PLoS One. 2012;7(11):e49726 - PubMed
  45. Int J Mol Med. 2016 Nov;38(5):1359-1366 - PubMed
  46. J Extracell Vesicles. 2017 Apr 4;6(1):1308779 - PubMed
  47. PLoS One. 2012;7(6):e38885 - PubMed
  48. J Extracell Vesicles. 2013 Feb 15;2:null - PubMed
  49. Nat Commun. 2015 Jun 18;6:7321 - PubMed
  50. PLoS One. 2014 Sep 17;9(9):e107259 - PubMed
  51. Cancers (Basel). 2016 Dec 10;8(12 ): - PubMed
  52. J Extracell Vesicles. 2014 Sep 18;3:null - PubMed
  53. Philos Trans R Soc Lond B Biol Sci. 2014 Sep 26;369(1652):null - PubMed
  54. Food Chem Toxicol. 2017 Dec;110:229-239 - PubMed
  55. J Extracell Vesicles. 2015 Jul 17;4:27031 - PubMed
  56. JCI Insight. 2016 Nov 17;1(19):e89631 - PubMed
  57. Nanomedicine. 2017 Aug;13(6):2061-2065 - PubMed
  58. J Extracell Vesicles. 2016 Jun 20;5:31655 - PubMed
  59. Nat Rev Immunol. 2015 Jun;15(6):375-87 - PubMed
  60. Nucleic Acids Res. 2016 Jul 27;44(13):5995-6018 - PubMed
  61. Genome Biol. 2009;10(3):R25 - PubMed
  62. Liver Int. 2015 Apr;35(4):1172-84 - PubMed
  63. Inflammation. 2012 Aug;35(4):1308-13 - PubMed
  64. Front Oncol. 2014 Jun 04;4:129 - PubMed
  65. PLoS One. 2017 Jan 23;12 (1):e0170628 - PubMed
  66. World J Methodol. 2013 Mar 26;3(1):11-8 - PubMed
  67. J Cell Mol Med. 2017 Oct;21(10 ):2403-2411 - PubMed
  68. Proteomics. 2013 Nov;13(22):3354-64 - PubMed
  69. Crit Care. 2014 Dec 12;18(6):704 - PubMed
  70. RNA Biol. 2017 Jan 2;14 (1):58-72 - PubMed
  71. J Extracell Vesicles. 2017 Mar 7;6(1):1286095 - PubMed
  72. J Extracell Vesicles. 2016 Feb 19;5:29975 - PubMed
  73. J Extracell Vesicles. 2016 Feb 15;5:29497 - PubMed
  74. J Extracell Vesicles. 2015 Mar 26;4:27269 - PubMed
  75. Intensive Care Med. 2016 Dec;42(12 ):1980-1989 - PubMed
  76. RNA Biol. 2017 Feb;14 (2):245-258 - PubMed
  77. Sci Rep. 2017 Jul 13;7(1):5270 - PubMed
  78. Int J Mol Med. 2017 Sep;40(3):834-844 - PubMed
  79. Crit Care Clin. 2003 Jul;19(3):441-58 - PubMed
  80. J Immunol. 2007 Oct 15;179(8):5082-9 - PubMed
  81. J Extracell Vesicles. 2014 Mar 26;3:null - PubMed
  82. Crit Care Med. 2014 May;42(5):1096-104 - PubMed
  83. Sci Rep. 2016 Sep 19;6:33641 - PubMed
  84. J Extracell Vesicles. 2014 Dec 22;3:26913 - PubMed
  85. Kidney Int. 2012 Nov;82(9):1024-32 - PubMed
  86. Immunology. 2012 Jun;136(2):192-7 - PubMed
  87. Genome Biol. 2014;15(12):550 - PubMed

Publication Types