Isolation of Extracellular Vesicles in Saliva Using Density Gradient Ultracentrifugation

  • Kazuya Iwai
  • Satoshi Yamamoto
  • Mitsutaka Yoshida
  • Kiyotaka ShibaEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1660)


This chapter describes a method for isolating human salivary extracellular vesicles (EVs) using density gradient ultracentrifugation. Standard protocols established for isolation of EVs from blood or a conditioned medium of cultured cells do not work for whole saliva, due to its viscosity. Therefore, procedures including a pretreatment step and utilizing iodixanol as a gradient material enable EVs to be concentrated to a 1.1 g/ml density. This protocol is compatible with both swing and angle rotors. By employing an angle rotor, which enables high g-force, the centrifugation time was reduced to 4 h from the 17 h required when using a swing rotor.

Key words

Exosome Extracellular vesicles EVs Iodixanol Angle rotor Whole saliva 



This work was supported by the “Project for Private Universities: Matching Fund Subsidy” from the Ministry of Education, Culture, Sports, Science, and Technology and by a grant from the Vehicle Racing Commemorative Foundation.


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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  • Kazuya Iwai
    • 1
    • 2
  • Satoshi Yamamoto
    • 1
    • 2
  • Mitsutaka Yoshida
    • 1
    • 2
  • Kiyotaka Shiba
    • 1
    Email author
  1. 1.Division of Protein Engineering, Cancer InstituteJapanese Foundation for Cancer ResearchTokyoJapan
  2. 2.Department of Oral & Maxillofacial ImplantologyTokyo Dental CollegeTokyoJapan

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