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Human saliva-derived exosomes: comparing methods of isolation.
- Source :
-
The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society [J Histochem Cytochem] 2015 Mar; Vol. 63 (3), pp. 181-9. Date of Electronic Publication: 2014 Dec 03. - Publication Year :
- 2015
-
Abstract
- ExoQuick-TC(TM) (EQ), a chemical-based agent designed to precipitate exosomes, was calibrated for use on saliva collected from healthy individuals. The morphological and molecular features of the precipitations were compared with those obtained using the classical, physical-based method of ultracentrifugation (UC). Electron microscopy and immunoelectron microscopy with anti-CD63 showed vesicular nanoparticles surrounded by bi-layered membrane, compatible with exosomes in EQ, similar to that observed with UC. Atomic force microscopy highlighted larger, irregularly shaped/aggregated EQ nanoparticles that contrasted with the single, round-shaped UC nanoparticles. ELISA (performed on 0.5 ml of saliva) revealed a tendency for a higher expression of the specific exosomal markers (CD63, CD9, CD81) in EQ than in UC (p>0.05). ELISA for epithelial growth factor receptor, a non-exosomal-related marker, showed a significantly higher concentration in EQ than in UC (p=0.04). Western blotting of equal total-protein concentrations revealed bands of CD63, CD9 and CD81 in both types of preparations, although they were less pronounced in EQ compared with UC. This may be related to a higher fraction of non-exosomal proteins in EQ. In conclusion, EQ is suitable and efficient for precipitation of salivary exosomes from small volumes of saliva; however, EQ tends to be associated with considerably more biological impurities (non-exosomal-related proteins/microvesicles) as compared with UC.<br /> (© The Author(s) 2015.)
- Subjects :
- Adult
Aged
Blotting, Western
Chemical Precipitation
Enzyme-Linked Immunosorbent Assay
Exosomes ultrastructure
Female
Humans
Male
Microscopy, Atomic Force
Microscopy, Electron
Middle Aged
Saliva chemistry
Ultracentrifugation
Exosomes chemistry
Saliva cytology
Tetraspanin 28 analysis
Tetraspanin 29 analysis
Tetraspanin 30 analysis
Subjects
Details
- Language :
- English
- ISSN :
- 1551-5044
- Volume :
- 63
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- The journal of histochemistry and cytochemistry : official journal of the Histochemistry Society
- Publication Type :
- Academic Journal
- Accession number :
- 25473095
- Full Text :
- https://doi.org/10.1369/0022155414564219