Oral Presentation 24th Australian Conference on Microscopy and Microanalysis 2016

Isolating brain derived exosomes: A new method in the study of neurodegenerative disease (#108)

Benjamin J Scicluna 1 2 , Laura J Vella 3 , Bradley M Coleman 1 , Lesley Cheng 2 , Kevin J Barnham 3 4 , Andrew F Hill 2
  1. Department of Biochemistry & Molecular Biology, The University of Melbourne, Parkville, VIC, Australia
  2. Department of Biochemistry & Genetics, La Trobe Institute of Molecular Science, La Trobe University, Bundoora, VIC, Australia
  3. The Florey Institute of Neuroscience and Mental Health, Parkville, VIC, Australia
  4. Department of Pharmacology, The University of Melbourne, Parkville, VIC, Australia


Exosomes are small (30 – 150 nm) extracellular vesicles (EVs) released from cells that can transfer protein, lipid and nucleic acids to nearby and distal cells. It has been hypothesized that exosomes may play a role in the ‘prion-like’ spread of misfolded protein pathology in neurodegenerative diseases (ND), including Alzheimer’s, Parkinson’s, and prion diseases. While the majority of studies into the role of exosomes in ND have used in vitro models, the in vivo relevance of these vesicles in ND remains to be determined. We have used the example of transmissible prions to demonstrate that exosomes released from infected neuronal cells release exosomes with distinct structural properties determined with cryo electron microscopy with tomography. We have also shown that exosomes isolated from prion infected cells are capable of transmitting infection to healthy cells and animals.

Here we present a novel protocol for the isolation EVs and enrichment of exosomes from human and mouse frozen brain tissue. The isolation of exosomes from the extracellular space, relies on the use of differential centrifugation and an optimized sucrose gradient has been designed to protect the integrity of the exosome structure and cargo. Brain derived exosome and EVs are rigorously assessed by negative stained, transmission electron microscopy, and the detection of protein and nucleic acid vesicle markers. Furthermore, cryo-transmission electron microscopy allows us to detect internal ultrastructural features that may provide insight into how neurodegenerative diseases affect cellular pathways feeding into exosome biogenesis.

Isolation of exosomes from brain tissue will allow meaningful studies into understanding the role of the propagation of protein misfolding pathology in ND, and the potential discovery of new biomarkers. Rigorous validation of brain derived exosomes, including electron microscopy techniques, can also highlight differences in between EVs from healthy and diseased tissue, providing a potential mechanism to modulate propagation of disease.