Rapid and Gentle Immunopurification of Brain Synaptic Vesicles.
biochemistry
cryo-electron microscopy
lipid metabolism
neurodegeneration
proteomics
synaptic vesicles
Journal
The Journal of neuroscience : the official journal of the Society for Neuroscience
ISSN: 1529-2401
Titre abrégé: J Neurosci
Pays: United States
ID NLM: 8102140
Informations de publication
Date de publication:
27 04 2022
27 04 2022
Historique:
received:
23
12
2021
revised:
09
02
2022
accepted:
19
02
2022
pubmed:
18
3
2022
medline:
30
4
2022
entrez:
17
3
2022
Statut:
ppublish
Résumé
Current methods to isolate synaptic vesicles (SVs), the organellar quanta of synaptic transmission, require highly specialized materials and up to 24 h. These technical obstacles have thus far limited the study of SVs in models of synaptic function and pathophysiology. Here, we describe techniques for the rapid isolation of SVs by immunoprecipitation with widely available antibodies conjugated to magnetic beads. We report that the inexpensive rho1D4 monoclonal antibody binds SVs and show that elution with the 1D4 peptide yields native vesicles that are ≥ 10-fold purer than those obtained with classical techniques. These methods substantially widen the accessibility of SVs, enabling their purification in 60-90 min for downstream analyses including mass spectrometry and cryo-electron microscopy. Immunopurified SV preparations from mouse brain contained apolipoprotein E, the LDL receptor Lrp1, and enzymes involved in lipid metabolism, suggesting that SVs may play direct roles in lipid homeostasis and lipoprotein trafficking at the nerve terminal.
Identifiants
pubmed: 35296545
pii: JNEUROSCI.2521-21.2022
doi: 10.1523/JNEUROSCI.2521-21.2022
pmc: PMC9053850
doi:
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Langues
eng
Sous-ensembles de citation
IM
Pagination
3512-3522Subventions
Organisme : NIMH NIH HHS
ID : F30 MH116580
Pays : United States
Organisme : NIGMS NIH HHS
ID : P41 GM108538
Pays : United States
Organisme : NINDS NIH HHS
ID : R35 NS097362
Pays : United States
Organisme : NIGMS NIH HHS
ID : T32 GM140935
Pays : United States
Organisme : NIMH NIH HHS
ID : R01 MH061876
Pays : United States
Informations de copyright
Copyright © 2022 Bradberry et al.
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