Separation of Single Core and Multicore Lytic Granules by Subcellular Fractionation and Immunoisolation.
Methods Mol Biol
; 2654: 159-167, 2023.
Article
in En
| MEDLINE
| ID: mdl-37106182
ABSTRACT
Subcellular fractionation is an important tool used to separate intracellular organelles, structures or proteins. Here, we describe a stepwise protocol to isolate two types of lytic granules, multicore (MCG), and single core (SCG), from primary murine CTLs. We used cell disruption by nitrogen cavitation followed by separation of organelles via discontinuous sucrose density gradient centrifugation. Immunoisolation with a Synaptobrevin 2 antibody attached to magnetic beads was then used to harvest Synaptobrevin 2 positive granules for immunoblotting, mass spectrometry, electron, and light microscopy.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Proteins
/
Vesicle-Associated Membrane Protein 2
Limits:
Animals
Language:
En
Journal:
Methods Mol Biol
Journal subject:
BIOLOGIA MOLECULAR
Year:
2023
Document type:
Article
Affiliation country:
Alemania