May 25, 2022

Public workspaceGUV preparation

 Forked from GUV preparation and assay
DOI
dx.doi.org/10.17504/protocols.io.3byl4b398vo5/v1
  • 1Hurley Lab, University of California, Berkeley;
  • 2Team Hurley, University of California, Berkeley
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DOI: dx.doi.org/10.17504/protocols.io.3byl4b398vo5/v1
Protocol Citation: Liv Jensen, Chunmei Chang 2022. GUV preparation. protocols.io https://dx.doi.org/10.17504/protocols.io.3byl4b398vo5/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: May 02, 2022
Last Modified: May 31, 2024
Protocol Integer ID: 61817
Keywords: ASAPCRN
Abstract
GUV preparation for membrane tube assay application
GUV Preparation
Clean the coverglass.
Coat cleaned coverslips with 100 μL 5% (w/w) polyvinyl alcohol (PVA) with a molecular weight of 145,000 (Millipore).
Place the coated coverslip in a heating incubator at 60 °C to dry the PVA film for 30 min.
Spread a lipid mixture with a molar composition of 70% DOPC, 20% DOPE, 5% DO-PI(3)P, 5% DOPS, 0.3% Atto647N DOPE, 0.01% PEG2000-biotin-DSPE at 1 mg/ml uniformly onto the PVA film.
Put the lipid-coated coverslip under vacuum overnight to evaporate the solvent.
Use 100 μL 320 mOsm sucrose solution for swelling for 1 h at room temperature
Harvest the GUVs and use them with 12 h.