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. Author manuscript; available in PMC: 2019 Nov 21.
Published in final edited form as: Methods Mol Biol. 2019;1860:237–249. doi: 10.1007/978-1-4939-8760-3_15

Figure 3. Trans-SNARE assembly assay.

Figure 3.

Reconstituted t- and v-SNARE liposomes were incubated at 4 °C for the indicated time periods in the presence or absence of 5 μM Munc18-1 before 10-fold excess amount of inhibitory VAMP2 CD was added to block unpaired t-SNAREs. The liposomes were subsequently solubilized and the t-SNAREs were precipitated using nickel sepharose beads. Presence of FL VAMP2 in the precipitates was probed by immunoblotting, which was used as an indicator for trans-SNARE assembly between liposomes. The reactions were performed in the presence of 100 mg/mL Ficoll 70. Adapted with permission from reference (33). Copyright 2015 American Chemical Society.