FIGURE 1.

Purification of ACVs from whole mouse brain. (A) Purification protocol for isolating ACVs. At variance to all previous methods, the final step of the preparation involves elution with a GluA1 peptide that corresponds to the epitope of the GluA1 monoclonal antibody. Note that the same amount of protein (as assessed by BCA) was inputted into the same immunoisolation step for both wild-type and knockout preparations. (B) Western blots for GluA1 and VGLUT1 in WT isolated synaptosome content (LP2-WT), GluA1 KO isolated synaptosome content (LP2-KO), GluA1 peptide eluate from wild-type mice (E-WT), and GluA1 peptide eluate from GluA1 KO mice (E-KO). Original blots provided in Supplementary Figure 1. (C) Negative stain electron microscopy image of GluA1 peptide eluate of wild-type mice. Additional images are shown in Supplementary Figure 2. All images are provided as Source Data. (D) Histogram of vesicle diameters from wild-type eluate from three independent immunoisolations. (E) Negative stain electron microscopy image of GluA1 peptide eluate of GluA1 KO mice. Additional images are shown in Supplementary Figure 3. Additional images are provided in Supplementary Material. (F) Histogram of vesicle diameters from knockout eluate from three independent immunoisolations. (G) Representative western blots from three independent immunoisolations for GluA1 (102 kD), GluN1 (115 kD), PSD-95 (98 kD), LAMP1 (130 kD), and Golgin (100 kD) for homogenized whole brain pellet (HP), the second synaptosome wash step (SW), synaptosome (P3), synaptosome content (LP2), beads from immunoisolation prior to elution (B), flowthrough from immunoisolation (FT), and GluA1-peptide eluate off beads (E) for wild-type mice. Original blots provided in Supplementary Figure 4.