Super-resolution STORM imaging identifies synaptic Nrxn1 nanoclusters. (A) Representative image of cultured neurons at DIV18, showing multiple Homer1(+) synapses with a subset containing Nrxn1 nanoclusters. (B) Representative images of individual synapses from DIV18 neurons showing Homer1(+) synaptic junctions containing Nrxn1 nanoclusters. (C) Only a subset of Homer1(+) synapses contains Nrxn1 nanoclusters (NCs; 39%) in cultured neurons at DIV18. (D) Homer1 discs and Nrxn1 nanoclusters are separated by ∼85 nm across the synapse. The trans-axial distribution of Homer1 and HA-Nrxn1 localizations was determined by STORM imaging of hippocampal cultures at DIV18. (E) Nrxn1 nanoclusters occupy on average ∼19% area of the Homer1-defined synaptic junction. (F) Nrxn1 nanoclusters are on average labeled by approximately nine antibodies per nanocluster. (G–K) Same as A–E, except that cryostat sections of the hippocampal CA1 region from HA-Nrxn1 cKI mice were analyzed at P28. (L) Quantification of the axial distribution of STORM localizations of surface HA-Nrxn1 in hippocampal neurons or cryosections relative to those of Homer1, Piccolo, and Bassoon. Short vertical bars = SEMs; horizontal bars = SDs. For C–F, n = 54 synapses/three cultures; for I–K, n = 77 synapses/three mice. For further details, see Fig. S1.