Figure 11. Specific anti-SUMO1 immunolabelling in His6-HA-SUMO1 KI neurons permeabilised with 10 μg/ml digitonin does not correspond with synaptic boutons.
Hippocampal neurons from either His6-HA-SUMO1 KI mice or SUMO1 KO mice were fixed in 4% PFA and permeabilised using 10 μg/ml digitonin. Primary antibodies were 1:50 mouse anti-SUMO1 (green), 1:2000 rabbit anti-synapsin 1/2 (red), and 1:500 chicken anti-Map2 (blue). In His6-HA-SUMO1 KI neurons anti-SUMO1 labelling was generally absent in nuclei (arrows) as well as in SUMO1 KO neurons. Additional anti-SUMO1 labelling was observed similarly in both His6-HA-SUMO1 KI and SUMO1 KO samples, which did not appear to correspond with synapsin-positive puncta (arrowheads). Permeabilisation with 10 μg/ml digitonin resulted in incomplete immunolabelling with Map2, with dendrites appearing 'fragmented' (indicated by stars). Data are representative of 5 independent experiments. Scale bars, 10 μm.