Fig. 3.

Proximity ligation assay (PLA) supports a TSPO interaction with gp91^phox, p22^phox, and VDAC in primary microglia. a–c Representative PLA phase and pseudo-colored confocal images of protein pair interactions (colored dots) in microglia: TSPO + gp91 (a); TSPO + p22 (b); TSPO + VDAC (c). The images indicate that TSPO interacts with gp91^phox, p22^phox, and VDAC respectively supporting the co-immunoprecipitation and immunocytochemistry colocalization results. d Quantification of average number of protein pair signals per microglia in vehicle conditions at 18 h. PLA experiments confirms that TSPO interacts with the NOX2 subunits gp91^phox and p22^phox, as well as the mitochondrial protein, VDAC. Imaging and quantification included negative controls for labeling conditions of single antibody only, as well as no primary antibody, to ensure signal specificity. Data are expressed as mean ± s.e.m. n = 4–7 independent experiments with > 30 cells counted per treatment and per labeling condition. One-way ANOVA across labeling conditions was performed; (TSPO+gp91: F_3,17 = 8.193, p = 0.002; TSPO+p22: F_3,11 = 11.108, p = 0.003; TSPO+VDAC: F_3,14 = 9.979, p = 0.002). *p < 0.05 = significantly different relative to all other conditions within a protein pair group