Fig. 2
PLIC quantitative analysis of protein oligomerization in mTECs. a A graphical scheme illustrating PLIC analysis of Aire oligomerization using a single monoclonal antibody. b PLIC analysis showing representative images of the oligomerization of Aire in mTEChi and CD45+ populations, presented as (left to right): bright field (BF), PLA (green), staining of CD80 (PB), EpCAM (APC), CD45 (APC/Cy7), and BF/PLA/EpCAM overlay (merged). c Quantitative analysis of Aire oligomerization (presented as the percentage of PLA+ cells) in mTEChi or CD45+ cells isolated from Aire +/+ and Aire –/– mice, compared to probes background. Shown are values from three independent experiments. Each point on the graph (triangle for mTEChi, circle for CD45+, cross for probes only) represents an average value of single independent experiment, calculated from 2 to 4 biological replicates, +s.e.m. d Representative images of Aire oligomerization as detected by PLIC in mTEChi isolated from Aire –/– mice, prior to corrected analysis gating and post gating (filtering out false-positive signal). Images are presented as (left to right): bright field (BF), PLA (green), staining of CD80 (PB), EpCAM (APC), CD45 (APC/Cy7), and BF/PLA/EpCAM overlay (merged). Scale bar 7 μM. e Quantitative analysis of Aire oligomerization (presented as the percentage of PLA+ cells) in mTEChi cells isolated from Aire +/+ (n = 2, s.e.m) or Aire –/– (n = 1) mice, compared to probes background sample (n = 1), prior to (light grey) and post (dark grey) corrected gating
