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. 2022 Mar 8;13:842023. doi: 10.3389/fimmu.2022.842023

Figure 3.

Figure 3

Time course of C5b-9 puncta formation in Barrett’s esophagus and esophageal adenocarcinoma cell lines after exposure to immunopurified C9. Cell lines representing non-dysplastic BE (BAR-T, CP-A), high-grade dysplasia (CP-B, CP-D) and EAC (FLO-1 and OE-33) were incubated in serum-free medium for 4 hours before being incubated in medium containing C9-depleted human serum for 30 minutes (A), or medium containing C9-depleted human serum plus 3 µg/ml purified human C9 for 0, 30 or 75 minutes (B). Coverslips were removed and stained with C5b-9 antibody and imaged by confocal microscopy. Maximum intensity projection of C5b-9 signal at 0, 30 and 75 min in all cell lines, cell nuclei are represented by blue (DAPI) and C5b-9 by green. Images are all at the same magnification. All images ( Figure S4 ) were used to quantify the average number of puncta per cell (C, D) and the area positive for C5b-9 per cell (E, F) in untreated or cells incubated for 30 minutes in C9 depleted human serum (C, E), or with C9 for 0, 30 or 75 minutes (D, F). *P<.05. **P<0.01. ***P<0.001. ****P<0.0001.