Figure 2.

Developing a 96-well transwell TNF-α-based barrier damage system. (A) Barrier function was quantified in untreated monolayers (yellow) or monolayers challenged with 40 ng/mL TNF-α (red). At the time of treatment initiation, media on both apical and basolateral sides of the transwell was changed to media containing TNF-α. TEER measurements were taken immediately before challenge and then in 24-h intervals. Each data point represents the average of 8 cell culture replicates. (B) Barrier disruption following 72-h challenge with an 8-point dose curve with TNF-α. Values are reported as %untreated, which was obtained by averaging 5 cell culture replicates for each concentration of TNF-α and normalizing to the average TEER value of untreated controls (4 cell culture replicates). (C) Evaluation of anti-TNF-α antibody for the prevention of barrier damage. Monolayers were pre-treated for 1 h with 8-point dose curves of either anti-TNF-α neutralizing antibody or an isotype-matched control before challenge with 40 ng/mL TNF-α. TEER measurements from 5 wells per condition were taken at 72-h post-challenge and plotted in a similar manner to (B). All data in this figure were collected from cultures grown in 96-well transwells.