T84 human colon intestinal epithelial cells, used as a model to study the effects of enterotoxins on TJ proteins [29] (link), [30] (link), [39] (link)–[42] (link) were obtained from the American Type Culture Collection. Cells (passage 5 to 18) were maintained in equal volumes of Dulbecco’s Modified Eagle Medium (DMEM) and F-12 supplemented with 5% (vol/vol) fetal bovine serum (FBS) (Invitrogen). Cells were grown in T-75 culture flasks (Sarstedt) at 37°C, 5% CO2 in a humidified incubator. Cell viability of T84 cell line was measured using Trypan blue [43] . For maintenance purposes, confluent T84 monolayers were passaged weekly using trypsin-EDTA treatment in phosphate buffered saline (PBS: Invitrogen) free of calcium chloride and magnesium chloride.
For immunofluorescence experiments, T84 cells were seeded, on LabTek 8-well chamber slides (Fisher Scientific), at a density of 150,000 cells/ml and used after 2 days of growth. One hour before treatment with purified STb cell monolayers was washed using PBS and the medium was changed to medium without FBS.
For immunofluorescence experiments, T84 cells were seeded, on LabTek 8-well chamber slides (Fisher Scientific), at a density of 150,000 cells/ml and used after 2 days of growth. One hour before treatment with purified STb cell monolayers was washed using PBS and the medium was changed to medium without FBS.