E-GEOD-10070 - Gene Expression in MCF10A cells through Differentiation on Transwells
Released on 11 June 2008, last updated on 14 August 2013
To further understand the differences occurring in MCF10A cells as they polarize and differentiate in the Transwell® model, we performed gene expression profiling with Affymetrix Human Genome U133 Plus 2.0 Arrays. Four experimental time points, were sampled: conventional cultures of MCF10A cells grown on plastic (Monolayer) and MCF10A cells plated on Transwells® sampled at three TEER values, 200-300 Ω cm2 (Base), 1400-1600 Ω cm2 (Midpoint), and 3000-3200 Ω cm2 (Plateau). Experiment Overall Design: Cells are grown in monolayer to 90-95% confluency, trypsinized and counted for seeding onto permeable supports (Transwell®, 0.4 µm pores, polyester) in normal growth medium. MCF10A cells are seeded on 12-well Transwells® (Corning) at 105 cells/cm2. Both chambers of media were changed strictly on a 24-hour schedule. Transepithelial electrical resistance (TEER) is measured daily with Epithelial Volt-Ohm Meter (EVOM; World Precision Instruments), prior to media change. Total RNA was isolated at the indicated TEER listed above. Canonical monolayer MCF10A cells served as a control comparison. Each time point was performed on triplicate arrays except for Plateau (n=4).
transcription profiling by array
In vitro multipotent differentiation and barrier function of a human mammary epithelium. Marshall AM, Pai VP, Sartor MA, Horseman ND.