Transfection of epithelial cells.

Epithelial cells are found throughout the body, from skin to glandular formations within tissues. In vivo, these cells are attached to a three-dimensional basement membrane matrix. Normal epithelial cells are extremely difficult to transfect- especially when using liposome based transfection reagents. Previously, we only used electroporation (Amaxa) to transfect normal human colonic epithelial cells, and got 65% GFP+ cells. Thanks to SignaGen, we have now successfully transfected normal human colonic epithelial cells using GenJet Ver. II (Cat # SL100489) with up to 75% GFP+ efficiency. The following is a brief protocol for transfecting epithelial cells with GenJet Ver. II (Cat # SL100489):
1. Grow epithelial cell ~85% confluency at time of transfection. Epithelial cells must be freshly prepared and healthy.
2. For a 6-well plate, dilute 1.0 µg of DNA and 3.0 µl of GenJet Ver. II per well with serum free DMEM. Add the diluted reagent to diluted DNA and let the transfection complex form at room temperature for 15 minutes.
3. Add transfection complex to epithelial cells directly. Transfection is conducted in the presence of serum/antibiotics with transfection volume of 1.0 ml per well in a 6-well plate.
4. Remove the transfection complex 5 hours after transfection and change back to normal growth medium.
5. Check transgene expression 24~48 hours post-transfection.