Rabbit αsma

At times of sacrifice, mouse intestines were rinsed in phosphate-buffered saline (PBS) and fixed with 4% paraformaldehyde overnight, rinsed in PBS, and either dehydrated for paraffin embedding or immersed in 30% sucrose for 4 hours at 4°C, embedded in OCT and frozen for cryosectioning. Antigen retrieval was achieved using citrate buffer, pH 6.0, with a pressure cooker (PickCell Laboratories). Foxl1 staining required the use of cryosections ,antigen retrieval, and amplification of signal using tyramide (TSA systems PerkinElmer). Antibodies used were: Ki67 (BD Pharmingen 1:500), Lysozyme (Dako 1:1000), Sox9 (Millipore 1:300), Mucin 2, HBEGF (Santa Cruz Biotechnology 1:50) Guinea Pig Foxl1 (1:1500), Goat GFP (Abcam 1:200), Mouse E-cadherin (BD transduction 1:250), Mouse β catenin (BD transduction 1:200), Rabbit EpCAM (Abcam 1:100), Rabbit αSMA (Abcam 1:100), Rabbit Myh11 (Abcam 1:100). Cy2-, Cy3- and Cy5-conjugated donkey secondary antibodies were purchased from Jackson ImmunoResearch Laboratories, Inc. For immunohistochemistry, horseradish peroxidase (HRP)-conjugated secondary antibodies were incubated at room temperature for 2 hours. VECTASTAIN ABC kit (Vector Laboratories) was then used to detect the signal, and the slides were washed with PBS. For signal development, the DAB substrate was used.