Single and dual labeling of cells and tissue sections by immunofluorescence (IF) or immunohistochemistry (IHC) was performed according to previously described methods [16] [17] . The primary antibodies were: goat polyclonal anti-CCSP(T18) (1:50, Santa Cruz Biotech, Santa Cruz, CA), rabbit antiuteroglobin-related protein 1 (UGRP1) (1:100, a kind gift from Dr. Shioko Kimura, NCI/NIH, Bethesda, MD), rabbit anti-pan-cadherin (, 1:50, Abcam, Cambridge, MA), mouse anti-b-catenin (BD). rabbit anti-pan-cytokeratin (1:100, Dako, Carpinteria, CA), rabbit anti-pro-SPC (1:200, Millipore), rat anti-BrdU (1:100, Accurate Chemical & Scientific Corp, Westbury, NY), rabbit anti-sox2(1:2000, Seven Hill, Cincinnati, OH) and rabbit anti-ALDH1(1:500. Abcam). Approximately 1610 5 FACS sorted cells, were washed twice in PBS with 1% FBS and resuspended in 30 ml of Cell Adherence Solution (Crystalgen, Commack, NY). After standing for 2 minutes, 3 ml of the cell mixture was mounted on glass slides, dried for 2 minutes and fixed with 4% paraformaldehyde in PBS for 15 minutes. Both fixed cells and tissue sections were blocked with 1% goat or rabbit normal serum for 1 hour. The blocking solution was removed and 75 ml of primary antibody was added to cells. After 1 hour of incubation at room temperature, slides were washed in PBS three times. Secondary antibodies were added. For dual-labeling IF, additional primary antibodies were added after the third PBS wash, followed by incubation with secondary antibodies conjugated with Alexa fluor 488 or 594 (Invitrogen). All incubations were performed at room temperature and slides were washed in PBS (365 min) between each step and mounted with an anti-fading reagent with 49,6-diamidino-2-phenylindole (DAPI) (Invitrogen). Control slides were included in each analysis in which non-immune serum was substituted for primary antibodies and secondary antibodies individually. All IF images were taken with a Zeiss LSM 510 Meta Mk4 Confocal Microscope (Zeiss, Thornwood, New York). For IHC, signals were developed using 3,39diaminobenzidine (DAB).