Anti epcam

A tissue chip, HBla-U060CS-01, which consists of 30 paired tumor and paratumor samples, was purchased from Shanghai Outdo Biotech. To ensure the consistency of the analysis, all immunofluorescence analyses were performed using the same type of tissue chip.
The following antibodies were used to detect specific proteins: anti-HLA-DRA (rabbit, 1:50, Proteintech, Cat. No. 11221-1-AP), anti-EPCAM (rabbit, 1:50, Proteintech, Cat. No. 21050-1-AP), anti-RGS5 (rabbit, 1:50, Proteintech, Cat. No. 11590-1-AP), anti-PDGFRA (rabbit, 1:200, Abcam, ab203491), and anti-CXCL12 (rabbit, 1:100, Abcam, ab155090).

For histological analysis, endometrial tissues were inflated and fixed in 4% Paraformaldehyde Fix Solution at 4 °C for 24 h. After the fixation, tissues were subsequently dehydrated and embedded in paraffin. Each section was obtained at 5 μm, and was subjected to a series processes of hydration for the H&E staining and histopathological analyses. For immunofluorescence analysis, the sections were deparaffinized and rehydrated, and the antigen retrieval was accomplished in citrate buffer (10 mM, pH 6.0). After blocking for 1 h at room temperature with 5% bovine serum albumin, the endometrial sections were incubated with the primary antibodies overnight at 4°C. After removing the primary antibodies, all sections were incubated with the fluorescent secondary antibodies at room temperature for 1 h. The following primary and secondary antibodies were used: anti-LCN2 (proteintech, 1:100), anti-SAA1/2 (Abmart, 1:100), anti-DKK4 (Biorbyt, 1:100), anti-EPCAM (proteintech, clone number 2A2D5, 1:100), anti-Vimentin (abcam, clone number EPR3776, 1:200), goat anti-mouse Alexa Fluor 647 (abcam, 1:200), goat anti-rabbit Cy3 (abbkine,1:200). Images were acquired with a thunder imager DM6B (Leica) and the confocal microscopy (OLYMPUS).

The following anti-mouse antibodies were used: anti-Hai-2 goat polyclonal antibody (pAb) (R & D Systems, Minneapolis, MN); anti-Epcam (ProteinTech., Rosemont, IL), anti-claudin-7 (Thermo Fisher Scientific, Waltham, MA), anti-mouse β-catenin (Sigma, St. Louis, MO), anti-cleaved caspase-3 (Cell Signaling, Boston, MA), and anti-lysozyme (AVIVA Systems Biology, San Diego, CA) rabbit pAbs; anti-Ki67 rabbit monoclonal antibody (mAb) (Abcam, Cambridge, UK); anti-E-cadherin goat pAb (R & D Systems); anti-claudin-2 and anti-ZO-1 rabbit pAbs (Thermo Fisher Scientific); anti-β-actin mouse mAb (Sigma) and anti-ssDNA rabbit pAb (Dako, Carpinteria, CA).