Human PAUF/ZG16B Antibody (817310)

Detection of Human PAUF/ZG16B by Western Blot.

Western blot shows lysates of human placenta tissue and human prostate tissue. PVDF membrane was probed with 1 µg/mL of Mouse Anti-Human PAUF/ZG16B Monoclonal Antibody (Catalog # MAB7777) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for PAUF/ZG16B at approximately 22 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

PAUF/ZG16B in Human Pancreas Cancer Tissue.

PAUF/ZG16B was detected in immersion fixed paraffin-embedded sections of human pancreas cancer tissue using Mouse Anti-Human PAUF/ZG16B Monoclonal Antibody (Catalog # MAB7777) at 15 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using the Anti-Mouse HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS002) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm and plasma membranes of cancer cells. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of PAUF/ZG16B by Western Blot

Depletion of pancreatic adenocarcinoma upregulated factor (PAUF) inhibits TGF-beta -mediated increase in epithelial–mesenchymal transition (EMT) in Panc-1 cells. Panc-1 cells were transfected with scrambled (Ctrl) or PAUF siRNA and stimulated with or without TGF-beta for (C) 1 or (A,B,D–F) 24 h. (A,B) Successful knockdown of PAUF expression was verified using immunoblotting and sandwich ELISA (n = 4). (C) Phosphorylation levels of MEK1/2 and ERK1/2 were examined using immunoblot analysis. Densitometry of the phosphorylated proteins was performed using the ImageJ software (n = 3). (D) Protein levels of EMT-related signaling molecules were determined using immunoblotting. (E) Expression levels of E-cadherin and alpha SMA were assessed using immunofluorescence staining with Alexa Flour 488-labeled anti-E-cadherin or Alexa Flour 647-labeled anti-alpha SMA antibody and nuclei were stained with DAPI. Scale bar, 50 μm. Relative fluorescence intensity was measured in four fields per sample in a randomized manner using the ImageJ software (n = 4). (F) Changes in cell morphology were observed using an optical microscope, and cell migration and invasion were evaluated using Transwell migration and Matrigel invasion assays. Scale bar, 50 μm. The migrated and invading cells were counted in four fields per sample (n = 4). Statistical significance was calculated using two-way ANOVA followed by post-hoc multiple comparisons test with Bonferroni correction. Data are represented as the mean ± standard deviation (SD). ** p < 0.01, *** p < 0.001. Image collected and cropped by CiteAb from the following open publication (https://www.mdpi.com/1422-0067/25/21/11420), licensed under a CC-BY license. Not internally tested by R&D Systems.