Pon3 polyclonal antibody

Tissue microarray (TMA) containing 286 pairs of HCC and the adjacent non-tumor tissues was constructed in collaboration with the Shanghai Outdo Biotech Company (Shanghai, China). Immunohistochemistry was performed as previously described [16 (link)]. A PON3 polyclonal antibody (Abcam, MA, USA) diluted at 1:300 was used as the primary antibody.
Density of PON3-positive staining was quantified using a DFC420 CCD camera connected to a DM IRE2 microscope (Leica Microsystems Imaging Solutions Ltd, Cambridge, United Kingdom). Photographs of representative fields were captured under a high-power magnification (200×) using the Leica QWin Plus v3 software. The Image-Pro Plus v6.0 software (Media Cybernetics Inc, MD, USA) was used to count the integrated optical density (IOD) of each photograph as previously described [16 (link)], and the ratio of IOD to total tissue area (AREA) of each photograph was calculated as PON3 density.

Western-blot was performed as described previously [35 (link)]. Antibody dilutions were 1:1,000 for the PON3 polyclonal antibody (Abcam, MA, USA) and 1:5,000 for β-actin (Sigma-Aldrich, USA). Apoptosis antibodies were purchased from Cell Signaling Technology (#9915; MA, USA) and diluted at 1:1,000. Cell cycle antibodies (CCNB1, CCND1, p27, p57) were purchased from AbSci (MD, USA) and diluted as the manufacturer's instruction.