Pab8798

An IHC analysis was conducted as previously described [10 (link)]. Deparaffinized tissue sections were treated with 3% hydrogen peroxide in methanol for 10 min to remove endogenous peroxidase. Antigen retrieval was carried out with sodium citrate buffer (0.1 M) using the microwave method. The slides were incubated with normal serum to block nonspecific binding and then incubated for 1 h with primary antibodies (diluted 1:100 to 1:200), such as antibodies against IFN-γ (sc-74104; Santa Cruz Biotechnology, Dallas, TX, USA), TNF-α (MBS175453; MyBioSource, San Diego, CA, USA), TGF-β (MBS462142; MyBioSource), IL-8 (ab110727; Abcam, Cambridge, UK), CCL-2 (PAB16617; Abnova, Taipei, Taiwan), CXCL-1 (PAB8798,; Abnova), CXCL-9 (bs-2551R; Bioss, Woburn, MA, USA), CXCL-10 (bs-1502R; Bioss), and CXCL-11 (bs-2552R; Bioss). The slides were incubated for 10 min with biotinylated secondary antibodies (PK-7800; Vector Laboratories, Burlingame, CA, USA) and horseradish peroxidase-conjugated streptavidin. Signals were detected using the 3,3-diaminobenzidine tetrahydrochloride substrate chromogen solution, and the cells were counterstained with Mayer's hematoxylin. In order to evaluate the expression levels, after 5 circles were drawn with equal diameters in separate areas, without overlap, the positive cells were counted from 4 slides per group.

IHC analysis was conducted as previously described (Lee et al. 2018 (link)). Deparaffinized tissue sections were treated with 3% hydrogen peroxide in methanol for 10 min in order to remove endogenous peroxidase. Antigen retrieval was carried out with sodium citrate buffer (0.1 M) using the microwave method. Next, the slides were incubated with normal serum so as to block non-specific binding, then incubated for 1 h with primary antibodies (diluted 1:100 to 1:200) such as TNF-α (MyBioSource, San Diego, CA), IFN-γ (sc-74104, Santa Cruz Biotechnology, Dallas, TX), TGF-β (MBS462142, MyBioSource, San Diego, CA), CXCL1 (PAB8798, Abnova, Taipei, Taiwan), CCL-2 (PAB16617, Abnova, Taipei, Taiwan) and CCR2 (Biorbyt, orb137093, Cambridge, UK). The slides were incubated for 10 min with biotinylated secondary antibodies (PK-7800, Vector Laboratories, Burlingame, CA) and horseradish peroxidase-conjugated streptavidin. Finally, signals were detected using the 3,3-diaminobenzidine tetrahydrochloride substrate chromogen solution, and the cells were counterstained with Mayer’s haematoxylin.