Anti h pgds

Pam3CSK4 (TLR1/2) and LPS (TLR4) were obtained from InvivoGen (Carlsbad, CA, USA). Mouse recombinant IFN-β was purchased from PBL InterferonSource (now PBL Assay Science, Piscataway, NJ, USA) and mouse IFN-γ was from R&D Systems (Minneapolis, MN, USA). Antibodies used in this study were obtained as follows. Anti-H-PGDS and Anti-COX-2 antibodies were produced by Cayman Chemical (Ann Arbor, MI, USA). Anti-STAT1 and anti-pY701-STAT1 antibodies were from Cell Signaling Technology (Danvers, MA, USA). Anti-actin antibody was from BD Biosciences (Franklin Lakes, NJ, USA). PGs were purchased from Cayman Chemical (Ann Arbor, MI, USA). All other chemicals were from Sigma (St. Louis, MO, USA), unless stated otherwise.

After deparaffinization, the sections were antigen-activated with Histo VT One (Nacalai Tesque) and blocked using Blocking One Histo (Nacalai Tesque). Primary antibody reactions were performed with anti-HPGDS (Cayman Chemicals, Ann Arbor, MI, USA) at a dilution ratio of 1:2000 in Solution A (Toyobo) and anti-Tryptase (ab2378: Abcam), anti-NG2 (sc-80003: Santa Cruz Biotechnology, Inc., Dallas, TX, USA), anti-CD11b (66519-1-Ig: Proteintech Group, Inc., Rosemont, IL, USA), anti-Troponin T (MAB18742: Bio-Techne Corp., Minneapolis, MN, USA), anti-α-SMA (sc-53142: Santa Cruz Biotechnology, Inc.), and anti-CD31 (sc-376764: Santa Cruz Biotechnology, Inc.) at a dilution ratio of 1:500 in Solution A overnight at 4 °C. Secondary antibodies were reacted with goat anti-rabbit IgG H&L (Alexa Fluor^® 647) (ab150083: Abcam) for anti-HPGDS and goat anti-mouse IgG H&L (Alexa Fluor^® 488) (ab150113: Abcam) for others in Solution A (Toyobo) (dilution 1:1200) for 1 h at room temperature. Sections were mounted with Vectashield Hard Set Mounting Medium with DAPI (H-1500: Vector Laboratories, Inc., Newark, CA, USA). To confirm the specificity of the antibodies, we also verified that no fluorescence was observed in the negative control without the primary antibody.