Immunohistochemistry analysis of IL-18 on paraffin-embedded mouse intestine tissue. Antigen retrieval was performed using citrate buffer (pH6, epitope retrieval solution) for 20 mins. Sample was blocked using 10% goat serum, incubated with IL-18 polyclonal antibody (Product# PA5-79481) with a dilution of 1 µg/mL (overnight at 4°C), and followed by biotinylated goat anti-rabbit IgG (30 minutes at 37°C). Development was performed using Streptavidin-Biotin-Complex (SABC) with DAB chromogen method.

Western blot was performed using Anti-IL-18 Polyclonal Antibody (Product # PA5-79481) and a 22 kDa band corresponding to Interleukin-18 was observed in RAW 264.7 and not in Neuro-2a which is reported to be negative. A non-specific band around 85 kDa was also observed in RAW 264.7. Whole cell extracts (30 µg lysate) of RAW 264.7 (Lane 1) and Neuro-2a (Lane 2) were electrophoresed using NuPAGE™ 12% Bis-Tris Protein Gel (Product # NP0342BOX). Resolved proteins were then transferred onto a Nitrocellulose membrane (Product # IB23001) by iBlot® 2 Dry Blotting System (Product # IB21001). The blot was probed with the primary antibody (0.2 µg/mL) and detected by chemiluminescence with Goat anti-Rabbit IgG (Heavy Chain) Superclonal™ Recombinant Secondary Antibody, HRP (Product # A27036, 1:4000 dilution) using the iBright FL 1000 (Product # A32752). Chemiluminescent detection was performed using SuperSignal™ West Dura Extended Duration Substrate (Product # 34076).