Gb11017b

Marc-145 cells or pig tissues were lysed in RIPA buffer containing 1% (v/v) phenylmethylsulfonyl fluoride (PMSF) (Beyotime, Jiangsu, China). The antibodies and their dilutions were shown as follows, anti-CEBPB (GTX100675, GeneTex, Alton Pkwy Irvine, CA, USA, 1:1000), anti-PRRS virus Nucleocapsid (GTX129270, GeneTex, Alton Pkwy Irvine, CA, USA, 1:1000), anti-β-tubulin (GB11017B, Servicebio, Wuhan, China, 1:1000), anti-β-actin (AC026, ABclonal, Wuhan, China, 1:20 000). The protein expression levels were normalized to corresponding β-actin or β-tubulin. ImageJ software was utilized for the quantitative analysis of Western blot results.

Marc-145 cells and PAMs were collected and lysed using RIPA lysis buffer supplemented with fresh protease and phosphatase inhibitors. The protein concentration was determined by the BCA Protein Assay kit (Solarbio, Beijing, China). Equal amounts of each sample were loaded and subjected to SDS–PAGE, transferred onto PVDF membranes (Millipore, Darmstadt, Germany), and then incubated for 12 h at 4°C with the following primary antibodies: IL6 (sc-323975, Santa Cruz, USA), PRRSV nucleocapsid protein (GTX129270, GeneTex, Alton Pkwy Irvine, CA, USA), NF-κB p65 (A11201, Abclonal, Wuhan, China), ARID3A (A7668, Abclonal, Wuhan, China), β-actin (AC026, Abclonal, Wuhan, China), anti-β-tubulin (GB11017B, Servicebio, Wuhan, China), and H3 (A2348, Abclonal, Wuhan, China). After incubation, the PVDF membrane was washed with Tris-buffered saline Tween 20 (TBST) three times and then incubated with HRP-conjugated secondary antibodies (G1213, Servicebio, Wuhan, China). The protein on this membrane was visualized using enhanced chemiluminescence (ECL) (Servicebio, Wuhan, China) in a Western Blotting Detection System.