Ab236494

Aortic specimens from the experimental group and control group mice were stored in liquid nitrogen for Western blot assays to evaluate the expression of CD11b and MMP. Protein from the aortic specimens was extracted with radio immunoprecipitation assay (P0013B, Beyotime) and phenylmethylsulfonyl fluoride (ST506, Beyotime), and then measured with the bicinchoninic acid assay kit (P0012S, Beyotime). The lysates containing 40 μg protein were treated by SDS-PAGE and separated by electrophoresis. The isolated proteins were transferred to a polyvinylidene fluoride (PVDF) membrane. Then, the PVDF membrane was blocked in tris buffered saline with Tween (TBST) containing 5% skim milk, and a primary antibody solution was prepared with TBST containing 3% BSA. The membrane was incubated overnight at 4°C with one of the following primary antibodies each time: anti-CD11b (EPR1344, Abcam), anti-MMP-9 (ab236494, Abcam), and anti-β-actin (ab8227, Abcam) antibodies. Then, it was washed three times with TBST, incubated at room temperature for 1 h with secondary antibody (diluted 1:5000), washed three times with TBST again, and exposed.