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【文獻標題】Silencing ICAM-1 reduces the adhesion of vascular endothelial cells in mice with immunologic contact urticaria

【作者】Gaomei Lv, Jingang Fan

【作者單位】臨沂市人民醫(yī)院（Linyi People’s Hospital）

【文獻中引用產(chǎn)品】

生物素標記的羊抗兔抗體

【關(guān)鍵詞】Intercellular cell adhesion molecule-1; RNA inference; Immunologic contact urticaria; Vascular endothelial cells; Adhesion molecule; Cell-cell adhesion

【DOI】doi.org/10.1016/j.gene.2020.144965

【影響因子(IF)】4.80

【出版期刊】《Gene》

【產(chǎn)品原文引用】

Immunohistochemistry (IHC)

Fourteen days after model establishment, the vascular endothelium specimens were collected from executed normal and ICU mice (4 mice each group). The specimens were fixed in 4% paraformaldehyde for 24 h and then dehydrated with 80%, 90%, and 100% gradient ethanol and nbutyl alcohol before paraffin embedding at 60oC. Paraffin-embedded specimens were cut into 5 μmserial sections. Xylene dewaxing was performed after the specimens were paved at 45oC and later baked at 60oC for 1 h. The specimens were dehydrated with gradient ethanol and soaked in 3% H2O2 solution for 10 min, followed by 90-s high pressure antigen repair. Then, the specimens were cooled to room temperature. After that, the specimens were added with 100 μL of 5% bovine serum albumin (BSA) and received 30-min incubation at 37oC. The specimens were added with 100 μL primary rabbit anti mouse monoclonal antibodies (Abcam Inc., Cambridge, UK) to ICAM-1 (ab93952, 1 : 1000 - 1 : 4000) or CD31 (1 : 1000 - 1 : 4000, ab2836), and then overnight incubation was performed at 4oC. The 30-min specimen incubation was operated with biotin labeled goat anti rabbit secondary antibody working fluid (HY90046, 1 : 100; Shanghai Hengyuan Science & Technology Co., Ltd., Shanghai, China) at 37oC. Subsequently, the specimens were sliced, followed by 30-min culture with streptomycin-avidin-biotin-peroxidase complex (Beijing Zhongshan Biotechnology Co., Ltd., Beijing, China) at 37oC. The specimens were developed by 3,3′-diaminobenzidine (Beijing Bioss Biotechnology Co., Ltd., Beijing, China) at room temperature before 5-min staining with hematoxylin and 4-s soaking in 1% hydrochloric acid alcohol. The average OD value of the ICAM-1 positive staining was assessed using an Image-Proplus software (Media Cybernetics, Inc., Silver Spring, MD, USA) at high magnification. The ratio of ICAM-1 positive cells to total cells was calculated by randomly selecting 5 high-power visual fields (× 400) with 100 cells in each field from each section (Xiong et al., 2012).

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