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1 材料和方法1.1 材料①细胞:LoVo 人大肠癌细胞由第一军医大学消化研究所中心实验室提供;人脐静脉内皮细胞由本室按杨小平法传代培养;②抗体:小鼠抗人 VEGF(IgGl)为美国 Neuwork公司产品;小鼠抗人 KDR 抗体(IgG1)为美国 Sigma 公司产品.1.2 方法细胞培养及细胞毒性实验:将冻存的 LoVo 细胞进行复苏及传代,并制成细胞悬液,加入96孔培养板中,1×10~4/孔.同法加入传代培养的人脐静脉内皮细胞.将培养板移入 CO_2孵箱中于37℃,50 mL/L CO_2及饱和湿度条件下培养.培养1 d后按表1及表2分别加入不同浓度的抗 VEGF抗体及抗 KDR 抗体,每个浓度设6个复孔,并设对照孔(未加
1 Materials and methods 1.1 Materials 1 cells: LoVo human colorectal cancer cells were provided by the Central Laboratory of Digestive Research Institute, First Military Medical University; human umbilical vein endothelial cells were subcultured by our laboratory according to Yang Xiaoping; 2 Antibodies: mouse anti-human VEGF (IgG1) ) is a product of the United States Neuwork Corporation; mouse anti-human KDR antibody (IgG1) is a product of Sigma, USA. 1.2 Methods Cell Culture and Cytotoxicity Test: Frozen LoVo cells are resuscitated and passaged, and a cell suspension is prepared. In a 96-well culture plate, 1×10 4 /well was added. The cultured human umbilical vein endothelial cells were added in the same manner. The culture plates were transferred to a CO 2 incubator and incubated at 37° C., 50 mL/L CO 2 and saturated humidity. After 1 day of culture, add different concentrations of anti-VEGF antibody and anti-KDR antibody according to Table 1 and Table 2, set 6 replicates for each concentration, and set control wells (without