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目前已公认细胞死亡时有两种完全不同的细胞形态学变化.细胞坏死时细胞肿胀,胞内物质溢出,导致周围组织发生炎症.细胞正常死亡时细胞收缩,无炎症反应.后一种死亡称为细胞凋亡(Apoptosis)或称为细胞编程死亡(Programmed Cell Death).近年来对细胞凋亡的研究表明,在细胞进行上述死亡过程中,DNA断裂成约核小体长短的片段,本文介绍的原位缺口平移法(in situ nick translation ISNT)就是检测凋亡细胞中的DNA片段,从而在组织原位显示凋亡细胞.1 材料与方法材料为人神经胶质细胞瘤,手术后将瘤组织固定于4%多聚甲醛和0.3%饱和苦味酸混合液中24h,然后浸入20%蔗糖PBS至组织块下沉.恒冷冰冻切片机切片,厚10μm,切片贴于涂有铬明胶的载片上后,37℃烘箱烘烤过夜,然后再于4%多聚甲醛和0.3%饱和苦味酸混合液中12h.ISNT程序:(1)切片预处理:①0.1mol/L Tris-HCl(pH8.0)缓冲液漂洗3次,每次5min.
It has been recognized that there are two completely different cell morphological changes when cell death. Cell necrosis, cell swelling, intracellular material overflow, resulting in inflammation of the surrounding tissue. Cell death normal cell contraction, no inflammatory response .A late death Apoptosis or Programmed Cell Death In recent years, studies on apoptosis have shown that DNA breaks into fragments of about the length of nucleosomes during the process of cell death. This article describes (In situ nick translation ISNT) is the detection of DNA fragments in apoptotic cells, thus displaying apoptotic cells in situ.1 Materials and Methods The material was human glioma, the tumor tissue Fixed in a mixture of 4% paraformaldehyde and 0.3% saturated picric acid for 24 h, then immersed in 20% sucrose PBS to sink into a tissue block, frozen in a microtome section 10 μm thick and affixed to a slide coated with chromium gelatin After overnight baking at 37 ℃, and then in 4% paraformaldehyde and 0.3% saturated picric acid mixture 12h.ISNT program: (1) slice pretreatment: ① 0.1mol / L Tris-HCl (pH8.0 Rinse buffer 3 times for 5 minutes each.