剪碎法及匀浆法制备兔肌肉VX2肿瘤单细胞悬液的方法比较  

Preparation of single cell suspension from rabbit muscle VX2 tumor with scissors versus homogenate method

作　　者：高斌[1] 贺克武[1] 李嘉嘉[2] 

机构地区：[1]合肥市第一人民医院影像中心,安徽省合肥市230061 [2]安徽医科大学第一附属医院中心实验室,安徽省合肥市230032

出　　处：《中国组织工程研究与临床康复》2007年第41期8315-8317,共3页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：合肥市科技局重点资助项目(2006004)~~

摘　　要：目的:比较剪碎法和匀浆法制备兔肌肉VX2肿瘤单细胞悬液的效果。方法:实验于2006-10-25/2007-04-01在安徽医科大学第一附属医院中心实验室进行。切取兔肌肉VX2肿瘤组织块,分别采用剪碎法和匀浆法制备兔肌肉VX2肿瘤单细胞悬液。经苏木精-伊红染色,光学显微镜观察细胞形态学差异,并行AnnexinV/PI双染法,流式细胞仪分析细胞凋亡率和细胞坏死率的差异。结果:①细胞形态:匀浆法细胞产率高,细胞形态完整,细胞分布均匀,细胞碎片及细胞团较少;而剪碎法所得细胞悬液在光镜视野中有较多的杂质和细胞团,完整的细胞相对较少。②流式细胞仪分析结果:匀浆法制备的单细胞悬液左下象限正常细胞分布较集中,右下象限凋亡细胞较少,左上象限坏死细胞及细胞碎片较少;所得细胞凋亡率和细胞坏死率值明显低于剪碎法[细胞凋亡率:(8.90±1.24)%,(15.75±1.53)%;细胞坏死率(35.22±1.56)%,(40.16±1.68)%,P均<0.01]。结论:用匀浆法制备兔肌肉VX2肿瘤单细胞悬液优于剪碎法。AIM： To compare the effect of processing single cell suspension with rabbit muscle VX2 tumor by scissors and homogenate methods. METHODS： The experiment was performed in the Central Laboratory of First Affiliated Hospital of Anhui Medical University from October 25^th 2006 to April 1= 2007. A piece of rabbit muscle VX2 tumor tissue was harvested to prepare the single cell suspension by using scissors or homogenate making machine. The cell morphology was assessed by optical microscope after HE staining, and the cell apoptotic rate and necrotic rate were analyzed by flow cytometry after Annexin V/PI double staining. RESULTS： （1）By the homogenate making machine preparation, higher single cell output rate, more intact cells, litter cell fragments and litter cell conglomeration were found in cell suspension, but many foreign substance and cell groups were found in cell suspension by scissor-cut method. （2）In the cell suspension prepared by homogenate method, normal cells mainly distributed in left inferior quadrant, few apoptotic cells were found in right inferior quadrant, and necrotic cell and cell fragments were few in left superior quadrant; moreover, the apoptotic rate and necrotic rate were significantly lower than that by the scissor-cut method [apoptotic rate： （8.90±1.24）%, （15.75±1.53）%; necrotic rate （35.22±1.56）%, （40.16± 1.68）%, P〈 0.01]. CONCLUSION： Homogenate method is preferable to prepare single cell suspension from rabbit muscle VX2 tumor compared with scissor-cut method.

关 键 词：单细胞悬液 流式细胞术 ANNEXIN V/PI双染法 VX2肿瘤 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]