改进的瑞氏染色法对人与小鼠淋巴细胞自发凋亡形态的鉴定及其意义

目的以人和小鼠淋巴细胞为研究对象，建立一种在光学显微镜下快速而特异识别凋亡淋巴细胞的染色方法，为淋巴细胞凋亡的研究提供一种简便易行的实验方法。方法收集人或小鼠淋巴细胞制成细胞涂片，晾干，滴加瑞氏染液染1min，再于瑞氏染液上滴加等量的PBS，混匀，染1～2min，水洗晾干后用中性树胶封片，镜检。实验设立实验组（经体外培养的淋巴细胞）和对照组（未经体外培养的新鲜淋巴细胞），动态观察体外淋巴细胞自发凋亡情况。结果人和小鼠淋巴细胞形态基本相似，实验组可见不同程度的细胞凋亡，光镜下细胞呈典型凋亡特征改变：凋亡早期细胞形态不规则，体积变大，然后核染色质边聚、浓缩、细胞核固缩呈半球形、环形、月牙形等，继而细胞裂解成细胞质膜包绕的核碎片，即形成凋亡小体，凋亡细胞体积缩小，而对照组的凋亡细胞偶见。与人淋巴细胞相比较，小鼠淋巴细胞更易发生凋亡。结论本法将瑞氏染色法进行改进，省去了甲醇固定细胞步骤及调整了染色条件等，对淋巴细胞进行染色，在光学显微镜下识别凋亡细胞，具有特异性高，染色方法简便、快速，成本低廉等优点，易于在实验研究和临床推广应用。

Identification of Morphology of Lymphocytes Spontaneous Apoptosis in Human and Mice by Improved Wright Stain Method and Its Significance

Objective To establish a rapid and specific stain assay for identifying the apoptosis of lymphocytes from human and mice under the light microscope which would provide a reliable experimental technique for further study. Methods The lymphocytes were collected from human and mice,and unfixed smears of the cell were prepared. After drying, the smears were exposed to Wright stain solution for 1 min. Then,an equal amount of PBS was added and mixed gently. After 1 to 2 min,the smears were washed with flooding water,drained dry,sealed with neutral gum and observed under a microscope. Cells were divided into two groups. The lymphocytes experimental group were cultured in vitro while those in control group were not. The apoptosis of lymphocytes was observed dynamically. Results The morphology of lymphocytes from human was conformity with that of lymphocytes from mice. Various apoptotic cells were found in experimental group.The distinct morphological feature of apoptotic apoptotic cell showed irregular shapes, volume lymphocytes could be seen. Morphologically, the of anion firstly. Then it was charac clear fragmentation,chromatin condensation which gathered under the nuclear mem circle, ring, crescent, etc Thereafter membrane-bound ap the cell shrunk. Apoptotic cells were not easily observed terized by nu brane in semi optotic bodies formed,and the volume of in the control groups. Compared with human lymphocytes,apoptotic cells in mice were apt to occur. Conclusion Wright stain method may be improved in thisstudy. The step of fixing cells with methanol was omitted, and staining condition was adjusted. This method is specific,simple, rapid and economical for identifying apoptotic lymphocytes under light microscope,and can be applied to experimental research and clinical laboratory.