解剖学报

解剖学报 ›› 2019, Vol. 50 ›› Issue (3): 392-394.doi: 10.16098/j.issn.0529-1356.2019.03.021

• 技术方法 • 上一篇    下一篇

不同固定液对肿瘤细胞形态及细胞膜通透性的影响

王小杰1 张玉祥2*   

  1. 1.承德医学院基础医学研究所，河北 承德 067000; 2.首都医科大学生物化学与分子生物学系，北京 100069
  • 收稿日期:2018-08-06 修回日期:2019-01-21 出版日期:2019-06-06 发布日期:2019-06-06
  • 通讯作者: 张玉祥 E-mail:yxzhang@ccmu.edu.cn

Effects of different fixing agents on tumor cell morphology and membrane permeability

WANG Xiao-jie1 ZHANG Yu-xiang 2*   

  1. 1.Institute of Basic Medical Sciences, Chengde Medical University, Hebei Chengde 067000, China; 2.Department of Biochemistry and Molecular Biology, Capital Medical University, Beijing 100069, China
  • Received:2018-08-06 Revised:2019-01-21 Online:2019-06-06 Published:2019-06-06
  • Contact: ZHANG Yu-xiang E-mail:yxzhang@ccmu.edu.cn

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摘要：

目的 观察不同固定液对人胰腺腺泡上皮癌（HPAC）细胞及人宫颈癌HeLa细胞形态及细胞膜通透性的影响。 方法 采用2种固定液对HPAC细胞及人宫颈癌HeLa细胞进行固定：新鲜配制0.25%和4%多聚甲醛溶液；75%和90%乙醇溶液；固定时间均为30 min。以只加完全培养基的细胞作为对照组。光学显微镜下观察各组细胞形态，采用7-氨基放线菌素（7-AAD）荧光染色法观察细胞膜通透性的变化。 结果 与完全培养基组相比，0.25%多聚甲醛溶液组细胞形态最典型，且7-AAD荧光染色较弱；4%多聚甲醛溶液组细胞形态典型，但7-AAD荧光染色较强；90%乙醇溶液组细胞发生肿胀，较正常细胞体积增大，且7-AAD荧光染色强；而75%乙醇溶液组细胞肿胀不如90%乙醇溶液组明显，7-AAD荧光染色强；完全培养基组细胞形态规则，7-AAD荧光染色最弱。 结论 0.25%多聚甲醛溶液对肿瘤细胞不但能够起到固定作用，且能够较好地维持细胞膜的完整性，是较为理想的细胞固定剂。

关键词: 多聚甲醛, 乙醇, 人胰腺腺泡上皮癌细胞, HeLa细胞, 7-氨基放线菌素荧光染色, 人

Abstract:

Objective To observe the effect of different fixative solutions on cancer cell morphology and membrane permeability. Methods Human pancreatic acinar epithelial carcinona(HPAC) cells of human pancreatic cancer and HeLa cells of human cervical cancer were fixed with 4 fixation solutions: freshly prepared 0.25% paraformaldehyde solution; Freshly prepared 4% paraformaldehyde solution; 75% ethanol solution; 90% ethanol solution. The fixation time is 30 minutes. PBS solution and complete medium were used as the controls. Cell morphology of each group was observed under optical microscope. Changes in cell membrane permeability were observed by fluorescence staining with 7-aminoactinomycin (7-AAD), which is not cell membrane permeable in intact cells but permeable in damaged cells. Hoechst33342 was used for staining both intact and damaged cells. Results The cells in the complete medium group were similar to unfixed cells in morphology, and the fluorescence staining of 7-AAD was the weakest. The cells in the complete medium group have typical cell morphology and low 7-AAD permeability. The 0.25% paraformaldehyde solution group had similar cell morphology to the complete medium group, and the 7-AAD fluorescence staining was weak. The morphology of cells in the 4% paraformaldehyde solution group was typical, but the fluorescence staining of 7-AAD was strong. The cells in the 90% ethanol solution group showed swelling, with a larger volume than the unfixed cells and a stronger fluorescence staining of 7-AAD. The cell swelling in 75% ethanol solution group was not as obvious as that in 90% ethanol solution group, and the fluorescence staining of 7-AAD was strong. The cells in PBS group were round, and the fluorescence staining of 7-AAD was strong. Conclusion 0.25% paraformaldehyde solution can not only fix tumor cells, but also maintain the integrity of cell membrane.

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