北京生物医学工程

基于原子力显微镜力曲线阵列模式获得兔角膜基质细胞弹性模量的方法研究

Study on methods of obtain elastic modulus of rabbit keratocytes by force volume model of AFM

作者：陈昕妍曾正张海霞李林

单位：首都医科大学生物医学工程学院，临床生物力学应用基础研究北京市重点实验室(北京 100069)

关键词：细胞弹性模量; 原子力显微镜; 力曲线阵列; Snedden 模型; 角膜基质细胞

分类号：R318.01

出版年·卷·期（页码）：2020·39·6（574-581）

摘要：

目的细胞的力学特性与自身的生物学特性及某些疾病息息相关，原子力显微镜（atomic force microscope, AFM）可提供多种扫描模式及测试方法，而关于细胞弹性模量的获得方法目前尚未统一，本研究拟探讨通过原子力显微镜力曲线阵列模式（force volume）获得细胞弹性模量的方法。方法首先通过酶消化法提取正常兔眼角膜基质细胞，进行培养并通过波形蛋白（vimentin）免疫荧光染色进行鉴定。通过原子力显微镜力曲线阵列测试，选定9个100×100 μm2区域，获得每个区域均布的1 024点的力-压痕深度曲线，利用Snedden模型拟合曲线获得相应测试点弹性模量。结果波形蛋白（vimentin）免疫荧光染色鉴定结果证实提取细胞免疫荧光波形蛋白鉴定阳性，确定细胞为角膜基质细胞。通过原子力显微镜测试得到每个区域约有3~5个完整细胞，拟合得到细胞的弹性模量中央与边缘处略有差别。细胞中央处拟合得弹性模量约为53.1 kPa±6.35 kPa。结论原子力显微镜力曲线阵列模式可以较为简易方便地获得众多细胞数据，更为细致的获得细胞整体力学特性，有效地避免了实验操作与压入点不同带来的误差。

Objective The mechanical properties of cells are closely related to their biological characteristics and some diseases. AFM（atomic force microscope) provide several scan and test models, while there is no uniform method to obtain the elastic modulus of cells so far. In this study, the elastic modulus of cells was obtained by force volume model of AFM. Methods Normal rabbit keratocytes were extracted by enzyme digestion, then identified by Vimentin immunofluorescence staining. Through the force volume model of AFM, nine 100×100 μm2 regions were tested and the force-indent depth curves of 1024 points of each area were obtained. The elastic modulus was obtained through the fitting of force-indent depth curves via Snedden model. Results The immunofluorescence staining was positive, confirmed that the cell is keratocytes. There were about 3-5 complete cells in each tested region, and the elastic modulus of the cells was slightly different between the center and the edge area. The elastic modulus at the center of the cell was approximately 53.1 kPa±6.35 kPa. Conclusions The force volume model of AFM can obtain cell data in a simple and convenient way. And the overall mechanical properties of keratocytes are obtained in detail, which effectively avoids the errors caused by different experimental operations and pressing points.

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