A method for quantitative analysis of cell microtubules based on image analysis

Abstract

Objective To develop a method for objective quantitative analysis of microtubule structure based on image processing and analysis of the pixel ratio and particle size. Methods Human umbilical vein endothelial cells (HUVECs) cultured for 3 d were divided into control group, depolymerization group and polymerization group. Western blotting was performed to detect the changes in the content of acetylated α-tubulin in the cells. Immunofluorescence staining was used to observe the microtubule morphology under a laser confocal microscope, and the acquired images were analyzed using Matlab 7.0 for image segmentation, enhancement, and extraction of the feature parameters. Results ① The content of acetylated α-tubulin was significantly lower in cells with depolymerization and significantly higher in cells with polymerization than in the control cells (both P < 0.05). ②The control cells showed bundles of intact microtubules around the cell nuclei, with a few small fluorescent spots in the cytoplasm. In depolymerization group, the cells presented with broken microtubule fibers with numerous small fluorescent spots distributed diffusely in the cytoplasm. In the polymerization group, a large number of thick microtubule fibers were observed, showing multiple branches with an irregular and roughly radial alignment, and many large fluorescent spots were seen in the cytoplasm. ③The parameters for pixel ratio and particle size analysis, including the average particle size, sorting degree, skewness and kurtosis, all differed significantly among the 3 groups (P < 0.05). ④Based on quantification of these characteristic parameters of the microtubules, the accuracy of Fisher's multivariate discriminant analysis could reach 80.0%. Conclusion The pixel ratio and the characteristic parameters for particle size analysis can be used as objective indicators for quantitative analysis of the microtubes in fluorescence images of cells.

Keywords

• microtubule
• image processing and analysis
• pixel ratio
• particle size analysis