Comparison of three morphology methods for observing mineralization nodules of osteoblasts

Liao Nai-shun; Li Zuan-fang; Lin Ru-hui; Chen Wen-lie; Huang Yun-mei; Huang Mei-ya

doi:10.3969/j.issn.2095-4344.2014.33.003

Abstract：BACKGROUND:Mineralized nodules are the mature marker of osteoblast differentiation, and the observation methods mainly use alizarin red staining. <br> OBJECTIVE:To compare the observation results of mineralized nodules by three methods, and to explore their characteristics and advantages, as wel as further application in the research of bone disease. <br> METHODS:The rat osteoblast-like cellline UMR-106 were cultured in the fresh medium that was changed every day, for 14 days. Alizarin red staining-light microscope, tetracycline fluorescence labeling-laser confocal scanning microscopy, and scanning electron microscopy were used to observe mineralized nodules. The calcium content of mineralized nodules was quantified using scanning electron microscopy and energy dispersive spectroscopy. In addition, tumor necrosis factor alpha that could inhibit the proliferation and differentiation of osteoblasts was used as the control. <br> RESULTS AND CONCLUSION:The three morphology methods could be used to observe the mineralized nodules of normal osteoblasts. As for tumor necrosis factor alpha, no mineralized nodules of osteoblasts were observed by alizarin red staining-light microscopy;smal mineralized nodules were observed by tetracycline staining-laser scanning confocal microscopy and scanning electron microscopy, suggesting tetracycline staining and scanning electron microscopy were more sensitive in the observation. Scanning electron microscopy could be used to observe the submicroscopic structures of mineralized nodules in the osteoblasts, and the formation of mineralized nodules, including the calcium secretion. Additional y, scanning electron microscopy combined with energy dispersive spectroscopy analysis can successful y quantify and position the mineralized nodules, indicating a potential application in the research of bone diseases.