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Journal of Cellular Biochemistry

Purity, cell viability, expression of GFAP and bystin in astrocytes cultured by different procedures

Authors

  • Fang Du,

    1. Jiangsu Key Laboratory of Neuroregeneration, Department of Neurochemistry, Nantong University, Nantong 226001, PR China
    2. Faculty of Medicine, Institute of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong
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  • Zhong Ming Qian,

    1. Jiangsu Key Laboratory of Neuroregeneration, Department of Neurochemistry, Nantong University, Nantong 226001, PR China
    2. National Key Laboratory of CMMP (Shenzhen) and Laboratory of Brain Iron Metabolism, Department of Applied Biology & Chemical Technology, Hong Kong Polytechnic University, Kowloon, Hong Kong
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  • Li Zhu,

    Corresponding author
    1. Jiangsu Key Laboratory of Neuroregeneration, Department of Neurochemistry, Nantong University, Nantong 226001, PR China
    2. National Key Laboratory of CMMP (Shenzhen) and Laboratory of Brain Iron Metabolism, Department of Applied Biology & Chemical Technology, Hong Kong Polytechnic University, Kowloon, Hong Kong
    • Jiangsu Key Laboratory of Neuroregeneration, Department of Neurochemistry, Nantong University, Nantong 226001, PRC.
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  • Xiao Mei Wu,

    1. Jiangsu Key Laboratory of Neuroregeneration, Department of Neurochemistry, Nantong University, Nantong 226001, PR China
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  • Christopher Qian,

    1. Faculty of Medicine, Institute of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong
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  • Robbie Chan,

    1. National Key Laboratory of CMMP (Shenzhen) and Laboratory of Brain Iron Metabolism, Department of Applied Biology & Chemical Technology, Hong Kong Polytechnic University, Kowloon, Hong Kong
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  • Ya Ke

    Corresponding author
    1. Faculty of Medicine, Institute of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong
    • Faculty of Medicine, Institute of Biomedical Sciences, The Chinese University of Hong Kong, Shatin, NT, Hong Kong.
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  • Zhong Ming Qian, Li Zhu, and Ya Ke contributed equally to this work.

Abstract

Primary astrocyte cultures are the most commonly used in vitro model for neurobiological studies. We speculated that different protocols might induce differences not only in the percentage of astrocytes but also in their biological characteristics. In this study, we investigated the effects of four major protocols on the purity of astrocytes, cell viability, expression of glial fibrillary acidic protein (GFAP) and bystin of cultured astrocytes using MTT assay, immunocytochemical staining, and Western blot analysis. We demonstrated that the purity of astrocytes (98.9%) generated by the subculture (SC) procedure is significantly higher than those generated by primary culture (PC), shaken once culture (SK-1) or shaken twice culture (SK-2). We also showed that expressions of GFAP and bystin in astrocytes that are purified by the SK-2 or SK-1 procedures are significantly higher than those in astrocytes prepared by PC or SC. In addition, astrocytes cultured by SK-2 or SK-1 have a higher level of cell viabilities at most time points after ischemia compared with astrocytes cultured by PC or SC. These suggested that physical stimulation induced by “shaken” or culture operation might be able to activate astrocytes and implied that different procedures induce differences not only in the purity but also in the biological characteristics of astrocytes, such as the percentage of activated astrocytes, GFAP, and bystin expressions and responses to ischemia. A more detailed analysis about the effect of “culture protocol factor” on the biological characteristics of astrocytes is absolutely needed. J. Cell. Biochem. 109: 30–37, 2010. © 2009 Wiley-Liss, Inc.

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