Optimization of trypsin digestion intensity to obtain high-purity in vitro cultured astrocytes

The aim of the present study was to observe the effect of trypsin digestion on the purity of in vitro cultured astrocytes and optimize the culture methods. The cerebral cortical tissue from newborn Sprague Dawley (SD) rats was isolated and digested with 0.25% trypsin for 20, 30, or 40 min. The obtai...

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Bibliographic Details
Published inSheng li hsüeh pao Vol. 67; no. 1; p. 103
Main Authors Jin, Hui, Yang, Peng-Bo, Feng, Gai-Feng, Jia, Ning, Yang, Wei-Na, Wang, Wei-Xi
Format Journal Article
LanguageChinese
Published China 25.02.2015
Subjects
Animals
Astrocytes - cytology
Cell Culture Techniques
Cell Proliferation
Cell Separation - methods
Glial Fibrillary Acidic Protein - metabolism
Rats
Rats, Sprague-Dawley
Trypsin
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Summary:The aim of the present study was to observe the effect of trypsin digestion on the purity of in vitro cultured astrocytes and optimize the culture methods. The cerebral cortical tissue from newborn Sprague Dawley (SD) rats was isolated and digested with 0.25% trypsin for 20, 30, or 40 min. The obtained single cell suspension was then cultured. Once reaching confluence, the cells were shaken at a constant temperature. Then, each of 20 and 30 min groups was subdivided into two groups, the control group with normal digestion and two-time-digestion group, and the cells were passaged and purified. Through inverted phase contrast microscope and MTT assay, cell growth and proliferation were observed, respectively. Immunofluorescence for glial fibrillary acidic protein (GFAP) was used to observe the morphology of astrocytes and to assess their purity in different stages. Flow cytometric analysis was used to detect the apoptotic rates of purified astrocytes. The results showed that, the cells being digested for 20 min
ISSN:0371-0874