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Home > Archive>Volume 29, Issue 5, 2019 >109-115. DOI:10.3969/j. issn. 1671 -7856. 2019. 05. 017
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Isolation and culture of tree shrew primary corneal stromal cells
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(Department of Ophthalmology, the Fourth Affiliated Hospital of Kunming Medical University; Department of Ophthalmology,Yunnan Second People’s Hospital; Yunnan Ophthalmology Research Institute; Yunnan Key Laboratory of Ophthalmological Research, Kunming 650021, China)

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R-33

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    Abstract:

    Objective To explore a simple and easy method for the rapid isdation and culture of corneal stromalcells (CSCs). Methods Two experimental groups A and B were designated. The corneal stromal layer was removed fromtree horny cornea. In the group A, it was cut, digested with 1% type II collagenase for 60 min, and resuspended aftercentrifugation. In the group B, type II collagenase was used first,and then treated with neutral protease digestion. Thegrowth and passage time of primary and passage cells were recorded and compared. Vimentin was detected byimmunohistochemistry and immunofluorescence assays. Results The collagenase digestion method which was used in thegroup A successfully isolated primary cells. After 9 days, the cells were passaged and showed good cell morphology. Thepassage cells grew fast and were passaged for 2 to 3 days. The double enzyme digestion method used in the group B isolatedfewer CSCs by day 10. A small number of scattered cells were observed, which were passaged from 15 to 20 days.Immunohistochemistry and immunofluorescence assays showed positive vimentin expression of the corneal stromal cells inboth groups. Conclusions Both methods can successfully isolated CSCs for culture, but the digestion method with only collagenase is easier and more efficiently to isolat a large number of tree shrew CSCs.

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History
  • Received:October 15,2018
  • Online: June 05,2019
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