Isolation, Culture and Identification of Hybrid Calf Intestinal Epithelial Cells

LIU Yifan, TAN Xiuwen, YOU Wei, ZHANG Xianglun, JIANG Fugui, SONG Enliang, WAN Fachun

doi:10.3969/j.issn.1006-267x.2019.05.034

Chinese Journal of Animal Nutrition >

2019 , Vol. 31 >Issue 5: 2278 - 2286

DOI: https://doi.org/10.3969/j.issn.1006-267x.2019.05.034

Molecular Nutrition

Isolation, Culture and Identification of Hybrid Calf Intestinal Epithelial Cells

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1. Institute of Animal Science and Veterinary Medicine, Shandong Academy of Agricultural Sciences, Ji'nan 250100, China;
2. Shandong Key Lab of Animal Disease Control and Breeding, Ji'nan 250100, China;
3. Shandong Provincial Testing Center of Beef Cattle Performance, Ji'nan 250100, China;
4. Shandong Provincial Engineering Technology Center of Animal Healthy Breeding, Ji'nan 250100, China

Received date: 2018-10-09

Online published: 2019-05-15

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Abstract

This experiment was designed to establish a method for isolation, culture and identification of hybrid calf intestinal epithelial cells, in order to provide primary cell culture models for the study of intestinal nutrient absorption, immune regulation and intestinal barrier function. Jejunum tissue of newborn hybrid calf was cut down and digested with mixture of collagenase Ⅰ and dispase. The calf intestinal epithelial cells were purified by differential digestion and differential attachment methods. The methods of cell morphology observation, growth curve, immunofluorescence and karyotype analysis were used to identify the cells. The results showed as follows: 1) calf intestinal epithelial cells obtained using mixture of collagenase Ⅰ and dispase digestive method grown well. The calf intestinal epithelial cells could be purified to more than 90%, and could pass more than 10 generations stably. 2) The "S" shape of growth curve of calf intestinal epithelial cells was accordance with the pattern of cell proliferation. 3) The calf intestinal epithelial cells were stained positive for cytokeratin-13 and villin by immunofluorescence identification. 4) Morphological features such as microvilli structure and tight junctions between cells were observed under transmission electron microscope. 5) Karyotype analysis showed that the number of chromosomes with normal morphology and the karyotype was diploid. The above results show that calf intestinal epithelial cells has been successfully obtained by collagenase Ⅰ and dispase combined digestion and differential attachment purification, which can used for the study of intestinal nutrient absorption, immune regulation and the intestinal barrier function.

Key words： calf; intestinal epithelial cell; combined digestion method; cell identification; karyotype analysis

Cite this article

LIU Yifan, TAN Xiuwen, YOU Wei, ZHANG Xianglun, JIANG Fugui, SONG Enliang, WAN Fachun . Isolation, Culture and Identification of Hybrid Calf Intestinal Epithelial Cells[J]. Chinese Journal of Animal Nutrition, 2019 , 31(5) : 2278 -2286 . DOI: 10.3969/j.issn.1006-267x.2019.05.034

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