Schousboe L P, Ovesen T, Ottosen P D, Ledet T, Elbrönd O
Ear-, Nose and Throat Department, Aarhus University Hospital, Denmark.
Acta Otolaryngol. 1995 Nov;115(6):787-95. doi: 10.3109/00016489509139403.
During the last decade middle ear epithelium has been cultured from various species. Until now, subcultivation has been achieved only with the use of a feeder-cell layer or conditioned medium. These factors are possible confounders in the in vitro model. On the other hand, subcultivation is necessary for exact quantitative studies. We present a reproducible culture method allowing subcultivation without feeder-cells or conditioned medium. The main features in our method are a low-serum, hormone-supplemented medium, an incubation temperature of 34 degrees C, fixation of explants, gentle trypsinization and replating with high cell density. Cells were identified by immunohistochemistry through a battery of monclonal antibodies. The percentage of epithelial cells in the subculture was 99.2%. To our knowledge, this is the first report describing subcultivation of middle ear epithelial cells exclusively in a completely controlled environment. These are optimal circumstances for future investigation and quantification of various factors influencing proliferation and differentiation of middle ear epithelium.
在过去十年间，已从多种物种中培养出中耳上皮细胞。到目前为止，仅通过使用饲养层细胞或条件培养基才能实现传代培养。在体外模型中，这些因素可能会造成干扰。另一方面，传代培养对于精确的定量研究是必要的。我们提出了一种可重复的培养方法，该方法无需饲养层细胞或条件培养基就能进行传代培养。我们方法的主要特点是使用低血清、添加激素的培养基，培养温度为34摄氏度，对外植体进行固定，轻柔胰蛋白酶消化，并以高细胞密度重新接种。通过一系列单克隆抗体进行免疫组织化学鉴定细胞。传代培养中上皮细胞的百分比为99.2%。据我们所知，这是第一份描述在完全受控环境中单独对中耳上皮细胞进行传代培养的报告。这些是未来研究和量化影响中耳上皮细胞增殖和分化的各种因素的最佳条件。
