Miller B E, Chapin R E, Pinkerton K E, Gilmore L B, Maronpot R R, Hook G E
Exp Lung Res. 1987;12(2):135-48. doi: 10.3109/01902148709062837.
A light microscopic technique based on alkaline phosphatase histochemistry was developed to specifically quantitate Type II cells in the intact rat lung. Lungs were fixed in 4% neutral-buffered formalin containing 0.25 M sucrose and embedded in glycol methacrylate. Two micron thick sections were mounted on glass microscope slides. Alkaline phosphatase activity was localized by using naphthol AS-BI phosphate as substrate in 0.125 M 2-amino-2-methyl-1-propanol buffer containing 0.625 mM MgCl2 (pH 8.9). Sections were counterstained with Harris hematoxylin. Type II cells were the only cell type in the alveolar region containing alkaline phosphatase activity, an observation that was confirmed by using electron microscopic histochemistry. By combining the alkaline phosphatase staining technique with standard morphometric procedures, the proliferative response to a single intratracheal dose of 10 mg silica was followed as a function of time. Type II cells were significantly increased at all time points examined. Twenty eight days following silica, Type II cells had increased to (252 +/- 16) X 10(6) cells per set of lungs compared to a control value of (141 +/- 32) X 16(6) cells. The method presented is a simple and rapid technique for examining Type II cell population kinetics.
开发了一种基于碱性磷酸酶组织化学的光学显微镜技术，用于特异性定量完整大鼠肺中的II型细胞。将肺固定在含有0.25M蔗糖的4%中性缓冲福尔马林中，并包埋在甲基丙烯酸乙二醇酯中。将2微米厚的切片安装在玻璃显微镜载玻片上。以萘酚AS-BI磷酸酯为底物，在含有0.625mM MgCl2（pH 8.9）的0.125M 2-氨基-2-甲基-1-丙醇缓冲液中定位碱性磷酸酶活性。切片用Harris苏木精复染。II型细胞是肺泡区域中唯一具有碱性磷酸酶活性的细胞类型，这一观察结果通过电子显微镜组织化学得到证实。通过将碱性磷酸酶染色技术与标准形态计量学方法相结合，跟踪单次气管内给予10mg二氧化硅后的增殖反应随时间的变化。在所有检测的时间点，II型细胞均显著增加。二氧化硅处理28天后，II型细胞增加至每组肺（252±16）×10⁶个细胞，而对照值为（141±32）×10⁶个细胞。所提出的方法是一种用于检测II型细胞群体动力学的简单快速技术。
