Multiple organ infection and the pathogenesis of SARS

Gu, Jiang; Gong, Encong; Zhang, Bo; Zheng, Jie; Gao, Zifen; Zhong, Yanfeng; Zou, Wanzhong; Zhan, Jun; Wang, Shenglan; Xie, Zhigang; Zhuang, Hui; Wu, Bingquan; Zhong, Haohao; Shao, Hongquan; Fang, Weigang; Gao, Dongshia; Pei, Fei; Li, Xingwang; He, Zhongpin; Xu, Danzhen; Shi, Xeying; Anderson, Virginia M.; Leong, Anthony S.-Y.

To identify the cell types that were infected by the virus, double labeling was performed by combining in situ hybridization for SARS viral genomic sequences and immunohistochemistry for one of the cell-associated markers. The in situ hybridization signal was visualized with alkaline phosphatase, which results in a dark blue color, whereas positive immunostaining signal was visualized with diamenobenzodin, which gives a reddishbrown color. When the two signals are present in the same cell, the staining changes to a purplish color, which is easily distinguishable from either of the primary colors. In addition, in situ hybridization and immunohistochemistry were performed separately on consecutive sections to analyze the relationship between in situ hybridization signals and immunostaining signals to ascertain the cell types that were infected by the virus. Seven cell marker antigens were examined to identify epithelial cells, macrophages, T cells, B cells, and splenic dendritic cells. The specificities and the sources of these antibodies are shown in Table III .