MAPPING PROTEIN DISTRIBUTIONS IN EPIDERMIS BY ELECTRON LOSS SPECTROSCOPY

能量过滤电子显微镜和电子显微镜 能量损失谱（EELS）已被用于表征 富硫蛋白在分层上皮中的分布， 它们在皮层中的再分布，特别注意 组装皮质细胞包膜（CE）。测量 提供了定量信息，补充了高度具体的 而是来自免疫细胞化学的定性数据。空间 硫、磷和氮的分布图 小鼠表皮和前胃的未染色薄切片，使用 电子光谱成像氮素分布 而磷图像提供了一个 控件来验证用于计算元素的算法 地图硫被发现在高水平的圆形L-颗粒中， 粒细胞胞质和皮质中，与 存在蛋白质兜甲蛋白（约7% Cys/Met残基）。鳗鱼 扫描透射电子显微镜测量 定量地证实了这些观察结果。这些数据还 确认CE中约75%的蛋白质是兜甲蛋白， 如从其氨基酸组成的建模所推断的。 在皮质间也观察到细胞外沉积物 胎鼠表皮鳞状细胞。其中的硫含量 外周沉积物至少与L-颗粒一样高，但它们 不标记抗兜甲蛋白抗体，表明存在 另一种富含硫的蛋白质这是一个延续Intramural 研究项目Z 01-RR-10481-02 BEI。

Energy-filtered electron microscopy and electron energy loss spectroscopy (EELS) have been used to characterize the distributions of sulfur-rich proteins in stratifying epithelia and their redistribution in the cornified layer, with particular attention to assembly of the cornified cell envelope (CE). Measurements provided quantitative information that complements highly specific but qualitative data from immunocytochemistry. Spatial distributions of sulfur, phosphorus and nitrogen were mapped in unstained thin sections of mouse epidermis and forestomach, using electron spectroscopic imaging. Nitrogen distributions were indicative of total protein while phosphorus images provided a control to validate the algorithms used to calculate the elemental maps. Sulfur was found at high levels in round L-granules in the granulocyte cytoplasm and in the cornified CE, correlating with the presence of the protein, loricrin (~ 7% Cys/Met residues). EELS measurements in the scanning transmission electron microscope confirmed these observations quantitatively. These data also confirm that approximately 75% of the protein in the CE is loricrin, as inferred from modeling of its amino-acid composition. Extracellular deposits were also observed between the cornified squames in fetal mouse epidermis. The sulfur content of these peripheral deposits is at least as high as that of L-granules but they do not label with anti-loricrin antibodies, suggesting the presence of another sulfur-rich protein. This is a continuation of Intramural Research Project Z01-RR-10481-02 BEI.