Immuno EM Study of Parafibromin Distribution in Cells

为了补充先前在共聚焦光学显微镜中的免疫标记研究，进行免疫电子显微镜（EM），目的是以更高的空间分辨率确定蛋白质parafibromin在人胚肾细胞中的亚细胞分布。副纤维蛋白与甲状旁腺功能亢进-颌肿瘤家族性癌症综合征以及甲状旁腺癌和甲状旁腺功能亢进有关。将培养的细胞引入塑料毛细管中，并在高压冷冻机中在2000个大气压下冷冻以保存其细胞超微结构。将冷冻组织块在含有戊二醛固定剂的丙酮中冷冻置换，包埋在树脂中，切片，并用抗副纤维蛋白的一抗标记。将样本与缀合至金的二抗反应，并在能量过滤透射电子显微镜（TEM）中进行数字成像。电子显微照片显示，parafibromin主要集中在细胞核与一些标记在其他地方的细胞。

To complement previous immunolabeling studies in the confocal light microscope, immuno electron microscopy (EM) was performed with the aim of determining the subcellular distribution of the protein parafibromin in human embryonic kidney cells at higher spatial resolution. Parafibromin is implicated in the hyperparathyroidism-jaw tumor familial cancer syndrome, as well as parathyroid cancer and hyperparathyroidism. Cultured cells were introduced into plastic capillary tubes and frozen at 2000 atmospheres in a high-pressure freezing machine to preserve their cellular ultrastructure. The frozen blocks were freeze-substituted in acetone with glutaraldehyde fixative, embedded in resin, sectioned, and labeled with primary antibodies against parafibromin. Specimens were reacted with a secondary antibody conjugated to gold and were imaged digitally in an energy-filtered transmission electron microscope (TEM). Electron micrographs showed that parafibromin was concentrated mainly in the nucleus with some labeling elsewhere in the cell.