Sasaki J, Takehara Y, Fujii Y, Nomura T, Watanabe S
Am J Anat. 1987 May;179(1):1-9. doi: 10.1002/aja.1001790102.
The nonionic detergent Triton X-100 has often been used for the extraction of cytoplasmic materials. We used the detergent in a vascular perfusion medium when preparing rat lung in order to observe the cytoskeleton of the nonciliated bronchiolar epithelial (Clara) cells. To eliminate some cytoplasmic materials selectively and to maintain good fine cell structure simultaneously, the lungs were perfused sequentially with the detergent (0.2% Triton X-100) alone for 2 min, with a mixture of low-concentration (0.1%) glutaraldehyde and detergent (0.2% Triton X-100) for 15 min, and finally with 2.5% glutaraldehyde for 5-10 min. After fixation, the nonciliated bronchiolar epithelial (Clara) cells were observed by scanning and transmission electron microscopy. At the apical region of the cells, there were central cytoplasmic protuberances (apical caps) filled with microfilaments. These filaments were bound at one end to the cytoplasmic side of the cell membrane and ran into the interior of the cytoplasm at the other end. As a control, the Clara cells were observed by transmission electron microscopy after perfusion with 2.5% glutaraldehyde solution. The luminal surfaces of the cells were covered with short, thick microvilli. The apical caps also had microvillus-like protrusions. These results suggest that the apical cap is not an apocrine droplet but rather is a stable structure involved in the function of the Clara cells.
非离子去污剂曲拉通X-100常被用于提取细胞质物质。我们在制备大鼠肺时，在血管灌注培养基中使用了这种去污剂，以便观察无纤毛细支气管上皮（克拉拉）细胞的细胞骨架。为了选择性地去除一些细胞质物质并同时保持良好的细胞精细结构，先单独用去污剂（0.2%曲拉通X-100）灌注肺2分钟，再用低浓度（0.1%）戊二醛和去污剂（0.2%曲拉通X-100）的混合物灌注15分钟，最后用2.5%戊二醛灌注5 - 10分钟。固定后，通过扫描电子显微镜和透射电子显微镜观察无纤毛细支气管上皮（克拉拉）细胞。在细胞的顶端区域，有充满微丝的中央细胞质突起（顶端帽）。这些细丝一端与细胞膜的细胞质侧相连，另一端延伸到细胞质内部。作为对照，在用2.5%戊二醛溶液灌注后，通过透射电子显微镜观察克拉拉细胞。细胞的腔面覆盖着短而粗的微绒毛。顶端帽也有微绒毛样的突起。这些结果表明，顶端帽不是顶泌小滴，而是参与克拉拉细胞功能的稳定结构。
