Shimosegawa T, Uchida T, Kobayashi S, Ito S, Shibata A
Nihon Naibunpi Gakkai Zasshi. 1984 Jan 20;60(1):44-53. doi: 10.1507/endocrine1927.60.1_44.
Met-enkephalin-like immunoreactivity-containing nerve elements in the canine pancreas was investigated by immunocytochemistry. Anti-Met-enkephalin (Met-Enk) serum used as the primary anti-serum was raised against synthetic Met-Enk conjugated with bovine serum albumin by a glutaraldehyde method. For light microscopic immunocytochemistry, Bouin-fixed and paraffin-embedded sections not exceeding 20 micrometers in thickness were stained by the PAP method. For electron microscopy, the fixative used was 4% paraformaldehyde plus 1% glutaraldehyde in a 0.067 M phosphate buffer. Ultra-thin sections of Araldite-embedded materials were stained by a protein A-colloidal gold method. The main localization of Met-Enk-like immunoreactivity-containing nerve fibers and ganglion cell somas was as follows: a) on the surface of ganglion cell somas, b) among nerve fiber bundles in the connective tissue septum, c) around blood vessels in the exocrine parenchyma, and d) in the islet of Langerhans. Electron microscopic immunocytochemistry showed that colloidal gold particles representing a Met-Enk-like substance were concentrated on secretory type granules of the ganglion cell somas and nerve terminals around the blood capillaries. Met-Enk-like immunoreactivity was eliminated by preincubation of the primary anti-serum with Met-Enk (nerve fibers: 0.1 X 2(-3) g/l, ganglion cells: 0.1 X 2(-2) g/l of the diluted antiserum, 1:2000) before application to tissue sections. The functional significance of Met-Enk-like immunoreactivity-containing nerve elements in both the endocrine and exocrine pancreas was discussed.
采用免疫细胞化学方法研究了犬胰腺中含甲硫氨酸脑啡肽样免疫反应性的神经成分。用作一抗血清的抗甲硫氨酸脑啡肽(Met-Enk)血清是通过戊二醛法针对与牛血清白蛋白偶联的合成甲硫氨酸脑啡肽制备的。对于光镜免疫细胞化学,用PAP法对厚度不超过20微米的Bouin固定石蜡包埋切片进行染色。对于电子显微镜检查,固定剂为0.067M磷酸盐缓冲液中的4%多聚甲醛加1%戊二醛。用蛋白A-胶体金法对Araldite包埋材料的超薄切片进行染色。含甲硫氨酸脑啡肽样免疫反应性的神经纤维和神经节细胞体的主要定位如下:a)神经节细胞体表面;b)结缔组织间隔中的神经纤维束之间;c)外分泌实质中的血管周围;d)胰岛内。电子显微镜免疫细胞化学显示,代表甲硫氨酸脑啡肽样物质的胶体金颗粒集中在神经节细胞体的分泌型颗粒和毛细血管周围的神经末梢上。在将一抗血清应用于组织切片之前,用甲硫氨酸脑啡肽(神经纤维:稀释抗血清0.1×2(-3)g/l,神经节细胞:0.1×2(-2)g/l,1:2000)预孵育可消除甲硫氨酸脑啡肽样免疫反应性。文中讨论了内分泌和外分泌胰腺中含甲硫氨酸脑啡肽样免疫反应性神经成分的功能意义。