Frederick S E, Newcomb E H
J Cell Biol. 1969 Nov;43(2):343-53. doi: 10.1083/jcb.43.2.343.
Segments of mature tobacco leaves were fixed in glutaraldehyde, incubated in medium containing 3,3'-diaminobenzidine (DAB) and hydrogen peroxide, and postfixed in osmium tetroxide. Electron microscopic observation of treated tissues revealed pronounced deposition of a highly electron-opaque material in microbodies but not in other organelles. The coarsely granular reaction product is presumably osmium black formed by reaction of oxidized DAB with osmium tetroxide. Reaction of the microbodies with DAB was completely inhibited by 0.02 M 3-amino-1,2,4-triazole and was considerably reduced by 0.01 M potassium cyanide. These results, when considered in light of recent biochemical studies, strongly suggest that catalase is responsible for the reaction. Sharp localization of this enzyme in microbodies establishes that they are identical to the catalase-rich "peroxisomes" recently isolated from leaf cell homogenates. A browning reaction that occurred in leaves during the incubation step was inhibited by cyanide but not by aminotriazole and therefore could not have been caused by the same enzyme. This reaction and a slight deposition of dense material within primary and secondary walls are ascribed to oxidation of DAB by soluble and wall-localized peroxidases.
将成熟烟草叶片的切片固定在戊二醛中，置于含有3,3'-二氨基联苯胺（DAB）和过氧化氢的培养基中孵育，然后用四氧化锇进行后固定。对处理过的组织进行电子显微镜观察发现，一种高度电子致密的物质在微体中大量沉积，而在其他细胞器中则没有。这种粗颗粒状的反应产物可能是氧化型DAB与四氧化锇反应形成的锇黑。微体与DAB的反应被0.02 M的3-氨基-1,2,4-三唑完全抑制，被0.01 M的氰化钾显著降低。结合近期的生化研究结果来看，这些结果有力地表明过氧化氢酶是该反应的原因。这种酶在微体中的精确定位表明，微体与最近从叶细胞匀浆中分离出的富含过氧化氢酶的“过氧化物酶体”是相同的。孵育步骤中叶片发生的褐变反应被氰化物抑制，但不被氨基三唑抑制，因此不可能是由同一种酶引起的。这种反应以及初生壁和次生壁内致密物质的轻微沉积归因于可溶性和壁定位过氧化物酶对DAB的氧化作用。
