Koretz K, Leman J, Brandt I, Möller P
Histochemistry. 1987;86(5):471-8. doi: 10.1007/BF00500619.
3-Amino-9-ethylcarbazole (AEC) used as chromogen in immunoperoxidase techniques normally has an intense, red colour. However, as an inconstant phenomenon, a pale yellowish-green reaction product severely impairing the evaluation can be observed. In order to circumvent this undesired effect, factors such as tissue fixative, proteolytic digestion, antibody concentrations and incubation time of the primary antibody were analyzed. The most important factor inducing a change in colour is probably the inadequately high local peroxidase concentration arising as the consequence of high amounts of bound primary antibody. This high enzyme concentration might cause metachromasia of AEC by producing the yellowish-green quinone-di-imine form of the substrate. As could be shown by spectrophotometry in test tube experiments, AEC metachromasia was proven to be enzyme dependent. Thus, the best way to trigger the local enzyme concentration on a tissue section to adequate levels appears to be the dilution of the primary antibody.
3-氨基-9-乙基咔唑（AEC）在免疫过氧化物酶技术中用作显色剂时，通常会产生强烈的红色。然而，作为一种不稳定的现象，可以观察到淡黄色至绿色的反应产物，这严重影响了结果的评估。为了避免这种不良影响，我们分析了诸如组织固定剂、蛋白水解消化、抗体浓度和一抗孵育时间等因素。导致颜色变化的最重要因素可能是由于大量结合的一抗导致局部过氧化物酶浓度过高。这种高酶浓度可能通过产生底物的黄绿色醌二亚胺形式而导致AEC的异染性。正如试管实验中的分光光度法所示，AEC异染性被证明是酶依赖性的。因此，将组织切片上的局部酶浓度调节到适当水平的最佳方法似乎是稀释一抗。
