Langlois N E, King G, Herriot R, Thompson W D
University of Aberdeen, Scotland, U.K.
J Pathol. 1994 Jul;173(3):249-53. doi: 10.1002/path.1711730308.
The rabbit polyclonal antibody to protein gene product 9.5 (PGP9.5) will detect the L1 isoenzyme of ubiquitin carboxy-terminal hydrolase (UCH), which is a marker for neurones and neuroendocrine tissue. We re-evaluated this antibody using the technique of non-enzymatic antigen retrieval (boiling sections in citrate buffer, heated by microwave oven) followed by streptavidin-biotin-peroxidase staining. Due to the fortuitous choice of appendix as positive control material containing small nerves, we found strong, repeatable cytoplasmic and nuclear staining of lymphoid follicle centre cells in addition to neural tissue. This effect could be repeated on other lymphoid tissues and was not dependent on microwave heating, but did require boiling in an ionic buffer solution. Staining was also observed with a fresh batch of antibody and with four of the five different batches of antibody which were supplied to us. This pattern was not obtained in fresh tissue, in fixed material following trypsinization, or by increasing the primary antibody concentration. We suggest that the boiling of sections in citrate buffer is exposing an epitope for the anti-PGP9.5 antibody which is inaccessible in the native or fixed state and therefore we would recommend retesting of antibody specificity following non-enzymatic retrieval of antigen.
针对蛋白基因产物9.5（PGP9.5）的兔多克隆抗体可检测泛素羧基末端水解酶（UCH）的L1同工酶，UCH是神经元和神经内分泌组织的标志物。我们采用非酶促抗原修复技术（在柠檬酸盐缓冲液中煮沸切片，用微波炉加热），然后进行链霉亲和素-生物素-过氧化物酶染色，重新评估了这种抗体。由于偶然选择阑尾作为含有小神经的阳性对照材料，我们发现除神经组织外，淋巴滤泡中心细胞也有强烈的、可重复的细胞质和细胞核染色。这种效应在其他淋巴组织上也可重复，且不依赖于微波加热，但确实需要在离子缓冲溶液中煮沸。用一批新抗体以及提供给我们的五批不同抗体中的四批进行检测时也观察到了这种染色。在新鲜组织、胰蛋白酶消化后的固定材料中，或通过增加一抗浓度，均未获得这种染色模式。我们认为，在柠檬酸盐缓冲液中煮沸切片会暴露抗PGP9.5抗体的一个表位，该表位在天然或固定状态下是无法接近的，因此我们建议在非酶促抗原修复后重新检测抗体的特异性。
