Longinotti Gloria, Ybarra Gabriel, Vighi Susana, Perandones Claudia, Montserrat Javier, Yakisich Juan Sebastian, Grasselli Mariano, Radrizzani Martin
Nanomateriales Funcionales, INTI-Micro y Nanotecnologías, Instituto Nacional de Tecnología Industrial (INTI), Av. Gral. Paz 5445, San Martín B1650WAB, Argentina.
Centro de Anatomía Patológica, Ciudad de la Paz 353, Buenos Aires C1426AGE, Argentina.
Diagnostics (Basel). 2021 Jan 26;11(2):171. doi: 10.3390/diagnostics11020171.
Antibodies are the most used technological tool in histochemistry. However, even with monoclonal antibodies, their standardization is difficult due to variation of biological systems as well as to variability due to the affinity and amplification of the signal arising from secondary peroxidase detection systems. In this article we combined two synthetic molecules to facilitate the standardization of a detection protocol of protein markers in histological sections. The first molecule was an aptamer, a 50-base single-stranded DNA fragment, which recognizes a PTEN tumor suppressor. The second molecule used was also another single stranded 18-base aptamer DNA fragment, which forms a quadruplex structure guanine box. This G-quadruplex recognizes and attaches a molecule of hemin, increasing the catalytic capacity for the hydrogen peroxide. Our results show how the correct structural design of DNA combining an aptamer together with the peroxidase-like DNAzyme allows to detect proteins in histological sections. This tool offers the standardization of the detection of prognostic markers in cancer, in quality and quantity, due to its synthetic nature and its 1:1 antigen:enzyme ratio. This is the first time that reproducible results have been presented in histological sections staining a cancer marker using a single-stranded DNA molecule with dual function.
抗体是组织化学中最常用的技术工具。然而，即使是单克隆抗体，由于生物系统的差异以及二级过氧化物酶检测系统产生的信号亲和力和放大作用导致的变异性，其标准化也很困难。在本文中，我们结合了两种合成分子，以促进组织切片中蛋白质标记物检测方案的标准化。第一个分子是适体，一个50个碱基的单链DNA片段，它识别一种PTEN肿瘤抑制因子。使用的第二个分子也是另一个18个碱基的单链适体DNA片段，它形成一个鸟嘌呤盒的四链体结构。这种G-四链体识别并附着一个血红素分子，增加了对过氧化氢的催化能力。我们的结果表明，将适体与过氧化物酶样DNA酶结合的DNA的正确结构设计如何能够在组织切片中检测蛋白质。由于其合成性质及其1:1的抗原:酶比例，该工具在质量和数量上为癌症预后标志物的检测提供了标准化。这是首次使用具有双重功能的单链DNA分子在组织切片中对癌症标志物进行染色并呈现出可重复的结果。
