Kaiser Philipp D, Maier Julia, Traenkle Bjoern, Emele Felix, Rothbauer Ulrich
Natural and Medical Sciences Institute at the University of Tuebingen, Markwiesenstrasse 55, 72770 Reutlingen, Germany; Department of Pharmaceutical Biotechnology, University of Tuebingen, Auf der Morgenstelle 8, 72076 Tuebingen, Germany.
Natural and Medical Sciences Institute at the University of Tuebingen, Markwiesenstrasse 55, 72770 Reutlingen, Germany; Department of Pharmaceutical Biotechnology, University of Tuebingen, Auf der Morgenstelle 8, 72076 Tuebingen, Germany.
Biochim Biophys Acta. 2014 Nov;1844(11):1933-1942. doi: 10.1016/j.bbapap.2014.04.019. Epub 2014 May 2.
In biomedical research there is an ongoing demand for new technologies, which help to elucidate disease mechanisms and provide the basis to develop novel therapeutics. In this context a comprehensive understanding of cellular processes and their pathophysiology based on reliable information on abundance, localization, posttranslational modifications and dynamic interactions of cellular components is indispensable. Besides their significant impact as therapeutic molecules, antibodies are arguably the most powerful research tools to study endogenous proteins and other cellular components. However, for cellular diagnostics their use is restricted to endpoint assays using fixed and permeabilized cells. Alternatively, live cell imaging using fluorescent protein-tagged reporters is widely used to study protein localization and dynamics in living cells. However, only artificially introduced chimeric proteins are visualized, whereas the endogenous proteins, their posttranslational modifications as well as non-protein components of the cell remain invisible and cannot be analyzed. To overcome these limitations, traceable intracellular binding molecules provide new opportunities to perform cellular diagnostics in real time. In this review we summarize recent progress in the generation of intracellular and cell penetrating antibodies and their application to target and trace cellular components in living cells. We highlight recent advances in the structural formulation of recombinant antibody formats, reliable screening protocols and sophisticated cellular targeting technologies and propose that such intrabodies will become versatile research tools for real time cell-based diagnostics including target validation and live cell imaging. This article is part of a Special Issue entitled: Recent advances in molecular engineering of antibody.
在生物医学研究中，对新技术的需求持续存在，这些技术有助于阐明疾病机制，并为开发新型治疗方法提供基础。在此背景下，基于细胞成分的丰度、定位、翻译后修饰和动态相互作用的可靠信息，全面了解细胞过程及其病理生理学是必不可少的。除了作为治疗分子具有重大影响外，抗体可以说是研究内源性蛋白质和其他细胞成分最强大的研究工具。然而，对于细胞诊断，其应用仅限于使用固定和通透细胞的终点分析。另外，使用荧光蛋白标记报告基因的活细胞成像被广泛用于研究活细胞中的蛋白质定位和动态。然而，只能观察到人工引入的嵌合蛋白，而内源性蛋白质、其翻译后修饰以及细胞的非蛋白质成分仍然不可见且无法分析。为了克服这些限制，可追踪的细胞内结合分子为实时进行细胞诊断提供了新机会。在这篇综述中，我们总结了细胞内和细胞穿透抗体生成方面的最新进展及其在活细胞中靶向和追踪细胞成分的应用。我们强调了重组抗体形式的结构设计、可靠的筛选方案和先进的细胞靶向技术方面的最新进展，并提出此类细胞内抗体将成为用于基于细胞的实时诊断（包括靶点验证和活细胞成像）的通用研究工具。本文是名为：抗体分子工程的最新进展的特刊的一部分。
