Intracellular antibodies (intrabodies) are recombinant antibody fragments that bind to target proteins expressed inside of the same living cell producing the antibodies. The molecules are commonly used to study the function of the target proteins (i.e., their antigens). The intrabody technology is an attractive alternative to the generation of gene-targeted knockout animals, and complements knockdown techniques such as RNAi, miRNA and small molecule inhibitors, by-passing various limitations and disadvantages of these methods. The advantages of intrabodies include very high specificity for the target, the possibility to knock down several protein isoforms by one intrabody and targeting of specific splice variants or even post-translational modifications. Different types of intrabodies must be designed to target proteins at different locations, typically either in the cytoplasm, in the nucleus or in the endoplasmic reticulum (ER). Most straightforward is the use of intrabodies retained in the ER (ER intrabodies) to knock down the function of proteins passing the ER, which disturbs the function of members of the membrane or plasma proteomes. More effort is needed to functionally knock down cytoplasmic or nuclear proteins because in this case antibodies need to provide an inhibitory effect and must be able to fold in the reducing milieu of the cytoplasm. In this review, we present a broad overview of intrabody technology, as well as applications both of ER and cytoplasmic intrabodies, which have yielded valuable insights in the biology of many targets relevant for drug development, including α-synuclein, TAU, BCR-ABL, ErbB-2, EGFR, HIV gp120, CCR5, IL-2, IL-6, β-amyloid protein and p75NTR. Strategies for the generation of intrabodies and various designs of their applications are also reviewed.

A growing number of research consortia are now focused on generating antibodies and recombinant antibody fragments that target the human proteome. A particularly valuable application for these binding molecules would be their use inside a living cell, e.g., for imaging or functional intervention. Animal-derived antibodies must be brought into the cell through the membrane, whereas the availability of the antibody genes from phage display systems allows intracellular expression. Here, the various technologies to target intracellular proteins with antibodies are reviewed.

The use of antibodies to target their antigens in living cells is a powerful analytical tool for cell biology research. Not only can molecules be localized and visualized in living cells, but inter...

Challenges posed by complex diseases such as cancer, chronic viral infections, neurodegenerative disorders and many others have forced researchers to think beyond classic small molecule drugs, exploring new therapeutic strategies such as therapy with RNAi, CRISPR/Cas9 or antibody therapies as single or as combination therapies with existing drugs. While classic antibody therapies based on parenteral application can only reach extracellular targets, intracellular application of antibodies could provide specific advantages but is so far little recognized in translational research. Intrabodies allow high specificity and targeting of splice variants or post translational modifications. At the same time off target effects can be minimized by thorough biochemical characterization. Knockdown of cellular proteins by intrabodies has been reported for a significant number of disease-relevant targets, including ErbB-2, EGFR, VEGFR-2, Metalloproteinase MMP2 and MMP9, β-amyloid protein, α-synuclein, HIV gp120, HCV core and many others. This review outlines the recent advances in ER intrabody technology and their potential use in therapy.