Cancer is still a leading cause of mortality. Each year 18 million people worldwide get diagnosed with cancer. The current trend is increasing. This makes cancer research more important.
What’s a Chromobody?
Chromobodies® are fluorescent nanoprobes for real-time and live-cell imaging of intracellular endogenous proteins. A Chromobody is a small intracellular functional antibody. It consists of a VHH/Nanobody that is genetically fused to a fluorescent protein. Plasmids encoding for Chromobodies can be transiently transfected into cells and the Chromobody protein is intracellularly expressed.
Vimentin dynamics can be effectively visualized using the ChromoTek Vimentin-Chromobody, e.g. to monitor epithelial-mesenchymal transition (EMT). To learn more about this unique probe attend Julia Maier’s talk “Tracing EMT with fluorescent biosensors (Chromobodies) in living lung cancer cells” on April 3, 4:20 pm in Room 144, Level 1 at AACR 2017 in Washington, DC.
Everybody really wants to use a detection probe that does not influence its target. For measurement of actin and actin-related functions there are a number of actin visualization probes and techniques available that researchers can select. Hence, one should only apply such actin visualization probes, which do not alter actin dynamics. How do the actin probes for visualization perform? Which probes do interfere with function and which probes do not? Robert Grosse and his team from the Institute of Pharmacology at the Biochemical-Pharmacological Center (BPC), University of Marburg, Germany took the effort and systematically looked into detection technologies of actin filaments in order to avoid potential pitfalls. In the recently published mini review/poster “Actin Visualization at a Glance” (Melak et al. 2017) they have discussed the pros and cons of current probes to visualize actin filaments, i.e. Phalloidin, LifeAct, Utrophin, F-tractin, SiR-actin, GFP-actin, tagged actin, and Actin Chromobody® (see table 1 and figure 1 below).
The progression of epithelial cancers into metastatic stages is triggered by a cellular process called “epithelial-mesenchymal transition” (EMT). In response to cytokines, tumor cells shut down the expression of epithelial makers including occludin or E-cadherin and gain mesenchymal markers such as N-cadherin or vimentin. Such cells are losing their cell-cell contacts and apical-basal polarity and become highly migratory and invasive. Considering the importance of this process for cancer progression and formation of metastasis, reliable cellular EMT biomarker and versatile detection systems are necessary to develop novel anti-metastatic therapies.
This time last spring we started marketing our Chromobody® plasmids. Our customers realized the impact of the technology and were immediately beginning to work with the Chromobodies®. Now, one year later the first papers are being published demonstrating the broad applicability of this intracellular fluorescent antibody technology:
The first is a paper in Molecular Cell by the Lindqvist Lab at the Karolinska Institutet in Stockholm, Sweden. They developed a FRET-based system for accurate quantification of fluorescence from single cells. By visualizing endogenous PCNA at replication foci with the Cell Cycle Chromobody® they nicely show that the mitotic entry network is linked to the completion of S phase. It does not depend on protein accumulation through G2 but is activated by mitotic phosphorylations at the end of S phase. The method they present allows analyzing live-cell as well as extracting temporal information from fixed cells based on endogenous marker proteins. (Akopyan, K.; Silva Cascales, H.; Hukasova, E., et al. Assessing kinetics from fixed cells reveals activation of the mitotic entry network at the S/G2 transition. Molecular cell. 2014, 53, 843-853. http://dx.doi.org/10.1016/j.molcel.2014.01.031)
Chromobodies® are new intracellular functional antibodies visualising dynamic changes of target proteins in live cells. With the help of NMI`s outstanding lentiviral transduction technology, Chromobodies® will be introduced into difficult-to-transfect cell lines such as primary cells. The cooperation will establish innovative cell lines available for phenotypic screening, target validation and high content analysis (HCA). The unique combination of Chromobody® nanoprobes and a comprehensive collection of disease-relevant cellular test systems will lift preclinical compound screening and pathway analysis in the early drug discovery process to a new level of performance.
- Intracellular Fluorescent Alpaca Antibodies (Chromobodies):
Non-invasive Biomarkers for High-Content Analysis in Mammalian Cells
- Development of Comparative, Cell-based, Reversible p53:Mdm2 and p53:Mdm4 High-Content Fluorescent Two-Hybrid (F2H) Assays for Drug Screening