Pleiner, T.; Bates, M.; Trakhanov, S.; Lee, C.T.; Schliep, J. E.; Chug, H.; Böhning, M.; Stark, H.; Urlaub, H.; Görlich, D. (2015) Nanobodies: site-specific labeling for super-resolution imaging, rapid epitope-mapping and native protein complex isolation. eLife. 4; e11349.
This work describes multiple strategies to engineer nanobodies as precision tools for cell biology and is based on multiple interdisciplinary collaborations at the Max Planck Institute for Biophysical Chemistry. In close collaboration with the group of Stefan Hell, we used nanobodies as ultra-small immunolabels for super-resolution microscopy. For this, we developed a site-specific fluorescent labeling strategy that maximizes nanobody brightness, while maintaining specificity and stability. We also established a nanobody-based single-step strategy for the native purification of protein complexes e.g. for structural analysis by electron microscopy (in collaboration with the group of Holger Stark). We also started mapping conformational nanobody epitopes by crosslinking mass spectrometry in collaboration with the group of Henning Urlaub.