Cell Biology The Cell Biophysics & Imaging Group
at the Netherlands Cancer Institute (NKI-AVL)
Kees Jalink, PhD, Principal Investigator
Genomic- and high-throughput screening methods have identified tremendous amounts of biomedically relevant proteins. The functions of these proteins can not be fully understood without detailed knowledge of their localization, concentration, and particularly, their mutual interactions and activation state in living cells. Many of these interactions are short-lived or exist very locally within the cell and therefore techniques with high spatiotemporal resolution are required to study them in single living cells. Our lab focuses on biophysical techniques to provide this resolution. Our lab is well-equipped for both electrophysiological and advanced biophotonic studies, and we develop and implement new techniques. We apply biophysics in our ongoing research and in a number of collaborations within and outside of the institute.
Biophotonic techniques
Major techniques to obtain detailed spatiotemporal information on biomolecules rely on studying the interactions of cellular constituents with light, an area termed biophotonics. Proteins can be labelled with fluorescent tags like GFP or chemical fluorochromes and studied non-invasively by live cell imaging. However, due to the limited resolution of light microscopy many essential parameters escape detection: examples are kinetic parameters such as on- and off-rates (often in the ms or us time range), protein-protein interactions, and protein conformation/ activation state (both at nm-scale). Details on kinetic parameters and interactions are therefore derived indirectly by using approaches such as Fluorescence Recovery After Photobleaching (FRAP), Fluorescence Loss In Photobleaching (FLIP), Fluorescence Resonance Energy Transfer (FRET), Total Internal Reflectance Fluorescence (TIRF), Fluorescence Lifetime Imaging (FLIM), electrophysiological techniques, and Fluorescence Correlation Spectroscopy (FCS). A whole range of novel tools including fluorescent proteins in all colours, photo-activatable compounds (pa-GFP, light-inducible crosslinkers, and UV-releasable ‘caged’ second messengers have recently become available. These techniques aim to collect information about the funcion of molecules, rather than static pictures. Therefore, they have collectively been called ‘Functional Imaging’.
Ongoing research
FRET sensors are genetically encoded constructs engineered to report on intracellular signaling events. Our sensors for messengers like PIP2 (the first lipid FRET sensor ever!) and cAMP, and for activation of receptors (Estrogen receptor) and proteins-protein interactions.
We also develop new hardware, software and algorithms for fast readout of FRET using Fluorescence Lifetime Imaging Microscopy (FLIM).
We develop and apply Super Resolution Imaging (GS-DIM / dSTORM). 3-color STORM is a breeze with our OxEA imaging buffer.
We apply those in ongoing research lines studying signal transduction, migration/invasion, cell survival, the effects of hypoxic conditions, and much more. We carry our FLIM screens, using RNA silencing libraries and small molecule inhibitor libraries, and we study kinetic properties of signal transduction networks, using computer modelling to understand the results.
Embedding
Our group is part of the van Leeuwenhoek Center for Advanced Microscopy (LCAM) and embedded within the ESFRI Roadmap EuroBioImaging, as well as in the Dutch initiative NL-BioImaging-Advanced Microscopy. We do advisory work and beta testing for leading vendors of microscopy and FLIM equipment. We also welcome guests with challenging imaging / analysis tasks to our lab.
Come and discuss your imaging projects with us, or inquire about a student rotation.
Over the last twelve months Kees Jalink was a peer reviewer for eLife
