In my group we explore the structural, biophysical, and cellular outcomes of protein complex formation at membrane-bound receptors. We use Microscale Thermophoresis (MST) in addition to other methods including isothermal titration calorimetry and surface plasmon resonance and find a good agreement between the methods. We are very pleased with the low material consumption, short measurement times and broad application range of MST.

Prof. John E. Ladbury - Department of Biochemistry and Molecular Biology The University of Texas M. D. Anderson Cancer Center, Houston, TX, USA

"Our research is based on the development of small molecular compound based on or derived from linear peptides that have successfully been tested in animal models for inhibition of tumour angiogenesis and/or metastasis. These compounds are promising candidates to enter in clinical studies in the near future. We recently used NanoTemper Technologies' Microscale Thermophoresis to characterize the affinity of our compounds to their targets. Using this technology we obtained very convincing data that were in agreement with results obtained using other technologies like Surface Plasmon Resonance (SPR). We appreciated the relatively short time for the time to establish the assay compared to alternative methods to measure protein affinities as well as the comparatively low amounts of protein consumption. We conclude that this new technology gives rise to reliable quantitative affinity data."

Dr. Alexandra Matzke, CSO, amcure GmbH, Germany

"Our work focuses on the structure and function of proteins involved in signal transmission in the synaptic cleft. Upon arrival of an action potential in the synapse, ion channels open resulting in an increase in cytoplasmic calcium. This increase in calcium concentration is sensed by synaptotagmin-1 which mediates exocytosis of synaptic vesicles and release of neurotransmitter in the synaptic cleft. The precise mechanism of action of synaptotagmin-1 is unknown, but involves both binding to calcium and to the phospholipid PIP2 in the plasma membrane. In our MST studies, we used the technology to precisely quantify the cooperativity of PIP2 and calcium binding. PIP2-binding to synaptotagmin-1 resulted in a 40-fold higher affinity for calcium, and this result is of fundamental importance for our understanding of neuronal exocytosis. Given (i) the relatively high affinity of synaptotagmin-1 for both PIP2 and calcium, (ii) the low solubility of PIP2 and (iii) that synaptotagmin-1 is prone to aggregate at high protein concentrations, we cannot think of any technique other than MST that would allow studying this cooperative binding to such detail. Moreover, due to the very high sensitivity of MST, we estimate that the same amount of synaptotagmin-1 suffices to record about 10,000 times more calcium binding curves with MST compared to NMR or ITC."

Dr. Geert van den Bogaart, Max Planck Institute for Biophysical Chemistry, Department of Neurobiology, Göttingen, Germany

"One major focus of research in our institute is the dynamic organization of the microtubule cytoskeleton. An important approach for understanding the molecular mechanisms underlying tubulin assembly and microtubule motility has been the establishment of in vitro biochemical assays using purified components. Often, fluorescent microscopy, employing tagged or labeled microtubule associated proteins (MAPs) and motors, is used to study tubulin assembly and microtubule based motility.

We use Microscale Thermophoresis (MST) to investigate and quantify the interactions of these motors and MAPs with both un-assembled tubulin subunits as well as polymerized microtubules.

Previously, we have employed size exclusion chromatography, sucrose gradient sedimentation and immuno-precipitation to detect but not quantify these interactions as the necessary instrumentation was unavailable. MST has proven an ideal way for cell biologists in our institute to quantify interactions as, in most cases, the fluorescently tagged protein preparation is already available and, most importantly, only a limited amount of sample is required.

A second research focus in our institute is the assembly and dynamics of cell membranes, in particular, endosomes. We have started using MST to quality control the loading and exchange of guanine nucleotides on Rab5 protein, a small GTPase and a key regulator of endosome assembly. In this case, MST is used to monitor the nucleotide dependent association of Rab5 with proteo-lipid vesicles."

Dr. David Drechsel, Group Leader and Service and Facility Leader at the Max-Planck-Institute of Molecular Cell Biology and Genetics, Dresden, Germany

"The MicroScale Thermophoresis (MST) technology is a good complement to other biophysical techniques we are applying in the lab to investigate receptor proteins involved in inflammatory response. However, MST provides additional benefits since we are interested in elucidating interactions in close-to-native conditions: thus, we appreciate that the interactions are investigated free in solution and can be studied even in cell lysate. We are using MST for a number of different interactions, mainly to investigate protein-protein interactions but also to study protein-DNA interactions and protein binding to ions.

MST is a novel technique which is not related to any other established biophysical methods thus providing independent insights into biomolecular interactions. It is very easy to use with low time and material consumption."

Prof. Dr. med. Uffe Holmskov; 
Head of the Institute for Molecular Medicine, University of Southern Denmark, Odense, Denmark

"Our recent research is focused on the molecular basis of herbicide resistance and exploiting new target sites of herbicide, such as PPI of AHAS. Via MST, we measured the Protein-Protein interaction, that have been difficult to detect and quantify with ITC because of instability of the protein and very little heat changes of our samples. The MST technology can be used as a complementary technology to the traditional methods in these fields. We can strongly recommend the MST technology for its low material consumption, short measurement times and broad application range in the field of molecule interaction studies."

Prof. Zhen Xi, Chairman, Dean of Academic Affairs, School of Chemistry 
Fellow of the University Committee

"Our recent research has focussed on the interactions of proteins involved in the Hedgehog signalling pathway. We have used MST to develop a binding assay with fluorescently labelled peptide and unlabelled protein. The speed of the assay and low sample consumption has allowed us to quickly identify the critical residues in the binding site by rapidly screening a series of mutated proteins. Furthermore, the assay can easily be adapted to screen peptides and small molecules for competitive binding."

Luca Jovine, Ph.D.;
Professor of Structural Biology & EMBO Young Investigator, Karolinska Institutet, Huddinge, Sweden;
Amy Cherry, PhD;
Lecturer in Biochemistry at University of Worcester, UK

Past positions: Vice Director of the Institute of Biophysics; Director of the Center for the Molecular Biology, Institute of Biophysics; Vice Chairman of the Crystallographic Society of China; Vice Chairman of the Biophysical Society of China

"Our recent research is focused on the critical pathogenic protein & complex from pathogenic microbes and the Protein structure-functional basis for some endogenous disease. Via MST, we measured the Protein-DNA interaction, that have been difficult to detect and quantify with other standard biomolecule interaction technologies (like SPR and ITC) because of the conformation/immobilization-sensitive and very little heat changes of our samples. MST technology can be used as a complementary technology to the traditional methods in these fields. We can strongly recommend MST technology for its low material consumption, short measurement times and broad application range for molecule-interaction studies."

Prof. Dacheng Wang, MCAS, Member of Chinese Academy of Sciences

"We are interested in applying computational fragment-based strategy to identify lead compounds to block protein-protein interactions in RAS pathway. Generally, the identified fragments are weak binders with binding affinities ranging from millimolar to micromolar. For such weak interactions, it is difficult to set up reliable biochemical assays or binding assays using ITC or SPR techniques. Fortunately, MST technology turns out to be an excellent way to determine such weak interactions between small fragments and protein in our system of interest. We efficiently and reliably performed our experimental binding measurement using minimum amount of protein and fragments.

We are also interested in studying the regulation mechanism of a nuclear hormone receptor, which plays fundamental role in homeostasis and metabolism. The nuclear hormone receptor can be modified by an important enzyme to regulate the expression of specific genes and cause certain metabolic disease. We wanted to measure the binding affinity between these two proteins. Interestingly, this interaction is extremely difficult for the standard binding affinity measurement approach, with no exothermal or endothermal signals using ITC. While using MST technology, we have successfully detected this interaction. We strongly recommend MST technology for its low material consumption, short measurement time, and very high sensitivity."

Niu Huang, Ph.D.;
Associate Investigator, NIBS, Beijing, China

My group focuses on the molecular mechanisms of action of Receptor Tyrosine Kinases (RTKs) and Cell Adhesion Molecules (CAMs) in development and tumor progression. We have shown that the activation of RTKs relies not only on the binding of the ligand but also on the collaboration with CAMs such as CD44. We measured the binding of several growth factors to CD44 using the NanoTemper Technologies' Microscale Thermophoresis. Using this technology we obtained very convincing data that were in agreement with results obtained using other technologies like Surface Plasmon Resonance (SPR). This method is particularly useful to perform low affinity binding measurements using low amounts of proteins. We are extremely satisfied with the quantitative data obtained with this new technology.

PD Dr. Orian-Rousseau, Group Leader Institute for Toxicology and Genetic, Karlsruher Institut für Technologie (KIT)

"We use an integrative structural biology approach to study the regulation of gene expression by nuclear receptors. In this context, Microscale Thermophoresis (MST) is an extremely valuable tool to assess and quantify ligand-receptor, cofactor-receptor and nucleic acid-receptor interactions. By using MST, we determined the binding affinities of wild-type and mutant coregulator constructs for a homodimeric nuclear receptor that were in very nice agreement with those measured with isothermal titration calorimetry (ITC). However, in addition to ITC, MST allowed us to examine a range of concentration ratios between the two binding partners that could not be reached with ITC and to demonstrate the existence of allosteric control in the binding of the coregulator to the homodimeric nuclear receptor. We are using the Monolith NT.115 in our daily research work and we are very satisfied by the data obtained with this novel biophysical method. In addition, we appreciate the support and the competence from the NanoTemper Technologies Company."

Dr. Isabelle M.L. Billas, Team ‘Large complexes involved in gene expression’ led by Dr. Bruno Klaholz - Department of Integrative Structural Biology, France

"We routinely assess interaction affinity for both small molecule and biologics projects, with NanoTemper Technologies' Microscale Thermophoresis being the most recent addition to the pool of instruments we use to carry out these measurements. It has proved a valuable tool for characterising small molecule-protein and protein-protein interactions, as well as for the study of protein aggregation concentration determination. There is very good agreement with other technologies such as Surface Plasmon Resonance (SPR) and Isothermal Titration Calorimetry (ITC), and we are particularly appreciative of this new technology because of the extremely low protein consumption and relatively short time required for the assay setup. NanoTemper customer support has been a key factor in enabling us to familiarise ourselves with the new technology. We would like to deploy increasing numbers of applications based on MST technologies and continue to interact with NanoTemper Technologies Company, a dynamic, scientifically driven company." 

Dr. Alexey Rak, 
Head of Structural Biology & Biophysics, Sanofi R&D, France

"As part of our drug discovery projects we use Microscale Thermophoresis (MST) as an orthogonal method to measure the binding affinity of compounds to their protein target and apply this fragment hit ID through to lead optimisation. MST complements our biophysical platform and has correlated well with other more established technologies. Because it uses small amounts of protein, MST has proved to be particularly useful to look at molecular interaction involving proteins that are difficult to express or purify. MST requires a relatively short time to setup new assays and is a powerful technique for buffer optimisation. Using the NT.115 MST instrument we have successfully measured small molecule-protein and protein-protein interactions in complex media. Finally, LabelFree MST is one of the few true label-free/immobilisation-free instruments capable of measuring molecular interactions. We have appreciated the professionalism and support from NanoTemper Technologies."

Dr. Nicolas Basse, Department of Structural Biology, UCB-Celltech, UK

"We are interested in molecular architectures especially to stabilize and analyze challenging smell receptors that belong to G protein-coupled receptors (GPCRs). GPCR is a super-family of receptors linked to many diseases. We want to measure the interactions of smell receptors with odorant that are very low molecular weight ligands. In the past that has been extremely difficult to detect and quantify other common molecular interaction technologies including SPR, and nearly impossible to use ITC for odorant-binding measurement because of the conformation and immobilization-sensitivities, or difficult and costly to obtain sufficient amount of receptors. The MST technology is a superb technology, it not only is label-free, hence reducing the measuring artifacts, but it also is surface-free that provides true measurement of finest molecular interactions. Thus it allows us much better analysis of the GPCR with their small-molecule ligands. We strongly recommend MST technology for its low material consumption (a few micrograms in a few microliters), short measurement times, inexpensive consumables, and broad application range in all molecule-interaction study."

Dr. Shuguang Zhang, Center for Bits and Atoms

“The main focus of our work lies on structure based-drug design. In this context, we study the interaction of the bacterial tRNA modifying enzyme tRNA-guaninetransglycosylase (Tgt) with small molecule inhibitors and with tRNA. Althoughwe were unable to measure any interaction of this enzyme with its small moleculesubstrates guanine and 7-deaza-7-aminomethyl guanine via MST, a strong influence on the thermophoretic behaviour of Tgt was noticed upon binding of the Macromolecular tRNA substrate allowing determination of the respective Kd value. In addition, MST yielded Kd values for lin-benzoguanine-based Tgt inhibitors (300-500 Da), which were excellently consistent with Ki values previously determined in enzyme kinetic studies. Accordingly, this fast and easy to use method provides a highly welcome alternative to the radioactive enzyme assay we used so far to figure out binding affinities of Tgt inhibitors. Furthermore, we investigate the interaction of a chaperone of the Shigella type III secretion system with its protein interaction partners or rather with synthetic peptides thereof to identify the respective recognition motifs. Via MST, we determined with low amounts of protein material Kd values which were in nearly perfect agreement with those measured via isothermal titration calorimetry.”

Prof. Dr. Gerhard Klebe,
Institut für Pharmazeutische Chemie, Philipps-Universität Marburg, Germany

"Together with Nanotemper Technologies we have optimized microscale thermophoresis for fragment screening applications which are difficult to access with standard technologies. Access to accurate information on fragment Kd, binding mode and aggregation behaviour of compounds enable informed decision making. Together with a proprietary fragment collection, this technology forms now the core of the CRELUX fragment based drug discovery service offerings. "

Dr. Ismail Moarefi, Chief Scientific Officer of CRELUX GmbH - A WuXi AppTec Company in Martinsried, Germany

"One mission of the Structural Research group within the Lead Identification and Optimization department is to quantitatively validate and characterize interactions of small molecules as well as new biological entities with protein targets. We employ various traditional biophysical methods such as SPR, Thermal Shift, ITC, NMR and X-ray crystallography. 
Most recently we included Microscale Thermophoresis (MST) in our standard project support workflow and extended its application from affinity determinations to fragment screening approaches. MST is a versatile and valuable tool which we quickly adapted in our repertoire of methods. The impressive advantages of MST, namely the low sample consumption, the broad application range, and swift assay development make it a unique biophysical method. The measurement in free solution without the need of surface coupling saves time and avoids a potential source for false positive or negative results. 
Our Structural Research group now also added the label-free version of the Monolith to our MST instrumentation portfolio, which gives us the possibility to choose to measure with high selectivity and sensitivity (NT.115). The labelfree version (NT.LabelFree) allows us to measusre without any additional sample modification depending on the need of the particular assay. In some cases, LabelFree MST allowed us to perform assays with otherwise "very ill" behaved proteins which were not amenable to any other biophysical technique. Generally, we find very good consistency between quantitative MST measurements and results stemming from other biophysical methods."

Dr. Markus Zeeb, Principal Scientist, Structural Research, Boehringer-Ingelheim, Germany

"The work of our group gave rise to the novel concept that under certain conditions matrix components may act as endogenous "danger" signals, which are recognized by innate immunity receptors, and are capable to trigger an inflammatory response reaction. We are using MST to measure the interactions of endogenous and in vitro mutagenized proteoglycans of the extracellular matrix with their receptors and are very satisfied with the results obtained so far. The affinities are consistent with the biological readouts and confirm previous results gained with immunprecipitation and binding assays. MST is a new technology we can definitely recommend for obtaining robust quantitative affinity data."

Prof. Dr. med. Liliana Schaefer, Pharmazentrum /ZAFES, Frankfurt a.M., Germany

 "Membrane interaction and fusion plays a fundamental role in many cellular processes such as intracellular trafficking, fertilization, tissue formation or viral infection. With the new MST technology we were able to measure the interaction of membrane vesicles, mediated by coiled coil-forming peptides. The technology requires only little sample material and is enabling for this type of experiments"

Prof. Dr. Andreas Janshoff, Dept. of Physical Chemistry. University of Göttingen, Germany

 "We are studying the processing of microRNA precursors In Drosophila and are using MST to measure the binding of nucleolytic Enzymes and their specificity factors to RNA substrates. The small amount of protein sample needed and the much faster measuring time compared with e.g. gel-shift assays are particular strengths of this new technique. It enables us to ask questions that we could not address before." 

Prof. Dr. Klaus Förstemann, Gene Center of the LMU, Munich, Germany

Yi Zheng, PhD, Director, Division of Experimental Hematology and Cancer Biology; Katherine Stewart Waters Endowed Chair, University of Cincinnati College of Medicine, USA

"Part of our research focus on the protein structural-function basis for iron metabolism of pathogenic microorganisms. We measured the dissociated constant for periplasmic binding protein and siderophore through MST technology. Because siderophore is extremely insoluble, it's difficult for us to complete that task via other methods such as ITC and SPR. We strongly recommend MST technology for its low material consumption, short measurement time and very wide application range."

Prof. Li-chuan GU, Principal Investigator, Shandong University, China

"We are interested in understanding the interactions between cells and organisms by investigating the role of polysaccharides exposed on cell surfaces and secreted polysaccharide signal molecules. We are applying MST to determine structural requirements for recognition of complex polysaccharides and the role of ligand-receptor interactions in the relationships between different cells and organisms.

MST is very useful for my lab since it allows to measure interactions in solution even in complex samples of membrane proteins. Also the small amount of sample material needed and the broad range of applications are very advantageous."

Prof. Jens Stougaard, Director of CARB, Department of Molecular BiologyLaboratory of Gene Expression, Aarhus University, Denmark

"Our lab is developing diagnostic tools based on protein biochips and is studying the basic mechanisms underlying herpes virus infections" We are using MST for elucidating the function of viral proteins and its interaction with other viral and host cell proteins. MST is simple to use and was quickly established in our lab. In particular we do appreciate the low instrument, consumables and maintenance costs associated with this new technology"

Dr. Maria  G. Vizoso Pinto, Max-von-Pettenkofer Institute, Munich, Germany

“We study the structural and molecular biology of bacterial adhesins and cell-surface filaments with respect to their function in bacterial pathogenesis, with the ultimate aim of developing a new generation of virulence-targeted antimicrobials. MST really is opening up the easy determination of some Kd's that were hard to get to with other methods. So far, we measured protein-protein, protein-glycan, protein-small compound and protein-cofactor interactions with the Monolith NT.115.”

Prof. Han Remaut, VIB Laboratory of Structural and Molecular Microbiology, Vrije Universiteit Brussel, Belgium

"My lab focuses on the elucidation of the structure and function of membrane proteins. This topic includes the investigation of protein-protein and protein-substrate interactions. We are employing MST in binding studies of membrane sensors to their transducers as well as soluble transcription factors to DNA. We also plan for investigations of receptor - ligand interactions with this exciting new technique. MST turned out to be an extremely useful method in our lab to determine binding constants. It is advantageous that the fast and handy experiments require only few amounts of protein."

Prof. Dr. Joachim Heberle, Exp. Molecular Biophysics, FU Berlin, Germany

"Our lab focuses on the mechanisms and organization of DNA packaging inside the cell nucleus. We study the biochemical properties of molecular machines, the chromatin remodelers that regulate DNA accessibility and switch between 'on' and 'off' states of genes. We are using MST for studying the activity of chromatin remodeling enzymes and to quantify their affinities towards DNA, RNA and other proteins. MST is a big step forward towards quantitative biochemistry. MST is simple to use and was quickly established in our lab. It is a good alternative to the traditional electrophoretic mobility shift assays".

Prof. Dr. Gernot Längst, Biochemistry III, University of Regensburg, Germany