 So, good morning everyone. So, it's my great pleasure to talk about, you know, lipid membranes or cell surfaces and membrane mimics at this interesting sweetness course. And before I start talking about science, I would like to briefly introduce you myself because I'm not a Swedish professor. Yeah. So, as Tommy mentioned, I was born and grown up in Kyoto and got my PhD in Kyoto University. But during my PhD, I spent some time in Helsinki and also in Copenhagen, precisely speaking, in detail. After my PhD, I moved to Germany 23 years ago and kept my career here mainly. So, I started as a postdoc in the physics department in Munich under the guidance of Erich Sackmann. He was one of the pioneers of membrane physics in Germany and Europe. Then later on, I got an independent grant to run my team myself for five years in Munich. Then made my habilitation degrees in physics at Munich. Right afterwards, I got promoted as a full professor of chemistry and later on physics at Heideberg. Yeah, then since 2013, I revived my activities after 15 years of absence in Kyoto, leading a team there. So, the reason why I'm here is because of your academic coordinator, yeah, Tommy. So, he mentioned already that we met for the first time, I think 15, 16 years ago. And afterwards, I don't know how many different places and occasions we met and had beer and talking about science in different cities and countries. So, we're here, Tommy and me. Yeah. Yeah. One was the summer school held in the island, Uito, 10 years ago. And the second one I showed you is the conference that I organized in Kyoto, where also Harbour is in the middle. So, in my team, I have three scientific pillars. I'm a sophomore physicist by training. So, the main pillar is dealing with physics or biological soft interfaces, where I set light on structure, mechanics and dynamics using both real space techniques as well as reciprocal space techniques, like scattering. This stems out into two directions. The light left one is suited for Heideberg, which is strongly in biomedicine, where I study the non equilibrium physics of life, especially focusing on the physics of diseases and development. Another direction that I'm working still on is to generate biosimical data hybrid materials, such as depositing membrane mimics on the surface of gallium nitride quantum nodes or two dimensional nuclear gases and quantum wires to detect the smart bio functions as an active optorextric system. So, of course, today, I would like to focus sharply on my main field, physics of biological interfaces. So, that's the reason why I chose this or I like this topic to talk. So, to start from, you know, let me start from a historical question about molecules and interface. In 1774, Benjamin Franklin made a very short report at the Royal Society meeting. He reported that when the tin spoonful of olive oil was spread on the pond, it stills away. But most importantly, the effect reached up to one quarter acre, but not more. So, that's a very short message and you may ask yourself what one can learn out of it. And of course, this is not an experiment recommended nowadays for the sake of environment. When you pour oil on the water surface of the pond, I think some people will call the police. The answer is given by Lord Rayleigh, another famous guy about 100 years later. What he did was he just divide volume by area and gained the thickness of the film of olive oil being one nanometers. Now we know that this is a very simple calculation, but giving us a right guess about the size or thickness of monolayer of fatty acids. This is so simple, but we tend to forget that the interface is not just a boundary that separates two bulk phases, but they have structures as well as functions. This is manifested in biological systems. So we can learn why does interface matter by visiting nature again. So I took a very famous cartoon from Strayer's textbook. It's showing that in biological cells, many key reactions are confined near the interface, which is nothing but the membrane separating interior and exterior worlds. So one can think of the importance of membrane from the economical viewpoint. So we can start from a classical, classical, most costly equation of diffusion and calculate. The diffusion time required for a particle collide with another one located at the distance B. We do it in 2D and 3D spaces. This was firstly shown by Stefan Hart in Weizmann in the late 70s, and the solution shows that the dependence of tau on 2 and 3D is quite different. So the tau, at least the diffusion time, in 2D scales with natural lock of R, while the one in 3D scales with 1 over R, clearly indicating that the dependence of diffusion time on R is minor in 2D. And this explains indeed the confinement of reaction in or near 2D membrane is very much an economically clever solution to run the reaction involving biomacromolecules such as proteins and enzymatic complexes. As you learn already from Ilpo's lecture this morning, membranes are not just boundary of all, but membranes know actively control the biological function, such as protein recombinitions, receptor activation, and so on, so forth. So here I show you one of one picture I presented in one of the most recent review together with my teacher, Erik Sakman, where we showed the proteins involved in cell migration, which are shown in this cartoon. If you zoom into the region near the spreading front of a migrating cell, you find many protein machineries involved. But all these proteins are initially swimming in such a specific space and they are not active unless they are recruited to the membrane surface. And binding of the PI3 kinase is indeed the master switch producing a lot of PIP3 lipids, so phosphorynostero 3-phosphate lipids, that connects proteins to the membrane. Also, if you see from the mechanical viewpoint, cell membranes possess very unique mechanical properties which are playing instrumental roles in sustaining our life. As I show here with the scheme, membrane consists of consisting of amphiphilic lipid molecules, and lipid itself has a polar head and hydrophobic tails, sharing common features with symmetric liquid crystals. So, symmetric liquid crystal phase is the leftmost phase on the right panel. Actually, in 70s, many physicists working on liquid crystals become aware of unique mechanics of lip membranes and they actually build up the fundament of membrane biophysics. So, in this slide, I show you three principle mechanical parameters describing the mechanical properties of lipid membranes. The first mode is an isotropic extension and contraction, and the middle one is a sharing deformation, which is a deformation which is not accompanied by the change of the area. And the third one is the bending deformation, which comes from the asymmetric tension balance between interior, inside, and outside of the field. So, everything can be represented by the Hukian energy representation I show in the lower part, which is even by one half times the modulus times the strain to the power of two. So, this is exactly what you learn in high school about the Hukian spring. So, as one of the prominent examples of biological membranes mechanics, I selected one very well-known example, human red blood cells. Here, I show you some statistics. About 10 to the power of 13 cells are running inside our body, and actually, when I was a student, I was astonished to see the production rate. One million cells per second are produced in our bone marrow, and they are traveling over 400 kilometer distance in 12 days, keeping finite volume of area in order to avoid the clotting. So, they are traveling long distance, patterned through many narrow capillaries. So, what I show here is a movie taken by my student showing healthy red blood cells passing through a constriction, which has the cross-section of 3 by 4 micrometers, meaning the capillaries. What is intuitive is the healthy guys are able to go through these narrow channels and easily recover their original shapes. But the disease cells as well as aged cells cause stiffer than the healthy one, and they are often rupture when they are encountering through this kind of constriction. In fact, the membrane has extremely low bending modulus, center power minus 20 joule, and low shear modulus, center power minus 6 Newton per meter, which are like 1,200,000 times softer compared to polyethylene fumes, which has a similar thickness. So, compared to plastic forms, they are 1,200,000 times softer, and this is a secret why they are able to keep their structure integrity against the strong shear stresses. So, now we look into the multicellular systems, and the first question that I can ask myself is how are these cell-cell and cell tissue contacts are precisely controlled? And when we look into the biological systems, we find that many cell-cell or cell tissue contacts keep a very finite and well-controlled spacing to avoid non-specific contact with an aid of many biopolymers. What I show here on the left is taken from Albert showing two plant cells, two neighboring plant cells, whose contact is mediated by a 200 nanometer thick cellulose layer. So, if I draw a cartoon showing this, the skin looks like on the right. In fact, many cell-cell contacts are mediated by biopolymers containing sugars, and they are controlling the generic and specific interaction balances in order to find and adjust the sensing contact. Actually, from the viewpoint of physics, cell-cell contacts can be treated as a certain property of working physics. There are two important points for us to keep in our mind. The first point is the stratified layers, namely layer by layer structure, like you saw in the previous slide. So, phytosodic polymer and membrane, excessibular polymer, membrane and cytosol. They are stable only if the complete wetting conditions are fulfilled at the interface, namely the presence of an additional layer. For instance, in this cartoon, the additional layer means lipid membrane. This must result in the gain of the surface free energy, which is characterized by the spreading coefficient as being positive, which was described by Dijon and Francois Rochauer quite some time ago. Second, when you look at the vertical force interactions, to keep a finite inter-cellular distance of typically tens to hundreds of nanometers, the interaction potential at the interface should be kept weakly repulsive. So, if you think about the interfacial forces contributing to these biological interfaces, you can name some, like ecostatic interactions, van der Waals, hydration repulsion and entropic forces generated by conformational fluctuation of polymers, as well as thermal fluctuation of membranes. But if you think about cell-cell contacts, it's very difficult or almost impossible to dissect each force contributor. Therefore, we can grab the net force active per unit area, which is nothing but the discerning pressure. This is a concept written by Teriagin from Russia for quite some time ago. And from the dynamic viewpoint, the discerning pressure is nothing but the first derivative of the work. You have to invest to change the distance between two membranes with respect to the distance. As you can imagine, the minimum can be found. So, corresponding to the equilibrium corresponds to the discerning pressure equals zero can be found only if the second derivative goes to positive. On the other hand, if you have negative discerning pressure, this causes the continuous thinning of the interlayer, which eventually results in the dissipation or rupture. And this is nothing but the scenario of wetting-dewetting transitions. So, therefore, in order to understand the physics behind, the design of defined cell membrane models with less complexity would be helpful to reveal how biopolymer modulate interfacial interactions. So, now we come to see the very important model of cell surfaces called supported membranes. There was a very historically important, I think, review published by Eric Suckman 25 years ago in Science. Entitled Supported Membranes. This is now cited over 2,000 times, and now it's really widely spread all over the world. So, the first report came from Adrian Bryan and Harden McConnell from Stanford 36 years ago. The systems look like that. You have a planar lipid bilayer deposited on planar substrate. And the membrane is separated from the substrate with a very thin water reservoir, which has a thickness of 5 to 10 nonsomes. Compared to the other membrane models, like, you know, black lipid membrane, liposomes, you know, the advantages of the supported membrane is, first, they can stably coat macroscopically large surfaces, which can go easily beyond tens of square centimeters. And also, you can study the structure and function using various surface-tensive techniques, such as AFM, you know, because of the planar geometry of the sample. Actually, in the review written in 1996, Eric already mentioned that this model of dark membrane that allowed the application of a manifold of surface-tensive techniques. And they form versatile models of low-dimensionality complex roots, which can be used to study interfacial forces and waiting for pneumonia and adhesion. And even he started talking about practical applications for, towards a design of biosensors on electro-optical devices. So about 30 years ago, related to neutral science, there are two important papers published by Swedish, British, and German physicists. The first paper I picked was published by Adrian Reigning, who was, at that time, ILL, and both Thomas in Oxford and Eric Sackman from Munich. They did the speculative neutral reflectivity of lipid bilayer deposited on quartz substrate using ILL D17, which is still one of the top runners in the field of neutral reflectability. The second paper I picked was also published by these people. Well, they were very active in this year, obviously, where they performed the new speculative neutral reflectivity of lipid monolayer at the outer interface. There they used crisp beam line at ISIS, the Rutherford-Uppleton Laboratory. So these systems have been used like a gold standard, and still we are using this lipid monolayer and supported membranes as a cell surface mimics. But when you want to go towards more biological questions, there is a fundamental problem, especially for supported lipid membranes, which is a direct protein substrate contact. The drawback of the supported membrane is when you incorporate transmembrane proteins, they are sticking out of the membrane typically by 10 nanometers. So when you deposit them on solid substrate, they always encounter the risk of protein denaturation, and also ill-defined density and orientation of membrane proteins. On the right, I show one of the data presented in my old old paper, where we stained platelet integrin alpha to be beta-3 and incorporate them into supported membranes and deposited on quartz substrates. As you can see, the picture looks very ugly, where the protein distribution is very homogeneous, and when we performed the fluorescence recovery after photobleaching experiments, we found that almost all proteins are completely pink. So to overcome this problem, we developed new models of membrane called polymer-supported membranes. So in this field, I published a review in Nature 15-16 years ago together with Eric, which was a time when I graduated from his school and moved to Heidelberg. And this morning, I checked and found that this paper was now cited more than a thousand times. In this review, we focused on the influence of soft interlayers and wedding interactions, as well as in the typical technological applications. So what is important is this polymer-supported membrane can be considered as a half model of cell-cell contact. So again, I showed this cartoon of cell-cell contact mediated by polymers, and if you cut in the middle plane and deposited on solid support, you find polymer-supported membranes. You can consider this solid substrate as another membrane, or you can consider that this is a half model. In 2007, I published quite a heavy paper in physical review letters, short, where we control the density and the thickness of the polymer layers and measured the diffusion of the integrin protein, which I showed in the previous slides. Now you see the proteins are completely uniformly distributed, unlike the one I showed here. So they are the same proteins, but now they are deposited on 10 nanometer 6-cellulose fields. So starting from the classical Einstein's equation modified by introducing the two-dimensional viscosity, as well as a dimensionless particle radius, epsilon. One can actually precisely calculate the frictional coefficient exerted on proteins, as well as the interlayer viscosity, which is regulated by the density and the thickness of the polymer interlayers. In the following accounts, we demonstrated that actually about 10s of nanometer 6 ultra thin polymer interlayers enables the maintenance of that little diffusivity, as well as the receptor functions for various transmembrane proteins. So now we come to examining the structure aspects of these polymer-supported membranes, and here we use, of course, neutral reflectivity. So what I show here in panel A and B are taken from our previous paper, showing the reflectivity of SOPCs, so sterile, oleo, and phosphatidic colline membrane deposited on thin and 6-cellulose fields. So the advantage of our cellulose is we deposit them layer by layer, and each layer has a thickness of five angstroms in dry state. So we know exactly or we can control the thickness by the layer numbers. As you can see with lines presented in each panel, the data will fit it with the slab models, and the neutral reflectivity of membrane exhibited very clear fringes in Fresnel plot, where we plotted the reflectivity multiplied by Q to the power of 4, indicating the formation of a layer structure with clear SND contrast, scattering density contrast. This is a very good sign of the formation of high-quality membrane, because we did the experiment in D2O, and since the hydrophobic interior of liquid membrane now hates water, so this gives us a clear SND contrast. Also what we found was a Gaussian roughness of membrane is much higher compared to that on solid supports, indicating the fluctuation of membrane under shallower confinement, which means indeed the theoretical prediction made by Draynkowski and Leibler in the 80s. So to see confirm that the answers or best fit results we got are unique and solid, you know, we performed also specular x-ray reflectivity. So here, you know, you note that the specular x-ray reflectivity of systems under water is in principle not possible, because unless you use an high-energy x-ray, so in these experiments we used a high-energy x-ray at ESR ref, now using the beam with an energy of 22 kilo-electron volt, which is far beyond of the elastic absorption peak or absorption band of water existing at around 8 kilo-electron volt. For the use of high-energy x-ray guarantees the high enough transmittance of beam through bulk water, the advantage we're using, say, or the difference from the neutron reflectivity is that for x-ray the contrast of the scattering length density is much poorer between the elements existing in biological systems, but the resolution is much higher. So if you compare the Q-range, you know, presented in this and the previous slide, you see that in the previous slides, you know, with neutrons you can go at most up to 0.3 reciprocal constants typically, but with x-ray you can go up to 0.5 with no problems of the counting statistics. So despite of the fact that they give different resolution and the contrast, the best fit results converged each other confirming that, you know, our solution was unique. So this brought us to the point to nail down the physical roles of biopolymers in fine adjustment of interfacial interactions. Again, you know, the disioning pressure equals zero is found at the equilibrium cell substrate distance. Since SOPC, so phosphatidicolin is petylionic and cellulose is neutrally charged, so we ignore the electrostatic interactions and focus on three major force contributors. Fundamental walls, hydration repulsion, and the hair free repulsion, which was originating from, which is originating from the thermally activated membrane fluctuations. So the important thing is all these three major forces can be quantitatively calculated. I show you two examples because van der Waas is far much complicated. So I show you the case of hair free and hydration repulsion. So hair free repulsion was described by Wolfgang Heflich, a German theorist in 1978, you know, which indicates the scaling of the repulsive pressure, you know, which is proportional to 1 over D to the power of 3. So what you can plug, what you should plug in is only the copper. This is a bending rigidity, which I mentioned, you know. This is measurable and also you can get easily from the literature in the range of 20, 30 kT. The hydration repulsion is a bit more tricky because you need to get two numbers, the offset pressure as well as the scaling. The scaling is very simple, it's an exponential decay function. These two values required to quantify the hydration repulsion can be calculated from the measurement of the film thickness as a function of osmotic pressure. This can be done by ellipsometer or, you know, X-ray nutrient effectometer coupled to a humidity channel. So what we performed or what we did here is to compare the theoretical prediction versus the experimental data. What I show here on the left and right are the results, computed results on thick, thin and thick cellulose. On thin cellulose, when we added up the van der Waas, black, hair free, green, and hydration repulsion blue, we get the net force showing in red. You see a clear crossing of the force zero. So corresponding to the equilibrium distance, which is about 140 angstroms. And if we compare this value to the values we obtained from experiments, they are in pretty much good agreement. In contrast, when you look at the data on thick cellulose, first you note that the position of zero force is much less prominent. For instance, when you look at the y-axis skating of left and right, it's different by a factor of 30. So although it looks similar, it's 30 times shallower than it appears when you put it here. So this means that on thick cellulose, hydration repulsion is more dominant so that the point of zero force is very vaguely confined compared to the syndrome. So as a result, we demonstrated that the counterplay of attractive van der Waas interaction shown in this massive black curve and hydration repulsion, which is an exponential repulsive force, confines indeed the membrane position. But in this case, the role of the sample and relation of membrane is less common. So along this line, we also made a small breakthrough where we succeeded in controlling the density and orientation of protein by spreading of native cell membranes directly onto banana supports. What we did first was the use of human red blood cells. So we took the blood of Stefan Kaufmann in my lab and also my blood. 20 years ago, it was allowed. Nowadays, it's no longer allowed. But in that time, we just took our blood and prepared the ghost cells simply by removing the cytosomes from the cell. We just throw them onto banana substrate with and without polymer and see the orientation of cells by putting antibodies, recognizing inside or outside of the cell. What we first observed was on growth and blood substrates, we saw nothing. No adhesion, no rupture, which is a kind of understandable because the surface of adult animal cells are covered with negatively charged sugars called cyanic acid. But when we did the same procedure on 10 nanometer six cells and applied the antibody recognizing the cytoplasmic domains of band three proteins, we obtained such a picture. It's a very frequently asked question, so I want to tell you that it's not the paint, so it's not a paint in a photo shop, but it's an immunofluorescence picture of my blood cells stained from inside. You see uniform and continuous coverage. On the other hand, when you apply the antibody going to the outside site extracellular domains, or glycopherin for instance, we saw nothing. So this means that now we have the scenario of complete waiting of cell membranes simply by having 10 nanometer thick polymers. The cell first adheres on the surface pulled by tension and rupture. Expose their inside and fuse with each other to form a continuous film, which is exactly as I mentioned in the introductory part about the scenario of complete waiting. This was indeed the first example of two-dimensional cell membranes, where the three D cells were transformed into quadruped two-dimensional films with a perfectly defined membrane orientation. Also we scratched the membrane from the surface and threw them into SDS page and found that the protein compositions were also kept very well. This enabled us actually to go to X-ray neutron because we now can cover macroscopically large substrates with native cell membranes spread in 2D. You can imagine that's a very nice tool to look into the structure of real cell membranes. Here I show you on the left the high-magic X-ray reflectivity data for healthy human red blood cell membranes and on the right the neutron reflectivity data. So the right was shown in Flanelplot just to show you a bit better features. We actually demonstrated that the same principle works for other native membranes such as microsome, sarcoplasmic reticulum, and plasma membrane extras. So now we are going to a very realistic biological same-surface remix and this is very intuitive that the wetting concept or wetting physics concept is very important. Like the bare glass no adhesion no wetting on soft polymer or on cellulose films no complete wetting. So this kind of wetting contrast between solid substrate and polymers supports brought me to another step to utilize wetting contrast to make a micro pattern of cell membranes. So now I have been talking about the macroscopically large substrate. For neutron reflectivity we cover the substrate with the size of five by eight centimeters 40 square centimeters substrate. Now I want to make them like a micro pattern. To do so we micro pattern cellulose support either by UV lithography or by micro contact screen the BSA, so bobbin serum algorithm which avoids the adhesion and spreading of cell membranes. Both worked out beautifully. So the top one is the same immunofluorescence picture of human red plus a membrane grid say printed onto the substrate. So only these grid parts are showing immunofluorescence positive signal and these are all backgrounds. The bottom picture shows the sarcoplasmic reticulum extracted from rabbit muscles and the green is showing the BSA you know stained with FITC and the right shows the grid of sarcoplasmic reticulum membrane which was stained by the antibodies to causing ATPs. So as you saw these papers have been published already quite some time ago and actually afterwards quite some followers came up but up to my knowledge maybe I'm wrong but you know we are the first one in the only one team that can replicate both microscopically large as well as micro patterned two-dimensional cell membranes. So now I come to the last part of my lecture where I want to show you how one can model the interactions between neighboring cell membranes mediated by a membrane-bound carb hydrate which is nothing but glycocomics you know. So as a model system we designed the stacks of synthetic and natural glycolipines which are shown on the left with structures and when we deposited thousands of these membranes you know membrane stacks and solid supports I call them as multiple supported membranes or polymer supported membranes. So as I show you know indicate with red characters or the red text so I was very lucky to get an outstanding synthetic chemist as well as microbiologist who can offer me very unique and well-defined sugar lipids. So either mimicking the lipids in animal and plant systems or in bacterial systems. As an experimental technique we utilize off-specular neutral scattering you know where we detect the scattered beam you know impinging to the plantar stocks of membrane now using 2D detectors. And what I show on the right here is the say raw detector readout you know presented in the angular coordinates gamma and omega. This experiment can be done in ILL D16 beamline you know and this was strongly supported by the local scientist Bruno de Me. So the advantage of this system is that owing to the planar geometry of the sample one can discriminate the momentum transfer perpendicular and parallel to the membrane plane. Simply from the geometric consideration so you see only gamma and omega are in this equation so this means that the specular signals reflects the vertical structure information and inter-membrane potential while the off-specular intensity reflects the lateral structure ordering and membrane mechanics. So after you measure the signals using nice 2D detector you have to work a little bit. So here we have to compute the scattering signals. To do so we have to revisit we had to revisit the basic framework of discrete symmetric Hamiltonian which was theoretically written by Leibniz and Lipposti in the 80s. Here the Hamiltonian contains two key parameters. First is the compression model B so that's a vertical compression model B and the bending rigidity copper. On the other hand when you see the data from the analytic viewpoint the scattering function from stratified rough interfaces within the first bond approximation is written in that form. Now this was written by Sina in the mid 90s and although this looks very complicated most important bits here is this G as a function of R called displacement correlation function and this displacement correlation function is determined by two key mechanical parameters lambda and eta. Lambda is named the jump parameter and no eta carrier parameter and what's notable is both lambda and eta are the products of copper and B. So this means from the analysis if you can find the best combination of lambda and eta you can calculate the copper and B which are corresponding one by one. What I want to emphasize here is there is an important the number we need to introduce called R that's a cut-off radius which represents the finite size effect. If you do not limit this lower limit the displacement correlation function becomes dispersive so you know the roughness becomes infinitely large when you make the separation between two points larger and larger. This size can be determined by the size of the membrane patches which you can determine by optical microscopy or atomic force microscopy. So plotting these numbers you know we actually computed the scattering signals. What I show in this slide is the direct comparison of the second black sheet from experiments and simulations. As you can agree probably yeah this is a very nice agreement and what I want to emphasize here is the introduction of the cut-off radius indeed allows a full calculation of scattering functions with no floating parameters. So this means if you plug in instrument resolution function and also the cut-off radius you can be fully quantitative. This is an advantage compared to the previous approach where you have to approximate something to fit the signal. So this technique was applied to shed light on the influence of the carb-hydrate confirmation on the mechanical properties. We started from a simple glycolipids possessing glucose and galactose unit on the head. One is gentioburus which is mimicking the lipotheic acid in gram-positive bacteria and the other one is lactose lipid and the difference only difference between two lipids are the glycosylic junction either 1.6 or 1.4. By comparing two modally you find that simply by bending the cylindrical lipid head groups through the bent one the bending rigidity decreases by a factor of 4. On the other hand when we look at and extend this idea to a bit more complex lipids you know we performed experiments with Daniel Veratz. He's an organic chemist in Göttingen and now in Branschweich. He designed the GB3 lipids which are known to be the glycolipids or galactoside accumulating in the eyes of patients suffering from fabric disease. So here you have the lactose unit plus one more branch here. So by adding just one branch you see that the bending rigidity decreased by a factor of 2 and the bending rigidity decreased by a factor of 2 and compression modulus as well. This was extended to more complex glycolipids by taking the bacterial lipopolysaccharides. That's what I collaborated with Klaus Brandenburg from Research Center of Bostel. That's a northern part of northern edge of Germany where he could actually extract the high impunity lipopolysaccharides from various bacterial mutants by chopping the sugar chains at the position of interest like I indicated with black lines. One can test a very short one called lipid A up to the rough mutants with the inner and outer membrane cores as well as the wild type lipopolysaccharides including polysaccharides chains. So what we found, so Rafael my fellow postdoc found was the bending rigidity and compression modulus is motivated by the genetic mutation of sugar chains. For example if you compare the shortest lipid A and the longest mutant APSRA which has I think 8 or 10 additional sugars, we see that the compression modulus decreased by a factor of 4 because these sugars are softer so that they are say you know the compression modulus gets smaller. So we showed that the off-specular nutrient scattering is a good tool or powerful tool that enables the influence of chemical genetic and epigenetic modification on membrane mechanics. To conclude so I hope that I could give you a flavor that cell membranes are just not a boundary but doing more you know. So they act as biological reaction hubs regulating many critical functions and the unique mechanical properties of cell membrane shown by red blood cells are tightly coupled to biological functions you know which can be described within the framework of smectically crystals. Then the recent development as I showed in the last part of both biochemical and instruments enables to design more precise biological relevant models of cell membranes. I started from very simple supported membrane developed 35 years ago and now we are dealing with much more realistic and complex mimics of cell surfaces. The combination of neutron and x-ray reflectivity is also a very good tool to probe the detailed structure of complex and buried interfaces under water enabling the interplace of interfacial forces regulating the wetting contents. And finally the specular scattering though is a good tool that you know quantitatively unravels the how chemical genetic and epigenetic changes in membrane boron carb hydrates would affect the mechanics of membranes. So since this is a lecture so I do not want to give a acknowledgement to the funding sources but you know I want to convince you that the near future development of instruments will enable us to tackle more challenging problems using neutron. So instead of talking to the money I want to thank my students postdocs and colleagues now many of them got the permanent faculty positions outside Germany or inside Germany as well and you know the supports by the large facilities like INL and ASF were instrumental and as you see the interdisciplinary collaboration with chemist medical scientists as well as microbiologists made us very unique and strong and I thank for your attention. Tommy I'm done. Okay thank you very much and I think there is time for a lot of questions. I hope there are some. Yeah you have some questions here waiting waiting waiting. So I think it's very interesting that you actually can look at real cell membrane. I think we always are struggling we always are doing the UPC with 5% the UPS or whatever so I think it's very useful to be able to look at the real real membrane and so on and sometimes one can argue it's complex but complexity is sometimes necessary I think. Yeah because you have quite a sense of biology. Yeah. So how much? Who has the question? Who is the question? Yes thank you very much for a great presentation and I just have a general question maybe to hear your point of view like what do you think are the main challenges right now where the research is going in the this like interface lipid kind of systems because it feels like you are also a pioneer in this in this field right now so maybe some comments. Yes so one direction that I think is interesting is to going through the influence of disease and aging on the membrane you know because you know many biochemical studies suggested that you know the structure as well as the composition of lipid membranes are modulated by aging and diseases and indeed you know what I plan to gather with this form is to spread the red blood cells infected by malaria parasites on polymer supports and look at how the malaria parasites modulate the site skeletons as well as membrane proteins and for the mechanics using microfluidics and other methods I really demonstrated that the disease actually modulated the mechanics of the membranes a lot so now I want to shed light on the structure in another direction that I would be very keen on testing or going myself you know until the end of my career so I'm not too old yeah so is you know to go into the dynamics so now I'm looking more or less now on the equilibrium systems not because of the time window that we can assess but maybe you heard that lecture of Vicky yeah about the inelastic neutral scattering and spin echo so back in the days you know using ILM I mean if we want to look at the mechanical properties of cell membranes you know using spin echo I mean I needed two weeks or a month not in the reality and we need to have tons of samples but no not if we imagine the situation of using the spreadation source I think that's something that I'm definitely interested in doing myself there as soon as the instrument is ready and that's actually the one of the reasons why you know I am coming in close contact you know so I bring some strange systems with a bit of idea of physics and you know the experts like Tommy and you know Vicky and no these people can support us to realize this idea yeah and in synchrotron it's also interesting look at the dynamics using x-ray photocollation spectroscopy and other methods and this was also very much limited in the past because of the coherency of the x-ray as well as the intensity and I was luckily involved in the upgrade committee of ESRF and we used as influence the panel to strengthen the coherence so in two years now or five years now I think they should have 30 times higher coherence which enables us to really do xpcs for also biological memory so you know to conclude you know my interest was to aging diseases so as well as to dynamics I think these two are what I think to be an exciting field in the future thank you very much and I'm also thinking that this new like methods with time time resolved experiments and free electron lasers are also like enabling like another level of this dynamic and structural motion analysis yes yeah so if FEL is a nasty method you know because you just burn the stuff with an intensive pulse and observing the shot but on the other hand I mean this is a very you know cute methods which I didn't try myself yet but I think this opens up also the future yeah thank you so much you're welcome and any more comments oh there is a hand Ying I have a basic question about the interplace of interfacial forces regulating the wetting contact and I remember you mentioned many forces including the hydrogen force and can you give a comment on what is hydrogen force is it only and persists between the bilayer membranes and what is it originated from yeah so hydration repulsion is a generic force you know and I think in Lund Hogan Venastro is one of the pioneers and another one in the US is no adrian precision and I think I just mentioned about adrian which I felt a bit dangerous here yeah okay so this is how the hydration project decays so D is a distance and lambda is a characteristic decay length and this is a long set so I didn't get your question so this is not a special thing for lipid bilayer or anything you know for instance you know when you say measure the hydration of polymers in controlled humidity what you see is nothing but the hydration repulsion you know so it exists between and hydrophobic hydrophobic surfaces hydrophilic hydrophilic surface because we are looking at water or hydrophilic parts and so it is not necessary to be surface it can also be like hydrophobic on some parts yeah but it is decaying over distance from the surface you see it's not the bulk osmotic pressure so of course now in bulk you know we can describe the chemical potential of water simply by osmotic pressure well when you go back to high school not high school but the first class physical chemistry you know you find found Hof's law you know the osmotic pressure is given by her kt divided by molar volume of water times natural lock of reactive humidity well done yes that's how i remember from the textbook yeah so there you know you can tune the osmotic pressure namely the chemical potential water in the system by changing the humidity what i'm talking here is how the repulsive force now decays in the presence of water yeah i hope you get it yes and so so what you find here is that in combination of all these forces that's why you can get the blood cell rupture in the surface no here i'm talking how the distance between the membranes are kept constant because now you know when the force is balanced there is no motion newton's first law yeah yeah so in order to have an stable equilibrium distance you need to achieve a balance of different forces if this is destroyed then you have continuous sinning which results in the collapse or your membrane is just separated more and more and they are gone and that's what i showed by computation here as i said so you know on thick cell loss you see the scaling of y so this is a pressure is very shallow you know so the deep here is compared no negatively small compared to this one yeah but still you know one can see how this osmotic repulsion is pushing away this the system from the deep well or fundaverse attraction yeah this is what i showed yes so what is the magic that's rupture that make this break the cell and onto the surface yeah so in order to write you know you need to achieve the gain of the free energy so this is described by surface tension and second is this no no force balance uh where it is so what is at the very beginning i explained this balance oh wait a moment wait a moment i think you probably are not so much familiar with physics so yeah i mean okay so this is the first point this is from the viewpoint of surface free energy but if just look at this now you can imagine that every hydrophilic surface should do the job but here well okay so if you have quartz or glass substrates the interaction is just purely repulsive so those cells do not adhere on the surface okay so they are just repelled when you have some polymers no you have the good balance of attraction and repulsion they sit on the surface and since they are hydrophilic they wet yeah so this is very similar well not similar so the first part is very similar if you consider the spreading of lipids on hydrophilic substrate and hydrophobic substrate well if you have hydrophilic substrate you have the spreading of bilayer but if you have a hydrophobic surface you know you have the spreading of lipid monolayer because the chain of hydrophobic tears that loves to spread on the surface by this no balance of the spreading coefficient