# name
Nanoscale dielectric mapping of biomembranes with in-liquid Scanning Dielectric Microscopy

# metadata.dc.contributor.other[].value
Universitat de Barcelona. Departament d'Enginyeria Electrònica i Biomèdica

# metadata.dc.description.abstract[].value
[eng] The structure and physicochemical properties of biomembranes are fundamental for the functioning of cells, and many pathologies have been associated with their alteration (cancer, neurodegenerations, obesity, etc.) 1, 2. For this reason, biomembranes have been the subject of intensive research. Yet, there is still limited knowledge of biomembranes, which show a heterogeneous structure at the nanoscale that is naturally present in cells, and determines many of the phenomena occurring through them at the molecular level 3, 4.

Due to their prominent role in Electrophysiology, electrical properties are among the more relevant physical properties of biomembranes. Most often, attention is paid to biomembranes' conduction properties, and the role played in them by ionic channels. However, biomembranes' dielectric properties are also of central interest in bioelectric phenomena, and a powerful reporter of membranes' composition, which can be exploited to develop label-free mapping methods. Yet, most of the available techniques have addressed the dielectric membrane properties in bulk solutions and at the level of single cells (micrometers), thus lacking spatial resolution. In other cases, they make use of exogenous labels, as in the case of spin paramagnetic resonance 5, 6 and fluorescence microscopy 7, 8, 9, 10, 11.
In recent years, the Nanoscale Bioelectric Characterization group at IBEC, as well as other groups, have developed some Scanning Probe Microscopies (SPMs) based techniques to attempt the dielectrical characterization at the nanoscale 12,  13, 14, 15, 16 and
applied them to biomembranes 17, 18, 19, 20, 21 and other biosystems 22, 12, 23, 24, 25, 26, 27. Initially, these techniques were implemented to be operated in air environment, but lately they were also extended to liquid environment 28, 29. The implementation of in-liquid Scanning  Dielectric  Microscopy  (SDM)  paved  the  way  to  the  accurate  dielectric
 
characterization of biomembranes at the nanoscale, in their physiological environment and in a label-free way 28, 29.
In-liquid SDM is based on measuring the electrostatic force acting on a nanometric probe under application of a modulated voltage between the tip and a conductive substrate, on top of which the sample is sitting. As compared to the standard SDM in air, operation in liquid environment requires several modifications. In terms of set-up, one needs to apply frequencies above the dielectric relaxation frequency of the electrolyte. Significant changes are also necessary for the modelling part 30.

This work of thesis takes advantage of the latest developments of in-liquid SDM to characterize the dielectric properties of heterogeneous model and natural purified membranes systems in liquid. In this framework, new knowledge has been gained about imaging in liquid conditions with SDM, e.g. about the prominent electrostatic finite size effect and different models have been tested and optimized for the analysis of the measurements.

First, I focused on characterizing mono- and bicomponent planar supported bilayer lipid mixtures containing cholesterol, providing a first proof-of-concept of the label-free mapping capabilities of the technique in liquid media, extending earlier work done in air on nanoparticles 12. This study allowed gaining information on the composition of sub- micrometric membrane domains in liquid environment 31 and to provide reliable values of the intrinsic dielectric properties of DOPC and DOPC/cholesterol compositions, about which there was some debate in the literature. The low values obtained are responsible for membranes’ low permeability to ions, in agreement with previous studies on monocomponent biomembranes 29. Our results allow speculating on fundamental properties of lipid bilayers like viscosity and hydration of cholesterol-containing layers.

Afterwards, we extended the methods to deal with more complex biomembrane 3D structures, such as liposomes 32. Liposomes with few hundred nanometers in height have
 
been successfully imaged by in-liquid SDM, showing a sensitivity comparable to the one for flat biomembranes only a few nanometers thin. Once again, the dielectric properties of the liposomes’ membrane were precisely extracted, this time in a more natural configuration of the biomembrane. This study also highlighted the technique’s sub- surface capabilities in the liquid environment, demonstrated earlier only in air measurements 33, 34, 35, 36, 37, 38, 39. This capability enabled to obtain in a label-free way the lamellarity of liposomes, a crucial parameter in liposomes technology. The developed methodology has the potential to be used to screen a myriad of different compositions of liposomes (shell and core), since in-liquid SDM was shown to be sensitive to the dielectric properties of the membrane but also to the conductivity of the medium inside the liposomes. This accomplishment was essential to evaluate its future application to living cells and constitutes one of the main achievements of this work.

During the thesis, I also draw my attention to the dielectric characterization of natural purified membranes in liquid environment. As a test example, we focused on the case of the purple membrane (PM), which had previously been studied in air environment 40, 20,
19. PMs are constituted by the protein bacteriorhodopsin (BR) arranged in a crystal lattice, and lipids in a ratio 10:1 lipids:proteins. However, an unsolved uncertainty in determining the real topography of supported PM patches in the liquid environment, which can also display a concave surface, made the dielectric quantification problematic, and further explorations will be necessary to provide reliable values of the permittivity of these layers in liquid media.

In summary, the objective and common thread connecting all the chapters of this work has been implementing and demonstrating the capabilities of in-liquid SDM in the dielectric characterization of bio-membranes, model and natural systems, with nanoscale spatial resolution.
 
I believe that this work laid the ground for elucidating the structure and dielectric properties of more complex membranes systems and their associated electric phenomena, e.g. conduction. Preliminary studies of the cell membrane directly on living neuroblastoma cells, in low concentration MOPS buffer, were carried out in collaboration with Maria Elena Piersimoni, PhD student at Imperial College, London. The group is now collaborating with experts in the field and trying to develop new algorithms, fundamental to extend the methods to living cells.

In addition to the main objective of my thesis, I also participated in a side project concerning the in-liquid SDM characterization of an operative EGOFET transistor 41, crucial for optimizing the devices and gain a better understanding of the transduction mechanism with biological entities. One of the newest frontiers of such technology is indeed to use supported lipid bilayers for bio-sensing purposes 42.




References:

(1)	Lauwers, E.; Goodchild, R.; Verstreken, P. Membrane Lipids in Presynaptic Function and Disease. Neuron 2016, 90 (1), 11–25. https://doi.org/10.1016/j.neuron.2016.02.033.
(2)	Ashrafuzzaman, M., Tuszynski, J. Membrane-Related Diseases, Springer-V.; Springer- Verlag Berlin Heidelberg 2012, 2012.
(3)	Mueller, P.; Rudin, D. O. Resting and Action Potentials in Experimental Bimolecular Lipid Membranes. J. Theor. Biol. 1968, 18 (2), 222–258. https://doi.org/10.1016/0022- 5193(68)90163-x.
(4)	Hodgkin, A. L.; Huxley, A. F. A Quantitative Description of Membrane Current and Its Application to Conduction and Excitation in Nerve. J Physiol. 1952, 117, 500–544. https://doi.org/10.1109/ICCCT2.2017.7972284.
(5)	Kurad, D.; Jeschke, G.; Marsh, D. Lipid Membrane Polarity Profiles by High-Field EPR.
Biophys. J. 2003, 85 (2), 1025–1033. https://doi.org/10.1016/S0006-3495(03)74541-X.
(6)	Marsh, D. Polarity and Permeation Profiles in Lipid Membranes. Proc. Natl. Acad. Sci. U. S. A. 2001, 98 (14), 7777–7782. https://doi.org/10.1073/pnas.131023798.
(7)	Huang, H.; McIntosh, A. L.; Atshaves, B. P.; Ohno-Iwashita, Y.; Kier, A. B.; Schroeder, F. Use of Dansyl-Cholestanol as a Probe of Cholesterol Behavior in Membranes of Living Cells. J. Lipid Res. 2010, 51 (5), 1157–1172. https://doi.org/10.1194/jlr.M003244.
(8)	Parasassi, T.; De Stasio, G.; Ravagnan, G.; Rusch, R. M.; Gratton, E. Quantitation of Lipid Phases in Phospholipid Vesicles by the Generalized Polarization of Laurdan
 
Fluorescence. Biophys. J. 1991, 60 (1), 179–189. https://doi.org/10.1016/S0006-
3495(91)82041-0.
(9)	Signore G., Abbonato G., Storti B., Stöckl M., Subramaniam V., B. R. Imaging the Static Dielectric Constant in Vitro and in Living Cells by a Bioconjugable GFP Chromophore Analog. ChemComm 2013, 49 (1723). https://doi.org/10.1039/b000000x.
(10)	Demchenko, A. P.; Mély, Y.; Duportail, G.; Klymchenko, A. S. Monitoring Biophysical Properties of Lipid Membranes by Environment-Sensitive Fluorescent Probes. Biophys. J. 2009, 96 (9), 3461–3470. https://doi.org/10.1016/j.bpj.2009.02.012.
(11)	Epand, R. M.; Kraayenhof, R. Fluorescent Probes Used to Monitor Membrane Interfacial Polarity. Chem. Phys. Lipids 1999, 101 (1), 57–64. https://doi.org/10.1016/S0009- 3084(99)00055-9.
(12)	Fumagalli, L.; Esteban-Ferrer, D.; Cuervo, A.; Carrascosa, J. L.; Gomila, G. Label-Free Identification of Single Dielectric Nanoparticles and Viruses with Ultraweak Polarization Forces. Nat. Mater. 2012, 11 (9), 808–816. https://doi.org/10.1038/nmat3369.
(13)	Fumagalli, L.; Ferrari, G.; Sampietro, M.; Gomila, G. Dielectric-Constant Measurement of Thin Insulating Films at Low Frequency by Nanoscale Capacitance Microscopy. Appl. Phys. Lett. 2007, 91 (24), 15–18. https://doi.org/10.1063/1.2821119.
(14)	Gomila, G.; Toset, J.; Fumagalli, L. Nanoscale Capacitance Microscopy of Thin Dielectric Films. J. Appl. Phys. 2008, 104 (2). https://doi.org/10.1063/1.2957069.
(15)	Fumagalli, L.; Gramse, G.; Esteban-Ferrer, D.; Edwards, M. A.; Gomila, G. Quantifying the Dielectric Constant of Thick Insulators Using Electrostatic Force Microscopy. Appl. Phys. Lett. 2010, 96 (18), 88–91. https://doi.org/10.1063/1.3427362.
(16)	Fumagalli, L.; Ferrari, G.; Sampietro, M.; Casuso, I.; Martínez, E.; Samitier, J.; Gomila, G. Nanoscale Capacitance Imaging with Attofarad Resolution Using Ac Current Sensing Atomic Force Microscopy. Nanotechnology 2006, 17 (18), 4581–4587. https://doi.org/10.1088/0957-4484/17/18/009.
(17)	Gramse, G.; Schönhals, A.; Kienberger, F. Nanoscale Dipole Dynamics of Protein Membranes Studied by Broadband Dielectric Microscopy. Nanoscale 2019, 11 (10), 4303–4309. https://doi.org/10.1039/c8nr05880f.
(18)	Knapp, H. F.; Mesquida, P.; Stemmer, A. Imaging the Surface Potential of Active Purple Membrane. Surf. Interface Anal. 2002, 33, 108–112. https://doi.org/10.1002/sia.1172.
(19)	Gramse, G.; Casuso, I.; Toset, J.; Fumagalli, L.; Gomila, G. Quantitative Dielectric Constant Measurement of Thin Films by DC Electrostatic Force Microscopy. Nanotechnology 2009, 20 (39). https://doi.org/10.1088/0957-4484/20/39/395702.
(20)	Fumagalli, L.; Ferrari, G.; Sampietro, M.; Gomila, G. Quantitative Nanoscale Dielectric Microscopy of Single-Layer Supported Biomembranes. Nano Lett. 2009, 9 (4), 1604– 1608. https://doi.org/10.1021/nl803851u.
(21)	Dols-Perez, A.; Gramse, G.; Caló, A.; Gomila, G.; Fumagalli, L. Nanoscale Electric Polarizability of Ultrathin Biolayers on Insulating Substrates by Electrostatic Force Microscopy. Nanoscale 2015, 7, 18327–18336. https://doi.org/10.1039/x0xx00000x.
(22)	Cuervo, A.; Dans, P. D.; Carrascosa, J. L.; Orozco, M.; Gomila, G.; Fumagalli, L. Direct Measurement of the Dielectric Polarization Properties of DNA. Proc. Natl. Acad. Sci. U. S. A. 2014, 111 (35). https://doi.org/10.1073/pnas.1405702111.
(23)	Biagi, M. C.; Fabregas, R.; Gramse, G.; Van Der Hofstadt, M.; Juárez, A.; Kienberger, F.; Fumagalli, L.; Gomila, G. Nanoscale Electric Permittivity of Single Bacterial Cells at
 
Gigahertz Frequencies by Scanning Microwave Microscopy. ACS Nano 2016, 10 (1), 280–288. https://doi.org/10.1021/acsnano.5b04279.
(24)	Esteban-Ferrer, D.; Edwards, M. A.; Fumagalli, L.; Juárez, A.; Gomila, G. Electric Polarization Properties of Single Bacteria Measured with Electrostatic Force Microscopy. ACS Nano 2014, 8 (10), 9843–9849. https://doi.org/10.1021/nn5041476.
(25)	Van Der Hofstadt, M.; Fabregas, R.; Millan-Solsona, R.; Juarez, A.; Fumagalli, L.; Gomila,
G. Internal Hydration Properties of Single Bacterial Endospores Probed by Electrostatic Force Microscopy. ACS Nano 2016, 10 (12), 11327–11336. https://doi.org/10.1021/acsnano.6b06578.
(26)	Checa, M.; Millan-Solsona, R.; Blanco, N.; Torrents, E.; Fabregas, R.; Gomila, G. Mapping the Dielectric Constant of a Single Bacterial Cell at the Nanoscale with Scanning Dielectric Force Volume Microscopy. Nanoscale 2019, 11 (43), 20809–20819. https://doi.org/10.1039/c9nr07659j.
(27)	Lozano, H.; Fabregas, R.; Blanco-Cabra, N.; Millán-Solsona, R.; Torrents, E.; Fumagalli, L.; Gomila, G. Dielectric Constant of Flagellin Proteins Measured by Scanning Dielectric Microscopy. Nanoscale 2018, 10 (40), 19188–19194. https://doi.org/10.1039/c8nr06190d.
(28)	Gramse, G.; Edwards, M. A.; Fumagalli, L.; Gomila, G. Dynamic Electrostatic Force Microscopy in Liquid Media. Appl. Phys. Lett. 2012, 101 (21). https://doi.org/10.1063/1.4768164.
(29)	Gramse, G.; Dols-Perez, A.; Edwards, M. A.; Fumagalli, L.; Gomila, G. Nanoscale Measurement of the Dielectric Constant of Supported Lipid Bilayers in Aqueous Solutions with Electrostatic Force Microscopy. Biophys. J. 2013, 104 (6), 1257–1262. https://doi.org/10.1016/j.bpj.2013.02.011.
(30)	Millán, R.; Checa, M.; Fumagalli, L.; Gomila, G. Mapping the Capacitance of Self- Assembled Monolayers at Metal/Electrolyte Interfaces at the Nanoscale by In-Liquid Scanning Dielectric Microscopy. Nanoscale 2020, 12 (40), 20658–20668. https://doi.org/10.1039/d0nr05723a.
(31)	Di Muzio, M.; Millan-Solsona, R.; Borrell, J. H.; Fumagalli, L.; Gomila, G. Cholesterol Effect on the Specific Capacitance of Submicrometric DOPC Bilayer Patches Measured by In-Liquid Scanning Dielectric Microscopy. Langmuir 2020, 36 (43), 12963–12972. https://doi.org/10.1021/acs.langmuir.0c02251.
(32)	Di Muzio, M.; Millán, R.; Gomila, G. Electrical Properties and Lamellarity of Single Liposomes Measured by In-Liquid SDM. [in Prep.
(33)	Fumagalli, L.; Esfandiar, A.; Fabregas, R.; Hu, S.; Ares, P.; Janardanan, A.; Yang, Q.; Radha, B.; Taniguchi, T.; Watanabe, K.; et al. Anomalously Low Dielectric Constant of Confined Water. Science (80-. ). 2018, 360 (6395), 1339–1342. https://doi.org/10.1126/science.aat4191.
(34)	Fabregas, R.; Gomila, G. Dielectric Nanotomography Based on Electrostatic Force Microscopy: A Numerical Analysis. J. Appl. Phys. 2020, 127 (2). https://doi.org/10.1063/1.5122984.
(35)	Castañeda-Uribe, O. A.; Reifenberger, R.; Raman, A.; Avila, A. Depth-Sensitive Subsurface Imaging of Polymer Nanocomposites Using Second Harmonic Kelvin Probe Force Microscopy. ACS Nano 2015, 9 (3), 2938–2947. https://doi.org/10.1021/nn507019c.
(36)	Riedel, C.; Alegra, A.; Schwartz, G. A.; Arinero, R.; Colmenero, J.; Senz, J. J. On the Use of
 
Electrostatic Force Microscopy as a Quantitative Subsurface Characterization Technique: A Numerical Study. Appl. Phys. Lett. 2011, 99 (2), 99–101. https://doi.org/10.1063/1.3608161.
(37)	Zhao, M.; Gu, X.; Lowther, S. E.; Park, C.; Jean, Y. C.; Nguyen, T. Subsurface Characterization of Carbon Nanotubes in Polymer Composites via Quantitative Electric Force Microscopy. Nanotechnology 2010, 21 (22). https://doi.org/10.1088/0957- 4484/21/22/225702.
(38)	Cadena, M. J.; Misiego, R.; Smith, K. C.; Avila, A.; Pipes, B.; Reifenberger, R.; Raman, A. Sub-Surface Imaging of Carbon Nanotube-Polymer Composites Using Dynamic AFM Methods. Nanotechnology 2013, 24 (13). https://doi.org/10.1088/0957- 4484/24/13/135706.
(39)	Alekseev, A.; Chen, D.; Tkalya, E. E.; Ghislandi, M. G.; Syurik, Y.; Ageev, O.; Loos, J.; De With, G. Local Organization of Graphene Network inside Graphene/Polymer Composites. Adv. Funct. Mater. 2012, 22 (6), 1311–1318. https://doi.org/10.1002/adfm.201101796.
(40)	Casuso, I.; Fumagalli, L.; Gomila, G.; Padrós, E. Nondestructive Thickness Measurement of Biological Layers at the Nanoscale by Simultaneous Topography and Capacitance Imaging. Appl. Phys. Lett. 2007, 91 (6), 063111–063113. https://doi.org/10.1063/1.2767979.
(41)	Kyndiah, A.; Checa, M.; Leonardi, F.; Millan-Solsona, R.; Di Muzio, M.; Tanwar, S.; Fumagalli, L.; Mas-Torrent, M.; Gomila, G. Nanoscale Mapping of the Conductivity and Interfacial Capacitance of an Electrolyte-Gated Organic Field-Effect Transistor under Operation. Adv. Funct. Mater. 2020, 2008032, 1–8. https://doi.org/10.1002/adfm.202008032.
(42)	Lubrano, C.; Matrone, G. M.; Iaconis, G.; Santoro, F. New Frontiers for Selective Biosensing with Biomembrane-Based Organic Transistors. ACS Nano 2020, 14 (10), 12271–12280. https://doi.org/10.1021/acsnano.0c07053.

# metadata.dc.description.abstract[].value
[spa] La estructura y propiedades fisicoquímicas de las biomembranas son fundamentales para el funcionamiento de las células, y muchas patologías (cáncer, neurodegeneraciones, obesidad, etc.) 1, 2 se han asociado a su alteración . Por este motivo, las biomembranas han sido objeto de intensas investigaciones. Sin embargo, todavía existe un conocimiento limitado de las biomembranas, que muestran una estructura heterogénea a la nanoescala, que en realidad son las que están presentes de forma natural en las células y determinan muchos de los fenómenos que ocurren a través de ellas a nivel molecular 3, 4.
Las propiedades eléctricas, debido a su papel destacado en la electrofisiología, se encuentran entre las propiedades físicas más relevantes de las biomembranas. La mayoría de las veces se presta atención a las propiedades de conducción de las biomembranas y al papel que juegan en ellas los canales iónicos. Sin embargo, las propiedades dieléctricas de la biomembrana también son de interés central en los fenómenos bioeléctricos y, también, pueden considerarse como un poderoso indicador de la composición de la biomembrana, que puede aprovecharse para desarrollar métodos de mapeo sin marcadores.
Este trabajo de tesis aprovecha los últimos desarrollos de microscopía dieléctrica de sonda de barrido (SDM) en líquido para caracterizar las propiedades dieléctricas de sistemas de membranas de modelo heterogéneo y membranas naturales purificadas en líquido. En este trabajo, se han obtenido nuevos conocimientos sobre la técnica de SDM en líquido, como por ejemplo sobre el prominente efecto electrostático de tamaño finito. También se han probado y optimizado diferentes modelos para el análisis de las medidas.
Primero, nos concentramos en caracterizar mezclas de bicapa lipídicas soportadas mono y bicomponente que contienen colesterol, proporcionando una primera prueba de concepto de las capacidades de mapeo sin etiqueta de la técnica en líquido y ampliando el trabajo realizado anteriormente en aire sobre nanopartículas 5.
Posteriormente, ampliamos los métodos para tratar con estructuras 3D de biomembranas más complejas, como los liposomas 6. Mediante SDM en liquído, se han obtenido imágenes de liposomas de unas pocas docenas de nanómetros de altura. Una vez más, se extrajeron con precisión las propiedades dieléctricas de la biomembrana de los liposomas, esta vez en una configuración más natural de la biomembrana. Este estudio también destacó las capacidades subsuperficiales de la técnica en líquido,
 
demostradas anteriormente solo en medidas de aire 7, 8, 9, 10, 11, 12, 13, y permitió obtener de forma ‘label-free’ la lamelaridad de los liposomas, un parámetro crucial en esta tecnología.
Este trabajo sentó las bases para dilucidar la estructura y las propiedades dieléctricas de sistemas de membranas más complejos, incluidas células vivas, y sus fenómenos eléctricos asociados, como por ejemplo la conducción.


References:

(1)	Lauwers, E.; Goodchild, R.; Verstreken, P. Membrane Lipids in Presynaptic Function and Disease. Neuron 2016, 90 (1), 11–25. https://doi.org/10.1016/j.neuron.2016.02.033.
(2)	Ashrafuzzaman, M., Tuszynski, J. Membrane-Related Diseases, Springer-V.; Springer- Verlag Berlin Heidelberg 2012, 2012.
(3)	Mueller, P.; Rudin, D. O. Resting and Action Potentials in Experimental Bimolecular Lipid Membranes. J. Theor. Biol. 1968, 18 (2), 222–258. https://doi.org/10.1016/0022- 5193(68)90163-x.
(4)	Hodgkin, A. L.; Huxley, A. F. A Quantitative Description of Membrane Current and Its Application to Conduction and Excitation in Nerve. J Physiol. 1952, 117, 500–544. https://doi.org/10.1109/ICCCT2.2017.7972284.
(5)	Fumagalli, L.; Esteban-Ferrer, D.; Cuervo, A.; Carrascosa, J. L.; Gomila, G. Label-Free Identification of Single Dielectric Nanoparticles and Viruses with Ultraweak Polarization Forces. Nat. Mater. 2012, 11 (9), 808–816. https://doi.org/10.1038/nmat3369.
(6)	Di Muzio, M.; Millán, R.; Gomila, G. Electrical Properties and Lamellarity of Single Liposomes Measured by In-Liquid SDM. [in Prep.
(7)	Fumagalli, L.; Esfandiar, A.; Fabregas, R.; Hu, S.; Ares, P.; Janardanan, A.; Yang, Q.; Radha, B.; Taniguchi, T.; Watanabe, K.; et al. Anomalously Low Dielectric Constant of Confined Water. Science (80-. ). 2018, 360 (6395), 1339–1342. https://doi.org/10.1126/science.aat4191.
(8)	Fabregas, R.; Gomila, G. Dielectric Nanotomography Based on Electrostatic Force Microscopy: A Numerical Analysis. J. Appl. Phys. 2020, 127 (2).
 
https://doi.org/10.1063/1.5122984.

(9)	Castañeda-Uribe, O. A.; Reifenberger, R.; Raman, A.; Avila, A. Depth-Sensitive Subsurface Imaging of Polymer Nanocomposites Using Second Harmonic Kelvin Probe Force Microscopy. ACS Nano 2015, 9 (3), 2938–2947. https://doi.org/10.1021/nn507019c.
(10)	Riedel, C.; Alegra, A.; Schwartz, G. A.; Arinero, R.; Colmenero, J.; Senz, J. J. On the Use of Electrostatic Force Microscopy as a Quantitative Subsurface Characterization Technique: A Numerical Study. Appl. Phys. Lett. 2011, 99 (2), 99–101. https://doi.org/10.1063/1.3608161.
(11)	Zhao, M.; Gu, X.; Lowther, S. E.; Park, C.; Jean, Y. C.; Nguyen, T. Subsurface Characterization of Carbon Nanotubes in Polymer Composites via Quantitative Electric Force Microscopy. Nanotechnology 2010, 21 (22). https://doi.org/10.1088/0957- 4484/21/22/225702.
(12)	Cadena, M. J.; Misiego, R.; Smith, K. C.; Avila, A.; Pipes, B.; Reifenberger, R.; Raman, A. Sub-Surface Imaging of Carbon Nanotube-Polymer Composites Using Dynamic AFM Methods. Nanotechnology 2013, 24 (13). https://doi.org/10.1088/0957- 4484/24/13/135706.
(13)	Alekseev, A.; Chen, D.; Tkalya, E. E.; Ghislandi, M. G.; Syurik, Y.; Ageev, O.; Loos, J.; De With, G. Local Organization of Graphene Network inside Graphene/Polymer Composites. Adv. Funct. Mater. 2012, 22 (6), 1311–1318. https://doi.org/10.1002/adfm.201101796.

# metadata.dc.source[].value
Tesis Doctorals - Departament - Enginyeria Electrònica i Biomèdica

# metadata.dc.title[].value
Nanoscale dielectric mapping of biomembranes with in-liquid Scanning Dielectric Microscopy