Development of a method to investigate protein binding on nanostructured surfaces - fabrication of surfaces and evaluation of analysis techniques

Abstract

The nanoscale surface topography of artificial materials, independent of surface chemistry, is known to significantly influence protein adsorption. Analyzing protein behavior in the presence of nanostructured surfaces is important in the areas such as medical implants development and biosensors. Surfaces with well-defined nanoscale topography can be fabricated by the available nanofabrication techniques and thus be used as model systems for protein assays. Well-ordered arrays of nanoholes with different diameters and the same depth were fabricated in silicon in order to study the effect of surface nanotopography on protein adsorption systematically. Electron beam lithography and reactive ion etching were used to fabricate small nanoholes down to 45 nm in diameter and 50 nm in depth. We have developed a method to study the interaction of a monolayer of human fibrinogen with nanoholes. Our approach includes using the intrinsic fluorescence of proteins which allows us to observe the natural behavior of proteins induced by nanoholes, and atomic force microscopy for further understanding of the interaction. In addition, Raman spectroscopy was evaluated as a technique to characterize the proteins adsorption to the planar and nanostructured surfaces. However Raman spectroscopy was not sensitive enough to detect any signal at the protein coverage used in this study. The result of the fluorescence measurements suggests that as the size of the nanoholes approaches 45 nm, fibrinogen adsorption is significantly increased.

Popular Abstract

Interaction of human proteins with artificial materials is of significant importance in different areas such as biomedical implants development, biosensors and nanosafety. For instance when an implant is put into contact with the human tissue, protein adsorption is the first phenomena to occur spontaneously in the series of biological responses. The formed protein layer at the solid-liquid implant interface is a part of the processes which may lead to the acceptance or rejection of the implant. It is known that surface nanotopography of artificial materials independent of surface chemistry affect protein adsorption. Proteins respond to the surface nanotopography when size of the topographical features is comparable to the dimensions of proteins (size of a typical protein is in the nanometer range e.g. human fibrinogen has dimensions of 47× 9× 6 nm).
Recent advancements in nanotechnology enable us to create nanostructured surfaces to mimic the environment which proteins sense in the presence of e.g. implanted artificial materials. In other words fabrication of model surfaces consisting patterns of nanoholes, nanogrooves and in general nanoscale features comparable with dimensions of proteins help us to understand the influence of surface nanotopography on protein behavior. In this project arrays of nanoholes with different diameters down to 45 nm and depth of 50 nm were fabricated on silicon in order to investigate the effect of size of the nanoholes on protein adsorption.
One of the major challenges in this field of study is how to characterize protein behavior in the presence of nanostructured surfaces. In this work we have developed a unique method to study the influence of nanoholes with different sizes on a monolayer of fibrinogen adsorption. Our approach includes fluorescence technique using the intrinsic fluorescence of proteins and atomic force microscopy for further understanding of the interaction.
In this thesis we have fabricated the arrays of nanoholes on Si substrate, deposited a monolayer of fibrinogen molecules on the nanostructured area and used fluorescence technique to investigate the effect of nanoholes on fibrinogen adsorption qualitatively. Atomic force microscopy was used as a complimentary technique to understand the distribution of fibrinogen molecules attached onto the nanostructured area. According to the fluorescence measurements, fibrinogen adsorption is significantly increased as the size of nanoholes approaches 45 nm.

• author: Malekian, Bita
• supervisor: Dan Hessman
• organization: Solid State Physics; Department of Physics
• course: FYSM60 20142
• year: 2015
• type: H2 - Master's Degree (Two Years)
• subject: Physics and Astronomy
• keywords: nanostructured surfaces, fibrinogen, intrinsic fluorescence, proteins, fluorescence imaging, Raman spectroscopy, protein adsorption, atomic force microscopy, nanoholes, SEM, EBL, RIE, fluorescence spectroscopy, Albumin, laminin, IgG, protein binding, tryptophan
• language: English
• id: 7791678
• date added to LUP: 2017-07-04 15:47:02
• date last changed: 2017-07-04 15:47:02