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A semi-cylindrical lens in Kretschmann geometry combined with a flow cell was designed for a commercial rotating compensator ellipsometer to perform internal reflection spectroscopic ellipsometry measurements, while allowing the use of multiple angles of incidence. A thin glass slide covered with a gold film was mounted between the half-cylindrical lens and a small-volume flow cell ensuring an improved sensitivity for protein adsorption experiments. The performance of the system was investigated depending on the angle of incidence, wavelength range and thickness of the gold films for surface plasmon resonance enhanced ellipsometric measurements, and a sensitivity increase was revealed compared to ellipsometric measurements with standard flow cells, depending on the measurement parameters and configuration. The sensitivity increase was demonstrated for fibrinogen adsorption.
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Multiple angle of incidence, spectroscopic, plasmon-enhanced,
internal reflection ellipsometry for the characterization of
solid-liquid interface processes
P. Petrika,b, E. Agocsa,b , B. Kalasa, P. Kozmaa, B. Fodora,c, J. Nadora,b, C. Majora, M.
aInstitute for Technical Physics and Materials Science, Centre for Energy Research, Hungarian
Academy of Sciences, Konkoly Thege Rd. 29-33, 1121 Budapest, Hungary
bDoctoral School of Molecular- and Nanotechnologies, Faculty of Information Technology,
University of Pannonia, Egyetem Str. 10, 8200 Veszpr´em, Hungary
cDoctoral School of Physics, Faculty of Science, University of P´ecs, Ifj´us´ag Str. 6, 7624 P´ecs,
A semi-cylindrical lens in Kretschmann geometry combined with a flow cell was designed for a commercial
rotating compensator ellipsometer to perform internal reflection spectroscopic ellipsometry measurements, while
allowing the use of multiple angles of incidence. A thin glass slide covered with a gold film was mounted between
the half-cylindrical lens and a small-volume flow cell ensuring an improved sensitivity for protein adsorption
experiments. The performance of the system was investigated depending on the angle of incidence, wavelength
range and thickness of the gold films for surface plasmon resonance enhanced ellipsometric measurements, and
a sensitivity increase was revealed compared to ellipsometric measurements with standard flow cells, depending
on the measurement parameters and configuration. The sensitivity increase was demonstrated for fibrinogen
Keywords: Ellipsometry, Bioellipsometry, Internal reflection, Plasmon enhanced ellipsometry, Protein adsorp-
Optical methods are of primary importance for the development of sensors and measurement techniques to char-
acterize solid-liquid interfaces. Techniques utilizing waveguides, surface plasmon resonance (SPR) or ellipsometry
are being developed and improved intensively.1–3 Currently, the sensitivity of ellipsometry is less than waveguide
or SPR sensors by several orders of magnitude. Moreover, it provides spectroscopic information which helps
to build complex optical models and to measure complex structures in a more quantitative way.4Ellipsometry
has mainly been used in a configuration measuring the surface through the liquid using flow cells.4, 5 There
have been further configurations proposed and demonstrated, which measure from the substrate side, mainly
in order to utilize plasmon enhancement.6–11 We used a semi-cylindrical lens with the Kretschmann geometry
to use spectroscopic ellipsometry in a broad wavelength range at different angles of incidence, and studied the
performance of the tool for different gold layer thicknesses. A detailed sensitivity study is under preparation.12
We designed a flow cell covered by a semi-cylinder made of BK7 glass. The diameter of the semi-cylinder is 51
mm. A 0.5 mm thick glass slides covered by plasmonic gold layers were attached index-matched to the cylinder
from the bottom, with the gold-covered side facing down, pressed against the o-ring of the flow cell. The gold
layers were evaporated in a thickness range from 10 nm to 50 nm. Below the gold layer, 2 nm of Cr2O3was
Corresponding author: Peter Petrik,
Optical Methods for Inspection, Characterization, and Imaging of Biomaterials II, edited by Pietro Ferraro,
Simonetta Grilli, Monika Ritsch-Marte, David Stifter, Proc. of SPIE Vol. 9529, 95290W
© 2015 SPIE · CCC code: 0277-786X/15/$18 · doi: 10.1117/12.2184850
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Figure 1. Photograph of the flow cell with the semi-cylinder placed on the mapping stage of the ellipsometer. A schematic
drawing of the beam path in the cylinder is shown on the top of the graph.
evaporated to enhance the adhesion of gold. The volume of the cell is approximately 10 µl. The small cell is
advantageous not only because smaller amount of expensive materials has to be used for the experiment, but
also because the transient fluid mixing effect is shorter when switching from one solution to the other.
Using our construction, the mechanical unit with the flow cell and the cylinder can be placed onto the
mapping stage of our rotating compensator Woollam M2000DI spectroscopic ellipsometer to a reproducible
position (Fig. 1). The advantage of the semi-cylindrical arrangement is that the incidence of light is perpendicular
to the surface of the cylinder at each goniometer position (i.e. for each angle of incidence on the sample).
Therefore, in a well-aligned case, neither of the ellipsometric parameters of Ψ and ∆ (amplitude and phase,
respectively, of the reflection coefficients of light polarized parallel (rp) and perpendicular (rs) to the plane of
incidence) has to be compensated and calibrated for the aberration caused by non-normal incidence.
The M2000DI ellipsometer has a wavelength range from 193 nm to 1690 nm. However, the part of the
spectrum below approximately 350 nm is cut by the BK7 glass. Additional to the capability of a plasmon
enhanced measurement, a major advantage of measuring from the glass substrate is that there is no cut-off in
the infrared side of the spectrum, i.e. the wavelengths up to 1690 nm can be utilized, in contrast to a cut-off
above 1000 nm when measuring through water in a conventional cell.
In case of plasmon resonance, rpgoes to zero. From the ellipsometric point of view, this appears as a vanish-
ing absolute value of the complex reflection coefficient, |rp/rs|(also denoted as tan(Ψ) in the terminology of
ellipsometry). |rp/rs|maps are shown for different wavelengths and angles of incidence in Fig. 2. The sharpest
resonance is observed for the gold thickness of 40 nm, which decreases for thicker layers (see the 50-nm case in
the figure) due to the increased absorption, and broadens for thinner layers (see the 20-nm case) due to radiation
loss. Plasmon dispersion curves are also plotted (as described in the caption) for refractive indices of glass shifted
by 0.0125 – already indicating a large sensitivity to the refractive index values at the interface of the plasmonic
layer. Note that the instrument with the constructed cell can measure down to the angle of incidence of 45.
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60500 Wavelength (nm) 1650
Figure 2. |rp/rs|values measured on the layer structure of BK7/Cr2O3/Au/Air for different Au layer thicknesses shown in
the bottom right corner of each sub-graph (with the BK7 glass and air being the ambient and the substrate, respectively).
The thickness of the Cr2O3layer for adhesion enhancement was 2 nm. The lines correspond to plasmon dispersion curves
calculated using the equation of θ=asin(sqrt{mN2
2]}/N1), where N1and N2are the refractive indices of the
glass and the liquid, respectively, whereas mdenotes the real part of the dielectric function of the gold layer. For the
different curves, N1is shifted by 0.0125.
Figure 3 shows the difference in the Ψ and ∆ ellipsometric angles between the cases of measuring with
and without an adsorbed fibrinogen layer. For thicknesses below 40 nm, the resonances become broader and
the sharp peaks of large differences (most pronounced for the thickness of 40 nm) at the plasmon resonance
conditions get broadened due to radiation loss.8, 10 Note that because of measuring in a large wavelength range
simultaneously, the overall sensitivity, when fitting whole spectra for complex optical models, are not necessarily
higher for resonant gold layer thicknesses. An analysis of wavelength ranges and plasmon layer thicknesses will
be published elsewhere.12
Figure 4 shows the adsorption curve of fibrinogen using both standard and Kretschmann flow cells when
measuring through the liquid at a fixed angle of incidence of 75.5Both adsorptions are made on a gold surface.
The change of the ∆ value is approximately an order of magnitude larger in case of the internal reflection
plasmonic configuration than in a standard flow cell. The thickness of the fibrinogen layer is about 5 nm,
the change of ∆ during the formation of this layer is about 30, whereas the repeatability of both measured
ellipsometric values of Ψ and ∆ are typically in the range of 0.05. As a consequence, the variation in ∆ for a
5-nm fibrinogen layer is almost three orders of magnitude larger than the repeatability of its measurement.
A total internal reflection plasmonic flow cell in the Kretschmann geometry has been demonstrated. The effect
of different plasmonic gold layer thicknesses has been investigated. It was shown that the sharpest resonance is
obtained for layer thicknesses close to 40 nm. However, because of measuring and fitting in a broad wavelength
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d=10nm'11 IO nm
400 600 800 1000 1200 1400 1600 400 600 800 1000 1200 1400 1600
Wavelength (nm)
0 2 4 6 8 10 12 0 510 15 20 25 30 35 40
30 = Fibrinogen
20 = - Kretschmann
a - Standard
15 =
10 =700nm
Time (min) 8
Figure 3. Differences in the Ψ and ∆ ellipsometric angles (in degrees) measured with and without an adsorbed fibrinogen
layer for different gold layer thicknesses (denoted by don the graph).
Figure 4. Ellipsometric angle ∆ in degrees as a function of time measured during fibrinogen adsorption in standard and
Kretschmann configurations.
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range simultaneously, and measuring not only the amplitude but also the phase difference of the TE- and TM-
polarized reflections, the ideal measurement range for a complex, multi-parameter optical model applied in a
broad spectrum is not necessarily only the resonance value of the angle of incidence, wavelength, and gold layer
thickness. A detailed sensitivity study will be published elsewhere.12
Support from the ENIAC E450EDL project is greatly acknowledged.
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... This opens the way to many applications, including in situ infra red ellipsometry for solid liquid interface processes [37]. Our group designed a modified Kretschmann configuration with a semi-cylinder, in which the light can be coupled from angles ranging from 45° to 90° in a broad wavelength range extending from approximately 350 nm to the end of the range (1690 nm) of our ellipsometer [38]. In this article we describe a two-channel version of the new configuration, in which simultaneous in situ measurements can be made in the semi-cylindrical Kretschmann configuration in two differently prepared surfaces in the same liquid cell in the same run. ...
Plasmon-enhanced in situ spectroscopic ellipsometry was realized using the Kretschmann geometry. A 10-μL flow cell was designed for multi-channel measurements using a semi-cylindrical lens. Dual-channel monitoring of the layer formation of different organic structures has been demonstrated on titania nanoparticle thin films supported by gold. Complex modeling capabilities as well as a sensitivity of ~40 pg/mm2 with a time resolution of 1 s was achieved. The surface adsorption was enhanced by the titania nanoparticles due to the larger specific surface and nanoroughness, which is consistent with our previous results on titanate nanotubes.
Full-text available
The influence of substrate materials on protein adsorption was studied by spectroscopic ellipsometry (SE) and atomic force microscopy. For model proteins fibrinogen and flagellar filaments were chosen and their kinetics of adsorption, surface coverage and adsorbed amount on virgin and chemically activated SiO(2) and Ta(2)O(5) thin films were investigated. In case of flagellar filaments the SE data were analyzed with an effective medium model that accounted for the vertical density distribution of the adsorbed protein layer. Adsorption was measured in situ using flow cells with various fluid volume. Compared to commercially available cells, a flow cell with significantly smaller volume was constructed for cost-effective measurements. The development of the flow cell was supported by finite element fluid dynamics calculations.
Integrated planar optical waveguide interferometer biosensors are advantageous combinations of evanescent field sensing and optical phase difference measurement methods. By probing the near surface region of a sensor area with the evanescent field, any change of the refractive index of the probed volume induces a phase shift of the guided mode compared to a reference field typically of a mode propagating through the reference arm of the same waveguide structure. The interfering fields of these modes produce an interference signal detected at the sensor׳s output, whose alteration is proportional to the refractive index change. This signal can be recorded, processed and related to e.g. the concentration of an analyte in the solution of interest. Although this sensing principle is relatively simple, studies about integrated planar optical waveguide interferometer biosensors can mostly be found in the literature covering the past twenty years. During these two decades, several members of this sensor family have been introduced, which have remarkably advantageous properties. These entail label-free and non-destructive detection, outstandingly good sensitivity and detection limit, cost-effective and simple production, ability of multiplexing and miniaturization. Furthermore, these properties lead to low reagent consumption, short analysis time and open prospects for point-of-care applications. The present review collects the most relevant developments of the past twenty years categorizing them into two main groups, such as common- and double path waveguide interferometers. In addition, it tries to maintain the historical order as it is possible and it compares the diverse sensor designs in order to reveal not only the development of this field in time, but to contrast the advantages and disadvantages of the different approaches and sensor families, as well.
Fundamentals of SPR Sensors.- Electromagnetic Theory of Surface Plasmons.- Surface Plasmon Resonance (SPR) Sensors.- Molecular Interactions in SPR Sensors.- Implementations of SPR Biosensors.- SPR Sensor Instrumentation.- The Art of Immobilization for SPR Sensors.- Applications of SPR Biosensors.- Investigating Biomolecular Interactions and Binding Properties Using SPR Biosensors.- SPR Biosensors for Detection of Biological and Chemical Analytes.- SPR Biosensors for Environmental Monitoring.- SPR Biosensors for Food Safety.- SPR Biosensors for Medical Diagnostics.
Ellipsometry used in internal reflection mode exhibits enhanced thin film sensitivity if operated close to surface plasmon resonance conditions. Compared to conventional ellipsometry, the changes in the ellipsometric parameter Δ are several orders of magnitude larger. Here, the origin of this large sensitivity is discussed by analysing thin film approximations of the complex reflectance ratio. It is found that the thickness sensitivity in Δ is proportional to the inverse of the difference between the intrinsic and the radiation-induced damping of the surface plasmons. (© 2008 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim)
In this study, we have reconstructed the statistical 3D structure of hundreds of nanometers thick surface immobilized flagellar filament protein layers in their native environment, in buffer solution. The protein deposition onto the surface activated Ta2O5 film was performed in a flow cell, and the immobilization process was followed by in situ spectroscopic ellipsometry. A multilayer optical model was developed, in that the protein layer was described by five effective medium sublayers. Applying this method, an in-depth analysis of the protein layer formation was performed. Based on the kinetics in the distribution of the surface mass density, the statistical properties of the filamentous film could be determined computationally as a function of the measurement time. It was also demonstrated that the 3D structure of the protein layer can be reconstructed based on the calculated in-depth mass density profile. The computational investigation revealed that the filaments can be classified into two individual groups in approximately equal ratio according to their orientation. In the first group the filaments are close to laying position, whereas in the second group they are in a standing position, resulting in a significantly denser sublayer close to the substrate than at a larger distance.Highlights► Flagellar filaments were immobilized onto Ta2O5 substrates in flow-cell. ► The surface was monitored in situ with spectroscopic ellipsometry. ► New optical model was developed for the characterization of the filamentous layers. ► The depth profile of protein mass density was determined. ► The statistical 3D structure of the filamentous layer was reconstructed.
In this article, spectroscopic ellipsometry studies of plasmon resonances at metal-dielectric interfaces of thin films are reviewed. We show how ellipsometry provides valuable non-invasive amplitude and phase information from which one can determine the effective dielectric functions, and how these relate to the material nanostructure and define exactly the plasmonic characteristics of the system. There are three related plasmons that are observable using spectroscopic ellipsometry; volume plasmon resonances, surface plasmon polaritons and particle plasmon resonances. We demonstrate that the established method of exploiting surface plasmon polaritons for chemical and biological sensing may be enhanced using the ellipsometric phase information and provide a comprehensive theoretical basis for the technique. We show how the particle and volume plasmon resonances in the ellipsometric spectra of nanoparticle films are directly related to size, surface coverage and constituent dielectric functions of the nanoparticles. The regularly observed splitting of the particle plasmon resonance is theoretically described using modified effective medium theories within the framework of ellipsometry. We demonstrate the wealth of information available from real-time in situ spectroscopic ellipsometry measurements of metal film deposition, including the evolution of the plasmon resonances and percolation events. Finally, we discuss how generalized and Mueller matrix ellipsometry hold great potential for characterizing plasmonic metamaterials and sub-wavelength hole arrays. (C) 2011 Elsevier Ltd. All rights reserved.
Total internal reflection ellipsometry (TIRE) in spectroscopic mode in the wavelength range 400–1200 nm is employed in situ at a solid/liquid interface for investigation of protein adsorption on thin semitransparent gold films. In this configuration, the surface plasmon resonance phenomenon gives a large enhancement of the thin film sensitivity. Adsorption of a monolayer of the protein ferritin is monitored kinetically in situ and results in a change in the ellipsometric parameter Δ of more than 90° compared to 3° in similar ellipsometric measurements on gold substrates. This large sensitivity demonstrates a potential for sensor applications. The ferritin layer optical function is modeled with a Cauchy dispersion model resulting in a layer thickness of 9.2 nm, in good agreement with the dimension of the ferritin molecules. A transition layer between the protein film and the gold layer is necessary to introduce in the model to account for interactions between the protein layer and the gold film. The large sensitivity of TIRE for thin layers opens up a pathway to analyze in detail the structure of thin protein layers provided that a further development of the experimental setup and the model for the protein layer is carried out.
Recently developed method of Total Internal Reflection Ellipsometry (TIRE) represents a very successful combination of the spectroscopic ellipsometry instrumentation with the Kretchmann type Surface Plasmon Resonance (SPR) geometry of the experiment. The modelling shows much higher sensitivity of the TIRE method to small changes in optical parameters (thickness and refractive index) of thin films, as compared to both traditional external reflection ellipsometry and SPR. Considering another advantage of performing the measurements in media of different optical density (and even opaque media), TIRE becomes very convenient for different sensing applications in both gaseous and liquid media, as well as for thin film characterisation. This work presents examples of applications of the TIRE method for the study of DNA hybridization and the registration of low molecular weight toxins.