Transcript Slide 1

Characterization of biological thin films with
combinatorial spectroscopic ellipsometry and
piezoelectric nanogravimetry
J.
1,2
Gerasimov * K.B.
2,5
Rodenhausen ,
T.
3,4
Kasputis ,
2,6
Schmidt ,
D.
A.K. Pannier3, R. Lai1,2, and M. Schubert2,6
H.
2,6
Wang ,
1Department
of Chemistry; 2Center for Nanohybrid Functional Materials; 3Department of Biological Systems Engineering; 4Biomedical Engineering Program;
5Department of Chemical and Biomolecular Engineering; 6Department of Electrical Engineering; University of Nebraska-Lincoln, U.S.A.
*[email protected]
http://ellipsometry.unl.edu
Experimental Design
Abstract
The formation and function of biotechnologically relevant
functional thin films was studied in real-time using tandem
spectroscopic ellipsometry (SE) and quartz crystal
microbalance (QCM). Simultaneously monitoring changes in
film thickness given by each technique allows us to derive the
porosity of the bioactive layer under investigation and gives
clues about the morphology of the surface-confined
biomolecules. A correlation of structural morphology with film
function makes possible the directed optimization of bioactive
films based on measurable parameters. We present the
results of two investigations involving the formation of
bioactive hybrid materials. In the study of DNA-incorporating
monolayers for sequence-specific DNA detection, two DNA
probes were analyzed for their ability to specifically recognize
their complement in solution. The formation of protein layers
on smooth and nanostructured surfaces was studied for its
utility as a scaffold for whole-cell immobilization and future
work in gene transfer studies.
Instrument
Theory
•
light
source
liquid
inlet
SE thickness (dSE) gives the thickness of adsorbed organic layer, excluding solvent.
liquid
outlet
detector
optical
window
optical
window
polarizer
𝑑𝑆𝐸
•
QCM thickness is derived from total adsorbed mass
𝜌𝑜 , 𝑚𝑜 = 𝑑𝑒𝑛𝑠𝑖𝑡𝑦 𝑎𝑛𝑑 𝑚𝑎𝑠𝑠 𝑜𝑓 𝑡ℎ𝑒 𝑎𝑑𝑠𝑜𝑟𝑏𝑎𝑡𝑒
1 𝜌𝑜
𝜌𝑎
𝑑𝑄𝐶𝑀 = (
+
) 𝜌𝑎 , 𝑚𝑎 = 𝑑𝑒𝑛𝑠𝑖𝑡𝑦 𝑎𝑛𝑑 𝑚𝑎𝑠𝑠 𝑜𝑓 𝑡ℎ𝑒
𝐴 𝑚𝑜 𝑚𝑎
𝑖𝑛𝑐𝑜𝑟𝑝𝑜𝑟𝑎𝑡𝑒𝑑 𝑠𝑜𝑙𝑣𝑒𝑛𝑡
•
Assuming equivalence of solvent and adsorbate densities, porosity (𝑓𝑜 𝑉) may be
expressed as a ratio of the thickness measured in SE to the thickness measured in
QCM
analyzers
polarized
light
gasket
polarized
light
surface
affected
by sample
QCM-D
control
𝑓𝑜 𝑉
𝑑𝑆𝐸
=
𝑑𝑄𝐶𝑀
Sequence-specific DNA Detection
Electrochemical DNA Sensor
Target Detection
Probe Adsorption
1.0
4
3
0.6
noncomplement
buffer
complement
target
DNA
buffer
Flush Probe
ddSE
SE
ddQCM
QCM
0.4
1
0
0
-0.5
100
Fibronectin
200
300
-0.5
1500
Time (min)
0.0
100
200
0.8 300
0.3
400
buffer
noncomplementary
target
complementary
-0.1 target
0
100
200
300
0.2
3
0.6
2
0.4
1
100
200
300
Time (min)
0.1
0.0
0.5
0.4
-0.1
0.3
0.2
-0.2
-0.3
0
400
aligned
0.1
0.0
-0.1
100
200
300
400
500
Time (min)
0.8
0.8
12
0.6
10
0.6
v
0.4
1
0.2
0.6
8
0
2
ddSE
SE
dQCM
dQCM
f0v
1000
0.1
14
0.8
f0
3
500
v
Protein Fraction
(%)
f
QCM
0.4
0.4
6
4
0.2
0.2
2
0
100
200
300
400
0
100
200
300
400
0
0
Time
(min)
100
200
300
400
(min)
80
20
100
160 40 18060Time200
220 120
Study surface functionalization
for cell adhesion and gene
transfer
Aptamer-based small molecule
detectionº
14
XQCM
Protein Fraction
12
Protein Fraction (%)
XQCM (mg/m )
2
Substrate
1.0
Nanostructured Surface
Titanium Nanocolumns on Gold
0
In the near future, we will add an electrochemistry module to the SE/QCM to study
the relationship between electrochemical trends and film morphology. We will direct
further efforts toward the following goals:
Time (min)
20
18
16
14
12
10
8
6
4
2
0
140
400
Future Work
Protein Fraction
Thickness (nm)
Thickness (nm)
2
XQCM (mg/m )
• extracellular matrix protein
• facilitates cell adhesion,
spreading, and proliferation
• diverse applications in
biotechnology and engineering
of hybrid materials
4X
0.3
0.2
0.1
-0.1
0
ddSE
SE
ddQCM
QCM
0.4
0.0
0
0
Flat Surface
12
20
4
18
10
16
3
8
14
0.5
12
26 0.4
10
4
80.3
1 0.2
6
2 0.1
4
0
200.0
0
0-0.1
0 0
140
0.5
0.0
-0.2
0.2
-0.2
-0.3
-0.4
Potential (vs. Ag/AgCl)
0.2
Time (min)
4
-0.1
0.4
0.2
Biofunctionalizing Sculptured Thin Films
Structured
Thin Film
0.2
v
18
16
14
12
10
8
6
4
2
0
-2
-0.2
-0.3
-0.4
Potential (vs. Ag/AgCl)
Thickness (nm)
0.4
0.5
𝑓f𝑜0 𝑉
Decrease in eT efficiency is
monitored using alternating
current voltammetry
SE/QCM provides structural
information about DNA layer
formation
bound
-0.1
SSSS S SSSS
•
2
Thickness (nm)
0
Elevated
Probe
Electron transfer (eT) to the
labeled, surface-bound stem-loop
DNA probe is greatly impeded
upon hybridization with
complementary target.
0.6
0.3
Thickness (nm)
•
1
SSSS S SSSS
SSSS S SSSS
Current (nA)
•
unbound
3
v
𝑓
f0𝑜 𝑉
eT
SSSS S SSSS
2
Current (nA)
eT
Thickness (nm)
0.8
buffer
10
8
6
4
2
160
180
200
Time (min)
220
Nebraska EPSCoR RII External Reviewer Panel Visit, University of Nebraska-Lincoln, August 2011
0
SSSS S SSSS
SSSS S SSSS
Characterize peptide-based
sensors for antibody detection
Monitor conformational change in
surface-confined proteinsºº
º Helmholtz Association of German Research Centres (2009, April 7). Fitting Pieces For Biosensors. ScienceDaily. Retrieved August
7, 2011, from http://www.sciencedaily.com /releases/2009/04/090407105149.htm
ººadapted from http://webs.wichita.edu/mschneegurt/biol103/lecture16/normal_rogue.gif
Acknowledgements: The Procter & Gamble Co., J.A. Woollam Co., Inc., NSF EPS-1004094, NSF CAREER CHE-0955439
400