Page 1
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Glass Surfaces and Coatings
for Biotechnology
Carlo G. Pantano
Department of Materials Science and Engineering
Materials Research Institute
Pennsylvania State University
University Park, PA, USA
Biomaterials and Bionanotechnology
Page 2
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 3
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
lecture outline:
Biomaterials and Bionanotechnology
•relevant characteristics and properties
of glass surfaces and coatings>>> introduction
•surface charge on flat glass substrates>>>substrates for cell transfer assays
•silane and hybrid sol/gel coatings>>>DNA and other microarrays
•carbon-doped “oxycarbide” glass>>>blood contact materials
•nanostructured coatings>>>engineered surfaces for biology, biomedicine and biotechnology
discussion and other applications:•surfaces for pharmaceutical packaging
•superhydrophobic/superhydrophilic surfaces
•bio-active glasses and toxicity
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Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Biomaterials and Bionanotechnology
Characteristics and Properties of Glass Surfaces and Coatings
• composition
• chemical functionality
• contact angle/wettability
• surface charge and other surface forces
• porosity/roughness/specific surface
• cleanliness and chemical durability
• uniformity of ALL the above
Page 5
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 6
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Methods of Characterization- surface composition (XPS)
- depth profiling (SIMS)
- surface roughness (AFM)
- organic adsorbates (FTIR/Raman)
- chemical structure (NMR)
- ellipsometry
- surface charge (streaming potential)
- contact angle tensiometry
- adhesion (CFM)
Biomaterials and Bionanotechnology
Page 7
clean silica surface clean multi-component surface
hydroxylated silica surface
Glass Surface Structure Models
functionalized multi-component surface
Page 8
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Computer simulation of glass surfaces: their atomic/nanoscale
heterogeneity, hydroxlyation and organo-functionalization
-6 -4 -2 0 2 4 6
-6
-4
-2
0
2
4
6
X (ang.)
Y (
an
g.)
0.05000
0.1150
0.1800
0.2450
0.3100
0.3750
0.4400
0.5050
0.5700
0.6350
0.7000
Reactivity
Index
-6 -4 -2 0 2 4 6
-6
-4
-2
0
2
4
6
X (ang.)
Y (
an
g.)
0.05000
0.1150
0.1800
0.2450
0.3100
0.3750
0.4400
0.5050
0.5700
0.6350
0.7000
Reactivity
Index
Page 9
yellow – Si red – O purple – Na blue – O green – H
bulk
Surface
10 Å
Water Molecules Adsorbing on a Simulated Sodium Silicate Glass Surface
Molecular Modeling of Water Interactions with Silica and Silicate Surfaces
Elam A. Leed and Carlo G. Pantano
Page 10
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Leaching and surface layer formation:
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Cubic Cell (22 Å)3
800 atoms
Bulk Glass
Structure
Glass
“Surface” Structure
Leached Glass
StructureSimulated Surface
Layer Structure
“relaxed” from 8000 K to 300 K
(in 500 ps)
“relaxed” at 300 K (100 ps)
Removal of above periodic
boundary condition……..
“relaxed” at 300 K (200 ps)
Removal of: aluminum,
calcium, and sodium
Hydroxylation
(charge neut.)
“condensed” at
300 K (200 ps)
(2 OH’s ~1.5 Å)
1
43
2
1
2 3 4Hydroxylated
Leached Glass Structure
Page 12
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
surface roughening by dissolution
Page 13
Normalized dissolution rates vs. pH for sodium-aluminosilicate glasses in the NBO glass series.
0 2 4 6 8 10 12-16
-15
-14
-13
-12
-11
-10
-9
x = 0.0 glass x = 0.6 glass
x = 0.2 glass x =0.8 glass
x = 0.4 glass x = 1.0 (nepheline glass)
No
rmali
zed d
isso
luti
on r
ate
(m
ole
s gla
ss /
cm
2 /
s)
pH
Page 14
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Cell Transfer for Cervical Cancer Diagnosis
Page 15
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
GYN cell transfer layer by SEM
Page 16
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 17
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Electrical double layer at the glass-water interface
Page 18
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Streaming potential determination of surface charge
Page 19
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Fluid Reservoir
Streaming Cell
Electrometer
Adjustable
Stand
Outlet Inlet
Electrodes
Clamp
Spacer
Gasket
Electrode
Streaming
Potential
System for
Flat Glass
Page 20
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 21
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 22
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Page 23
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
(in water)
Page 24
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Zeta potentials determined for the air and tin surfaces of soda-lime-silicate glass slides for 10-3 KCl solutions
containing 100 ppm of AlCl3 at different pH’s.
Page 25
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Surface compositions (by XPS) for the aluminum-hydroxide sol/gel coated slides, and the tin
surface of an uncoated E slide for reference; Coating 5 was rinsed before the heat treatment.
Page 26
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
-80
-60
-40
-20
0
20
40
60
0 2 4 6 8 10 12 14
pH
Ze
ta P
ote
nti
al
Tin Side w/ 100 ppm AlCl3Air Side w/ 100 ppm AlCl3CoatedSoak 10,000 ppm AlCl3
Gold Superfrost Plus
Gold Rite On (APTES)
EMS Poly-L-Lysine
ESCO Polysine
Al-(hydr)oxide (pH=2.0)
Air side
Al-(hydr)oxide (pH=3.5)
Air side
Coated Glass Slides– inorganic and (commercial) organic coatings
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1 2 3 4 5 6a
bc
d
Glass Substrate
OrganoFunctional Coating
Single strands of
Oligonucleotides or DNA
IMMOBILIZED at
known locations
DNA Microarray: a glass-based biological sensor
glass substrates provide: chemical inertness
optical platform
low fluorescence background
flatness and smoothness
low cost!
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DNA Microarrays
Solid support:Glass slidesSilicon
Plastics, etc.
A planar device comprised of an array of DNA single strands immobilized on the surface of an insoluble solid support.
For DNA arrays: Each spot contains 106 to 109 of identical DNA fragments.
Molecules: oligonucleotides, proteins, cells or tissues
SEGMENT OF THE MICROARRAY
SPOT CONTAINING DNA Probes
Immobilized DNA Probe
AGCTC
AGA
T
1x2 cm2
# of spots = 100-500,000(10-250 m)
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DNA Structure: The Fundamentals
Base-pair appr. 3.5 Å
Sugar-Phosphate Backbone
20 Å
6.8 Å
• DNA is a linear polymer made up of a sugar and phosphate backbone with variable side groups of different nitrogenbases. (A, C, G, T)
• DNA may be single or double stranded.
• COMPLEMENTARY BASE PAIRING:Weak H-bonding between the base pairs
G C and T A(HYBRIDIZATION)
T-C-A-G-G-T-T
A-G-T-C-C-A-A
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DNA Microarrays (Gene Chips for sequencing)
Unknown DNA solution
with fluorescent dyes.
Apply to pre-made DNA Microarray
Each spot contains identical DNAprobes of different known sequence.
Laser Confocal Scan
The sequence of the unknown strand is now determined.
Unknown DNA molecules attach to their complementary probes.
AGC
TC
TCG
AG
Array probe
Unknown probei.e.
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Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
3) Array is washed after
hybridization*
Healthy Cancerous
1) Targets are isolated
and labeled
2) Labeled targets
combined with array
4) Hybridized array
is scanned
Use of Microarrays: Gene Expression Experiment
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Use of Microarrays: Drug Discovery
Image Analysis forQuantification
Extract RNA
Untreated cells
Drug-applied cells
Reverse transcript to cDNA
Fluorescent labeling
Apply to array
Scan
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PROTEIN FROM BLOOD
READOUT
Protein Arrays: Diagnostic Analysis
- Squares of antibodies able to bind aspecific protein representing a disease-causing agent.
- Apply blood to the arrayof antibodies proteins from blood attach
- Apply fluor-labeledantibodies recognizableby the attached proteins.forming a antibody “sandwich”
SCAN
- Dot indicating that the patient has anthrax.
Anderson and Valkirs, Scientific American, 2002
Page 34
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Multiple Surface Chemistries Provide Opportunities for
Immobilization of Various Probes
Amino
Epoxy
Aldehyde
Surface Coatings Recommended Probes
NH3
+NH3
+NH3
+
• PCR products
• Long oligos (size > 50 mers)
• Short and long oligos
• PCR products
• Peptides
• Short and long NH2-modified oligos
• NH2-modified PCR products
• Antibodies
and in addition to
DNA arrays/probes:
ELISA’s
Protein arrays
Carbohydrate arrays
Chem-Bio Sensors
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Unmodified DNA strands carry intrinsic (PO4)
3- groups; glass surfaces functionalized with protonated amino groups (NH2) can be used for their initialimmobilization.
electrostatic attraction
functional amine group-NH3
+
-
+
Phosphate-Sugar DNA backbone(carries negative charge)
GLASS GLASS
Immobilization of Unmodified DNA to glass substrates
Page 36
Physical Deposition of Modified and Unmodified DNA
Microspotting (Shalon ad Brown, Stanford, 1995 )
Because of the ease of use and affordability, microspotting has become the most common microarray technology for basic research.
Page 37
Materials Research Institute
Center for Glass Surfaces, Interfaces and CoatingsIMI – NFG Winter School,
January 2008, Kyoto, Japan
Physical Chemistry/Engineering of the Microspotting Process