3.051J/20.340J 1 Lecture 12: Biomaterials Characterization in Aqueous Environments High vacuum techniques are important tools for characterizing surface composition, but do not yield information on surface structure or chemistry in a water-based environment. Aqueous-based methods for surface characterization are limited. Here we will consider three common techniques: 1. water contact angle studies - surface reconstruction (a) - water absorption (b) - surface chemistry analysis 112 2cos cos cosffθ θθ=+ θadroplet volume advancing receding θθr droplet volume advancing receding θ(a)(b)Cassie’s eqn: use to determine fraction of surface area of components 1 & 2 (f1 + f2 = 1)3.051J/20.340J 2 2. in situ ellipsometry - degree of hydration of a film Ellipsometric angles Ψ and ∆ ⇒ thickness (df ) & refractive index (nf) ( 3-layer model) f water water material materialnfn f n=+ nwater nf df nsub where fwater and fmaterial are volume fractions.3.051J/20.340J 3 3. Atomic Force Microscopy (or Surface Force Microscopy): imaging method that exploits intermolecular interactions between a small (~atomic) probe and molecules on surface position sensitive photodetector He-Ne laser (Au-coated) Si, Si3N4, SiO2 cantilever; spring const k~0.1-1N/m “atomic” Si, Si3N4, C tip sample surfacepiezoelectric transducer piezo-controlled sample stage Intermolecular potential curve short-range: ion-ion repulsion Long-range (attractive): van der Waals, H-bonding, electrostatic, dipole-dipole,… rU(r) Scan area: 1×1 nm2 to 250×250 µm2 z-range: 8 µm force range: 10-13-10-6N Force generated: UFz−∆=∆3.051J/20.340J 4 Operation Modes 1. Contact mode (short-range) ¾ Tiny cantilever deflections detected by photodiode array ¾ Tip rastered over sample surface at fixed force (via photodetector-z-piezo feedback loop) generates topographical image ⇒ analogous to stylus on a record player ¾ Good for hard samples; can drag soft materials! force applied: nN x-y resolution: 1Å z resolution: < 1Å Contact mode images of TiO2 (rutile) film surface ¾ No contrast at low resolution—flat surface ¾ High resolution—atoms of (001) plane are revealed Figure 10 (a) and (b) from K.D. Jandt, Surf. Sci. 491 (2001) 303.Photo removed for copyright reasons.3.051J/20.340J 5 2. “Tapping” mode ¾ Tip oscillates in z-axis at high ω (~50-500 kHz in air, 10 kHz in fluids) with intermittent sample contact ⇒ eliminates shear forces ¾ Interactions between tip and sample cause amplitude attenuation (driven amplitude ~ 10 nm) ¾ Cantilever deflections used in feedback loop to maintain average applied force similar to contact mode oscillatory amplitude attenuation ⇒ “height” data ¾ Commonly used for soft samples, aqueous environments x-y resolution: 1-2 nm Figures 1 and 4 from C.H. Chen, D.O. Clegg & H.G. Hansma, Biochemistry 37, 1998, 8262. Tapping mode images in air (left) and water (below) of laminin (Ln-1) adsorbed onto mica. ¾ Cruciform molecular shape ¾ “Arms” can bend and fold Photos removed for copyright reasons.3.051J/20.340J 6 Phase imaging (in conjunction with tapping mode) ¾ Tip oscillated in z-axis, making intermittent sample contact ¾ Simultaneous measurement of amplitude attenuation & phase lag of cantilever signal vs. signal sent by piezo-driver oscillation amplitude attenuation ⇒ “height” data oscillation phase-shift ⇒ “elasticity” map hard soft hard Drive signal: Phase data: in phase out of phase in phase Figure 6 from M.J. Fasolka et al., Macromolecules 33, 2000, 5702. Figure removed for copyright reasons.AFM image of polystyrene-b-poly(lauryl methacrylate) block copolymer film. Height data: variation in film thickness seen at polymer droplet edge Phase data: microdomains of soft PLMA block (Tg~-35C) and hard PS block (Tg~100C) are distinguished3.051J/20.340J 7 3. Force modulation mode ¾ Tip oscillates in z-axis at ω < ωο = (k/m)1/2 (cantilever resonance frequency), making intermittent sample contact; ω ~3-120kHz. ¾ Interactions between tip and sample cause amplitude attenuation ¾ Contact force applied to sample is modulated, giving elasticity information cantilever deflection amplitude ⇒ “elasticity” map 4. Non-contact AFM ¾ Oscillation near resonance frequency without tip-surface contact (long-range forces in U(r) curve; r > 0.6 nm, typical F <1 pN) ¾ Force gradients from surface interactions shift resonance frequency 12odFdz kωω−−= ¾ Force gradients used to map secondary interactions (difficult in fluids due to damping; good for soft samples) hard soft hard Drive signal: Force modulation: dF/dz >0 ⇒ attractive force dF/dz <0 ⇒ repulsive force resolution: dF/dz ~ 10 µN/m (0.1 pN at a gap of 1 nm)3.051J/20.340J 8 5. Force-Distance Profiles ¾ As sample is brought towards probe tip, measured force: ∆F = k∆zc (∆zc = cantilever deflection) D > 10 nm hydrophobic interactions, electrostatic interactions, steric repulsion of polymer “brush” layer D < 10 nm van der Waals attraction ¾ Obtain F(z) of species w/ surface by coating tip with receptor, antibody, ligand, colloid, cell, etc. Colloidal Particle Force Spectroscopy Grafted hydrophilic chains(EO)22 ⇒ “cloaking” pure repulsion Mixed grafted chains (EO)22 ⇒ “cloaking” C16H37 ⇒ “binding” repulsion-attraction-repulsion - jump to contact seen for ()UDkDD−> (i) - further approach bends cantilever (ii) - on retraction, tip “sticks” from adhesion forces (iii) NOTE: ↑ tip size = loss of x-y imaging resolution after S.C. Olugebefola et al., Langmuir 18, 2002, 1098. Figure by MIT OCW.3.051J/20.340J 9 ¾ Measure height of hydrated surface layer via nonlinear regimes Sample height interval: ∆zs = zs,j−zs,j-1 Force increment from cantilever deflection: ∆F ≡ k∆zc Sample deformation: ∆zs - ∆zc F (nN) Hydrated layer thickness ∆F/∆zs (nN/nm) 0 - zs −zo (instant slope) ∆zs ∆zc No tip/surface interactions; ∆F ≡ k∆zc = 0 Full contact with hard substrate; ∆F ≡ k∆zc = k∆zs Deformation of hydrated surface; ∆F ≡ k∆zc ≠ k∆zs zs −zo = separation distance 03.051J/20.340J 10 Biomaterials-relevant SFM/AFM Studies