Instructor: Dr. Marinella Sandros

1. Nanochemistry NAN 601 Instructor: Dr. MarinellaSandros Lecture 20: LBL Assembly

2. Layer by Layer (LBL) Assembly • It was first reported by Iler in 1966 that multilayer thin film can be fabricated by alternative deposition of oppositely charged nanoparticles. • In addition to charged colloids, they also pointed out the potentials of using other charged species such as polyvalent ions, surfactants, water soluble polymer and even proteins to build the multilayer assembly.

3. LBL Assembly Awareness • Iler’s suggestive work did not get public attention until later. Decher and Hong rediscovered and established the work in this area. • In 1997, a feature article named “Fuzzy Nanoassemblies: Toward Layered Polymeric Multicomposites”, published in Science by Decher, systematically reviewed the work of LBL assembly.

4. Basis of LBL Assembly • LBL assembly can be built through conventional methods such as: • electrostatic interactions • hydrogen bonding • step by step reaction • molecular recognition and biorecognition

5. LBL Assembly Technique • The preparation of LBL is relatively easy without sophisticated procedures or instruments. Beakers containing dipping solutions and waters are all we need. • Substrates with cleaned hydrophilic surfaces such as glass, silica or mica, which exhibit a nonzero surface charge, must be employed.

6. Procedure • The deposition of a first layer is achieved by dipping the charged substrate into a solution of a polyion of opposite charge. • After a typical 20 min immersion, the substrate is washed in water to remove excess polyions. • During this process, the adsorption of a polyion layer leads to overcompensation of surface charge.

7. Procedure • Therefore, the sign of net charge on the surface is inversed, and the subsequent deposition of an oppositely charged polyion becomes possible. • In most cases, drying is performed after layer deposition. By repeating the process, dipping alternately in Beaker A and B with washing and drying in between, a multilayer assembly with the desired number of layers can be built up.

8. Layer Constituents

9. Advantages • There are many advantages using LBL assembly over other strategies for ultrathin film preparation such as the Langmuir-Blodgett (LB) technique and the self-assembled monolayer (SAM) method.

10. Langmuir-Blodgett (LB) The principle of Langmuir-Blodgett deposition. Amphiphile molecules are spread over the aqueous solution (A). The area of the surface is reduced by the movement of the barriers. This compresses the amphiphiles to form an ordered film (B). The amphiphile film can be transferred to a solid surface if the latter is slowly drawn through the solution/air interface (C). http://www.foi.se/surfbiotech/tt/img00007.gif

11. LB Method • LB method suffers from the requirement of expensive instruments and is not applicable with many kinds of non-amphililic materials. https://www.abo.fi/student/en/media/11284/lbbild2.jpg

12. Self-Assembled Monolayer (SAM) • Self-assembled monolayers, SAMs, are formed when surfactant molecules spontaneously adsorb in a monomolecular layer on surfaces. Two of the most widely studied systems of SAMs are gold -alkylthiolate monolayers and alkylsilane monolayers. http://upload.wikimedia.org/wikipedia/commons/2/28/SAM_schematic.jpeg

13. Self-Assembled Monolayer (SAM) • SAM can be built up on a wider range of materials; however, it can not form multilayers. http://www.oocities.org/thomas_r_bonner/sam/sam_afm.jpg

14. Advantages of LBL • (1) LBL is extremely cheap, no expensive instrument required. • (2) The fabrication process is simple, and can be achieved either manually or by machine. • (3) The film can form not only on planar substrates, but also on substrates with different shapes. The thickness increment per layer is self-regulating; similar surface roughness of LBL is expected regardless of the roughness of substrates. • (4) LBL can incorporate different functional groups into the system for different applications.

15. Advantages of LBL • (5) Applied materials can be used such as conventional polyelectrolytes as well as various functional polyelectrolytes. • (6) Water-soluble biomaterials with charged sites on their surfaces can also be incorporated, such as proteins, DNA, and charged viruses. • (7) Charged inorganic substances: Colloidal nanoparticles, clay, zeolite and polyoxometalates have been reported for use in LBL assembly. • (8) Charged supramolecular assemblies have also been incorporated into LBL assembly

16. Nanoporous Polyelectrolyte Films • A multilayer thin film is formed by adsorbing a blend of SiO2 NPs and poly(acrylic acid) (PAA) in alternation with poly(allyamine hydrochloride) (PAH) on a PEI modified substrate. • The film is then cross-linked, forming amide linkages between PAA and PAH, followed by the removal of SiO2 NPs with HF/NH4F to make the porous structure. • BSA adsorbed increased with increasing bilayer number.

17. Formation of polyelectrolytecapsules on an “air” core • The Tween:Span mixture, which is used to form air microbubbles (a), is further stabilized by the electrostatic assembly of PAH/PSS multilayers (b–d). Bottom: a photograph of aircontaining polyelectrolyte capsules in aqueous solution after centrifugation. • Potential use as contrast agents in ultrasonic diagnostics, gas dispersingcontainers and in chemical catalysis poly(allylamine hydrochloride) (PAH) and poly(styrene sulfonate) (PSS) layers Angew. Chem. Int. Ed. 2005, 44, 3310 –3314

18. Nano-encapsulation of furosemidemicrocrystals for controlled drug release • Illustration of furosemide microcrystal encapsulation and release study. In step 1, precursor layers of (PSS/PDDA)2 are assembled onto positively charged furosemide microcrystals. In step 2, (PSS/gelatin)2–6 layers are added. In step 3, drug release in aqueous solution is monitored at different pH values.

19. Biocompatible LBL films fortissue engineering • Cells are sensitive both to the micro/nano-topographic and chemical features of their surrounding environment. • The engineering of surface properties of biomaterials is critical to develop bioactive devices with which to elicit appropriate cellular responses. ---An example: Development of biocompatible films for bone tissue engineering

20. Human Osteoblast-Like Cells Response to NanofunctionalizedSurfaces for Tissue Engineering • Biomimetic coatings containing fibronectin (FN), an adhesive glycoprotein of the extracellular matrix, have been assembled by means of the LbL technique. • Three precursor bilayers of cationic poly(dimethyldiallyl ammonium chloride) (PDDA) and anionic poly (styrenesulfonate) (PSS) were deposited to impart an homogeneous and well-defined charge to the substrates. • Subsequently cationic poly-L-lysine (PLL) was employed in alternation with FN. J Biomed Mater Res Part B: Appl Biomater 84B: 249–255, 2008

21. Bone Tissue Engineering • To evaluate the biocompatibility of the developed coatings MG63 human osteoblast-like cells were used. • Cells were seeded onto functionalised and non functionalized silicon and Nickel/Titanium (NiTi) surfaces. • Cell dispersion and morphology was evaluated on the surface of treated and non treated glass coverslips by phase contrast microscopy.

22. Results I

23. Results II

24. Results II • The developed coatings resulted to be effective in improving cell response both on silicon and NiTi. • These experimental evidences demonstrates the high potential of the assembled ultrathin films for the optimization of the surface properties of bone implants.

25. LBL films for BiosensorsDevelopment • Multilayer containing functional proteins can be assembled directly onto the surface of a transducer for the setting up of a biosensor MOVIE http://www.chem.fsu.edu/multilayers/Multilayer%20Animation%20Fast.swf

26. Kinetics of Polyion Adsorption • For the time-dependent control of adsorption and monitoring of the assembly in situ, the quartz crystal microbalance method (QCM). • The kinetics of the adsorption process could be delineated by the QCM technique, which is indispensable for establishing proper assembly conditions • The method is based on a vibrating quartz crystal sensor (an acoustic resonator). Measurements are made based on changes in vibration frequency in response to reactions that occur on the sensor surface.

27. Quartz Crystal Microbalance(QCM) • A quartz crystal microbalance (QCM) measures a mass per unit area by measuring the change in frequency of a quartz crystal resonator. The resonance is disturbed by the addition or removal of a small mass due to film deposition at the surface of the acoustic resonator.

29. QCM-D • In liquid, an adsorbed film may consist of a considerably high amount of water, which is sensed as a mass uptake by all QCMs. • By using QCM-D and measuring several frequencies and the dissipation it becomes possible to determine whether the adsorbed film is rigid or water-rich (soft), which is not possible by looking only at the frequency response where Elost is the energy lost (dissipated) during one oscillation cycle and Estored is the total energy stored in the oscillator. http://www.q-sense.com/qcm-d-technology

31. QCM-D • 1. Binding of a small globular molecule Moderate frequency response, Δf (mass change), but low dissipation, ΔD(structural change). • 2. Binding of a large elongated moleculeForms a softer and thicker layer which can be seen by higher Δf and much higher ΔD levels. • 3. Rinsing with bufferThe elongated molecule is removed, frequency and dissipation reduce again. http://www.q-sense.com/applications

32. Biointerfacing Properties http://www.q-sense.com/applications

33. Application of QCM-D For all applications, Q-Sense systems enable: • Mass measurements. • Nanogram sensitivity, less than 1% of a protein monolayer is detectable. • Structural properties. Simultaneously determine the orientation of molecules at the surface. • Real time, in situ and label free. Study kinetics, pH dependence etc. using the native molecule. • Flexible choice of substrate. Metals, polymers; any material that can be applied as a thin film can be used.

34. QCM-D Movie • http://www.q-sense.com/download

35. Piezoelectric Immunosensor for Taxane Monitoring Journal of Immunological MethodsVolume 313, Issues 1-2, 30 June 2006, Pages 191-198

36. Taxane Monitoring by QCM

37. LBL Method Review • Typically a glass or silicon wafer surface that has negative charge is coated first with a layer of a cationic polyelectrolyte such as poly(ethylene imine) (PEI, which has + charge due to partial protonation) or poly(diallyldimethyl- ammonium chloride) by dipping the substrate into an aqueous solution of the polymer for a few minutes, withdrawing the substrate and washing off the excess polymer. • The dipping operation is repeated with an aqueous solution of an anionic polyelectrolyte such as poly(acrylic acid) (which has negative charge due to partial deprotonation) or poly(sodium 4- styrenesulfonate) (PSS).

38. LBL Method Review • Thus alternating layers of polymers of opposite charge are deposited, such as PEI/PSS/PEI/PSS. • The top layer of polymer always leaves an excess of one sign of charge that attracts the next layer of opposite charge. • Tens or hundreds of layers can be deposited much faster than by the LB method. • The oppositely charged polyelectrolytes become highly entangled, and the films can have exceptionally high mechanical strength. • The film structures are completely amorphous.

39. LBL Method Review • In modifications of the LBL method one of the polyelectrolytes can be replaced by colloidal particles or even carbon nanotubes of the right surface charge, enabling the preparation for example of films that fluoresce different colors, if CdTe particles of different size are embedded.