Phospholipid Langmuir-Blodgett Films Research Articles

Layer by Layer Optimization of Langmuir–Blodgett Films for Surface Acoustic Wave (SAW) Based Sensors for Volatile Organic Compounds (VOC) Detection

Rayleigh surface acoustic wave (RSAW)-based resonant sensors, functionalized with single and multiple monomolecular layers of Langmuir–Blodgett (LB) films, were thickness and density optimized for the detection of volatile organic compounds (VOC), which could impose a serious threat on the environment and human health. Single layers of a phospholipid (SLP), hexane dissolved arachidic acid (HDAA), and chloroform dissolved arachidic acid (CDAA) were used for the LB film preparation. Several layers of these compounds were deposited on top of each other onto the active surface of high-Q 434 MHz two-port RSAW resonators in a LB trough to prepare a highly sensitive vapor detection quartz surface microbalance (QSM). Frequency shift was measured with a vector network analyzer (VNA). These devices were probed with saturated vapors of hexane, chloroform, methanol, acetone, ethanol, and water after each deposited layer to test the behavior of the QSM’s insertion loss, loaded Q, vapor sensitivity, and to find the optimum trade-off between these parameters for the best real-life sensor performance. With 2200 ppm and 3700 ppm sensitivity to chloroform, HDAA and CDAA coated QSM devices reached the optimum sensor performance at 15 and 11–15 monolayers, respectively. Surface pressure optimized single monolayers of phospholipid LB films were found to provide up to 530 ppm sensitivity to chloroform vapors with a negligible reduction in loss and loaded Q. This vapor sensitivity is higher than the mass of the sensing layer itself, making SLP films an excellent choice for QSM functionalization.

Reversible Monolayer-Bilayer Transition in Supported Phospholipid LB Films under the Presence of Water: Morphological and Nanomechanical Behavior.

Mixed monolayer Langmuir-Blodgett (LB) films of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) and cholesterol (Chol) in the 1:1 ratio have been prepared onto solid mica substrates. Upon immersion in water or in an aqueous HEPES solution (pH 7.4) the monolayer LB films were spontaneously converted into well-organized bilayers leaving free mica areas. The process has been demonstrated to be reversible upon removal of the aqueous solution, resulting in remarkably free of defects monolayers that are homogeneously distributed onto the mica. In addition, the nanomechanical properties exhibited by the as-formed bilayers have been determined by means of AFM breakthrough force studies. The bilayers formed by immersion of the monolayer in an aqueous media exhibit nanomechanical properties and stability under compression analogous to those of DPPC:Chol supported bilayers obtained by other methods previously described in the literature. Consequently, the hydration of a monolayer LB film has been revealed as an easy method to produce well-ordered bilayers that mimic the cell membrane and that could be used as model cell membranes.

Nanolayers for early diagnostics of proteins involved in degenerative amyloidosis

FK-506 binding protein (FKBP12) is a protein of the family of immunophilins, involved in many neurodegenerative diseases such as Alzheimer's syndrome where FKBP12 is known to be over-expressed in early stages of the disease. We designed and built Langmuir-Blodgett nanostructures incorporating ligands with high affinity for FKBP12: Tacrolimus (FK506) and Rifaximin as candidate nanosensors to detect low FKBP12 concentration in the initial phase of the amyloidosis. The binding process of the different ligands has been studied by means of photophysical measurements investigating the fluorescence quenching of the tryptophan residue in the binding pocket of FKBP12 by addition of the ligand in solution. Immobilization of the ligands was achieved adopting biomimetic strategy: phospholipid Langmuir-Blodgett films are proposed as nanoscaffolds for ligand inclusion. Several phospholipid nanoarchitectures differing in lipid composition, fluidity, number of layers and method of production (incubation versus co-spreading) were screened. The results have shown that both FK506 and Rifaximin ligands penetrate the lipid matrix either as monomers or as aggregates depending on their initial concentration. More importantly, the experiments demonstrated that the ligands in the LB scaffolds efficiently quench FKBP12 fluorescence in solution as a consequence of ligand binding to the protein.

Self-Patterned Mixed Phospholipid Monolayers for the Spatially Selective Deposition of Metals

Metal-reactive organosulfur groups were patterned onto mica and silicon surfaces by dewetting instabilities during the Langmuir-Blodgett (LB) deposition of phase-separated mixed phospholipid monolayers. Monolayers were formed from binary mixtures of dipalmitoylphosphatidylcholine (DPPC), dilauroylphosphatidylcholine (DLPC), and their ω-methyldisulfide-dialkylphosphatidylcholine analogues, DSDPPC and DSDLPC. Patterns of highly parallel stripes of condensed DPPC or DSDPPC, protruding by 0.7-0.9 nm from a fluid matrix of DLPC or DSDLPC, were observed over areas extending at least 30 × 30 μm(2) in the LB films. The average stripe width varied from ca. 150 to 500 nm, depending on the lipid composition and deposition pressure. X-ray photoelectron spectroscopy confirmed that the phospholipid-monolayer-bound methydisulfides react with Au vapor to form a gold-thiolate species. The adsorption of thermally evaporated Au, Ag, and Cu onto DSDPPC/DLPC and DPPC/DSDLPC patterns was investigated by field emission gun scanning electron microscopy (FEGSEM) and atomic force microscopy (AFM). A change in phase contrast is observed in FEGSEM and AFM over the methyldisulfide-functionalized areas following metal deposition due to metal-thiolate bond formation. An increase in step height between the DSDPPC stripes and nonfunctionalized DLPC background following metal deposition, as well as the resistance of the metal-coated DSDPPC or DSDLPC regions to detergent extraction from the surface, attest to a selective metallization of the pattern. Our results indicate that the preferential adsorption of vapor-deposited metal onto the ω-methyldisulfide-terminated phase occurs at submonolayer coverages. The chemical reactivity exhibited by the organosulfur-modified phospholipid LB films make these templates potentially interesting for the fabrication of solid-supported patterns of metal nanostructures.

Calcium Carbonate Particle Growth Depending on Coupling among Adjacent Layers in Hybrid LB/LbL Films

There are practical and academic situations that justify the study of calcium carbonate crystallization and especially of systems that are associated with organic matrices and a confined medium. Despite the fact that many different matrices have been studied, the use of well-behaved, thin organic films may provide new knowledge about this system. In this work, we have studied the growth of calcium carbonate particles on well-defined organic matrices that were formed by layer-by-layer (LbL) polyelectrolyte films deposited on phospholipid Langmuir-Blodgett films (LB). We were able to change the surface electrical charge density of the LB films by changing the proportions of a negatively charged lipid, the sodium salt of dimyristoyl-sn-glycero-phosphatidyl acid (DMPA), and a zwitterionic lipid, dimyristoyl-sn-glycero-phosphatidylethanolamine (DMPE). This affects the subsequent polyelectrolyte LbL film deposition, which also changes the the nature of the bonding (electrostatic interaction or hydrogen bonding). This approach allowed for the formation of calcium carbonate particles of different final shapes, roughnesses, and sizes. The masses of deposited lipids, polyelectrolytes, and calcium cabonate were quantified by the quartz crystal microbalance technique. The structures of obtained particles were analyzed by scanning electron microscopy.

Influence of the glycosylphosphatidylinositol anchor in the morphology and roughness of Langmuir–Blodgett films of phospholipids containing alkaline phosphatases

The morphology of phospholipid Langmuir–Blodgett films containing a glycosylphosphatidylinositol-anchored rat osseous plate alkaline phosphatase was studied by atomic force microscopy. Two forms of the enzyme were used in this study: an anchor-containing detergent-solubilized form, and a phospholipase-solubilized form, without the hydrophobic moiety. Direct measurements of catalytic activity in enzyme/phospholipid mixed films, and nanogravimetric estimations by quartz crystal microbalance confirmed the adsorption of both forms of alkaline phosphatase. Atomic force microscopy analysis showed that both enzyme forms originated aggregates when adsorbed on phospholipid Langmuir–Blodgett films. However, the phospholipase-solubilized form showed a higher roughness when compared to the detergent solubilized alkaline phosphatase. A model suggesting different alignments of the major axis of the ellipsoid polypeptide moiety relative to the film surface is proposed. This model, supported by surface density and catalytic activity data, might explain the difference of roughness observed for films containing each enzyme form adsorbed onto a phospholipid-modified solid support.

Surface density as a significant parameter for the enzymatic activity of two forms of alkaline phosphatase immobilized on phospholipid Langmuir–Blodgett films

Rat osseous plate alkaline phosphatase, a glycosylphosphatidylinositol (GPI)-anchored phosphomonohydrolase, was immobilized on Langmuir–Blodgett (LB) films. Enzyme solubilization either with polyoxyethylene-9-lauryl ether or with a glycosylphosphatidylinositol-specific phospholipase C resulted in a GPI-anchor-containing and a GPI-anchor-depleted form, respectively. Both forms were adsorbed on dimyristoylphosphatidic acid LB films and restricted to the outermost layer. The surface density and enzyme activity were determined using a quartz crystal microbalance and p-nitrophenylphosphatase activity, respectively. The detergent-solubilized form was co-spread with dimyristoylphosphatidic acid on the air/water interface and transferred to solid supports, providing an enzyme maximum surface density of 530 ng/cm 2. Maximal phosphohydrolytic activity, corresponding to 43% of that observed in homogeneous medium, was obtained at a surface density of 179 ng/cm 2. The phospholipase C-solubilized form was adsorbed directly from solution, reaching a maximum surface density of 1541 ng/cm 2, although the phosphomonohydrolase activity was 10 times lower than that obtained for the anchor-containing form. The combined analysis of surface density and enzymatic activity suggests that the alignment of the protein molecules on the LB lipid films induced by the glycosylphosphatidylinositol anchor facilitates the access of the substrate to the active site. This access is hampered by increasing enzyme surface densities and depends on a specific orientation of the adsorbed enzyme.

Spectroscopic investigation of tetracycline interaction with phospholipid Langmuir–Blodgett films

A widely used veterinary antibiotic, tetracycline (TC), has been incorporated in Langmuir–Blodgett (LB) films of dipalmitoylphosphatidic acid (DPPA) by means of two different procedures: co-transfer and incubation in solution. The resulting structures were characterized by means of contact angle and ellipsometric measurements. The presence of the antibiotic in the phospholipid film was evidenced by means of UV–Vis electronic absorption and infrared vibrational spectroscopy. The two sets of measurements unambiguously indicated the presence of the drug in the LB layer films obtained with both methods, although incubation led to a smaller content of immobilized tetracycline. In both cases, the drug was found to reside in the hydrophilic portion of the layers due to specific interactions of the dimethylamino group of the molecule with the polar head groups of the phospholipid.

Total External Reflection X-ray Absorption Spectroscopy Reveals a Zinc Coordination Shell in Phospholipid Langmuir−Blodgett Films

We have studied the local environment of zinc in phospholipid Langmuir-Blodgett films with extended X-ray absorption fine structure spectroscopy in total reflection conditions (reflEXAFS). Mono- and multilayer films made of dilauroyl-phosphatidic acid (DLPA) and monolayer films made of dimyristoyl-phosphatidylcholine (DMPC) were deposited on quartz slides from ZnCl2 solutions, and their molecular order was checked by means of X-ray reflectivity measurements. A very good signal-to-noise ratio in the reflEXAFS spectra of both multi- and monomolecular films was obtained. The theoretical fits to the experimental data show that zinc coordinates four atoms about 0.20 nm distant in both acidic DLPA and neutral DMPC films, while in water solutions, zinc coordinates six atoms about 0.21 nm distant.

Imaging of various surface properties of fluorescently labelled phospholipid Langmuir-Blodgett films with a combined scanning probe microscope.

We present results of phase separation of a single-component system of 1,2-dihexadecanoyl-sn-glycero-3-phospho-[N-(4-nitrobenz)-2-oxa-1,3-diazolyl]ethanolamine in which a liquid-condensed (LC) phase co-exists with a liquid-expanded (LE) phase. Domain formation in the co-existence region was studied using a newly developed combined scanning near-field optical microscope-atomic force microscope (SNOM-AFM). We demonstrate for the first time that the topographic, friction, fluorescence and surface potential distributions for a phase-separated single-component Langmuir-Blodgett film between the LE and LC phases can be simultaneously observed using the SNOM-AFM with a thin-step etched optical fibre probe.

Impedance Measurement of Taste Responses of Biomimetic LB Films

Impedance changes in phospholipid LB (Langmuir-Blodgett) and protein-mixed phospholipid LB films by several taste substances were measured. The resistance of the phospholipid LB film increased by bitter and sweet substances, whereas the resistance decreased for salty, sour and umami substances. For sweetness the larger change was observed in the protein-mixed phospholipid LB film, and hence discrimination between sweetness and bitterness was possible.

Molecular images of phospholipid Langmuir-Blodgett films by atomic force microscopy

Abstract Langmuir-Blodgett films of dimyristoylphosphatidylcholine (DMPC) have been studied on both macroscopic and microscopic scales by atomic force microscopy in the LE-LC coexisting phase (liquid expanded and liquid condensed phase). Macroscopic scale images show that Langmuir-Blodgett films of DMPC have many similar domain structures as observed in fatty acids Langmuir-Blodgett films. In addition, it is observed that some phospholipid molecules crystalize out. A molecularly resolved image was only obtained in LC phase while in the LE phase, no molecularly resolved image was obtained because the films were destroyed by the tip interaction.

Characterization of mixed miscible and nonmiscible phospholipid Langmuir–Blodgett films by atomic force microscopy

Langmuir–Blodgett (LB) phospholipid films have proved to be efficient models for biological membranes. In order to acquire a better knowledge of the structure of these membranes, LB films of a mixture of miscible and nonmiscible phospholipids were studied by atomic force microscopy (AFM). Mixtures of various compositions were deposited on mica and graphite substrates at various pressures. AFM images showed two coexisting domains corresponding to the nonmiscible (DPPE-DOPE) phospholipids with difference in height of about 0.5 nm due to the structural differences between the two phospholipids. The area ratios between the two types of domains, measured on AFM images, were consistent with the molar fraction and area per molecule at the surface pressure during deposition. We have also reported on the topography of multilayers of nonmiscible mixtures. In conclusion, AFM allowed us to have a better insight into deposition mechanisms and the properties of binary mixtures of phospholipids LB films.

Electronic Control of Diffusion Through Langmuir-Blodgett Film Membranes

A preliminary investigation is reported into the diffusion of a quinine derivative through composite membranes obtained by the deposition of phospholipid Langmuir-Blodgett films on to porous nylon supports. The results suggest that the thin organic layers probably collapse into the pores in the membrane rather than bridge them. It is found that the diffusion of the drug can be influenced by the application of an electric field.

Analytical surface spectroscopy of phospholipid Langmuir-Blodgett films

Langmuir-Blodgett (LB) films composed of DPPC and DNP-cap-DPPE phospholipids were characterized by X-ray photoelectron spectroscopy (XPS) and static secondary ion mass spectrometry (SSIMS). These surface analytical techniques were systematically evaluated with respect to capabilities for the determination of film composition, orientation and thickness. Mixed monolayers were utilized to investigate quantitative compositional analysis capabilities. Orientation information was obtained using angle dependent XPS and by utilizing variations in the extent of Ag cationization of molecular ions in SSIMS of DPPC monolayers on Ag substrates. Film thickness determinations were provided by XPS studies of monolayer and multilayer DPPC films.

Observation of phospholipase A2activity towards the hydrolysis of phospholipid Langmuir–Blodgett films deposited on a quartz-crystal microbalance

Catalytic hydrolysis of phospholipid Langmuir–Blodgett (LB) films deposited on a quartz-crystal microbalance (QCM) by phospholipase A2 was followed by observing frequency changes of the QCM in an aqueous solution.

Stability and characterization of phospholipid Langmuir-Blodgett films

The regularity of the bilayer structure was compared between dipalmitoylphosphatidic acid (DPPA) Langmuir-Blodgett (LB) films and “oriented multilayers” of DPPA, using X-ray diffraction and electron spectroscopy for chemical analysis (ESCA). The angular variation of the (001) Bragg spot due to the irregularity of the lamellar repeat was remarkable in the system of oriented multilayers. In contrast, the spot of DPPA LB films exhibited only a slight angular variation, suggesting a regular structure of this film. The take-off angle dependence of the phosphorus:carbon ratio obtained by ESCA indicates that the regular layered structure in DPPA LB film is maintained after 49 layers are deposited.

Inhomogeneous translational diffusion of monoclonal antibodies on phospholipid Langmuir-Blodgett films

The translational mobility of fluorescent-labeled monoclonal antibodies specifically bound to supported phospholipid bilayers containing hapten-conjugated phospholipids has been measured as a function of the surface concentration of bound antibodies using fluorescence recovery after photobleaching. Fluorescence recovery curves are fit well by a model that assumes the presence of two populations of antibodies with different lateral diffusion coefficients. The larger diffusion coefficient equals 3.5 x 10(-9) cm2/s, the smaller diffusion coefficient ranges from 1.5 x 10(-9) cm2/s to 2.5 x 10(-10) cm2/s, and the fractional fluorescence recovery associated with the smaller coefficient increases from approximately 0 to approximately 0.7 with increasing concentration of bound antibody. These results suggest that complexes of haptenated phospholipids and antibodies in phospholipid Langmuir-Blodgett films form clusters or domains in a concentration-dependent fashion.

Scanning tunneling microscopy of lipid films and embedded biomolecules

We have observed the ordered molecular structure of fatty acid and phospholipid Langmuir-Blodgett films with a scanning tunneling microscope. Besides confirming earlier studies of cadmium arachidate bilayers on graphite, we have observed bilayers of di-myristoyl-phosphatic acid (DMPA) on cleaved graphite, and studied monolayers of DMPA on oxidized graphite and on gold-coated calcite in air. We present the first images of a bilayer containing the protein concanavalin A. These experiments confirm the possibility of obtaining STM images by electron transport through layers of organic molecules more than 50 Å thick. Sample preparation, microscopic technique, and results are discussed.

Steady state conduction in synthetic phospholipid Langmuir-Blodgett films

The results of an investigation into d.c. conduction in Langmuir-Blodgett films of dipalmitoyl phosphatidylethanolamine are reported. They show that, after long times under electrical stress in a vacuum, a final stable value of conductivity is achieved which depends superlinearly on applied voltage and very weakly on temperature. Comparison of the data with Chow's equation for electron tunnelling yields a value of about 2.7 nm for the tunnelling distance. Before the final steady condition is achieved, the current shows an additional component which has an activation energy of 0.36 eV. This component only contributes to d.c. conduction in positive bias conditions and is removed after 10 6 s under electrical stress. At short times, the current in both polarities is dominated by the a.c. conductance of the sample.