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Layer-by-layer coating of degradable microgels for pulsed drug delivery.

Recently, we reported on "self-rupturing" microcapsules which consist of a biodegradable dextran-based microgel surrounded by a polyelectrolyte membrane. Degradation of the microgel increases the swelling pressure in the microcapsules which, when sufficiently high, ruptures the surrounding polyelectrolyte membrane. The membrane surrounding the microgels is deposited using the layer-by-layer (LbL) technique, which is based on the alternate adsorption of oppositely charged polyelectrolytes onto a charged substrate. In this paper, we characterize with confocal microscopy, electrophoretic mobility, scanning electron microscopy and atomic force microscopy in detail the deposition and the properties of the LbL coatings on the dextran microgels. We show that by fine-tuning the properties of both the microgel core and the LbL membrane the swelling pressure which is evoked by the degradation of the microgel is indeed able to rupture the surrounding LbL membrane. Further, we show that the application of an LbL coating on the surface of the microgels dramatically lowers the burst release from the microcapsules and results in massive release at the time the microcapsules rupture.

Delayed-Action Preparations↗

Interactions between MC3T3-E1 cells and textured Ti6Al4V surfaces.

This paper presents the results of an experimental study of the interactions between MC3T3-E1 (mouse calvarian) cells and textured Ti6Al4V surfaces, including surfaces produced by laser microgrooving; blasting with alumina particles; and polishing. The multiscale interactions between MC3T3-E1 cells and these textured surfaces are studied using a combination of optical scanning transmission electron microscopy and atomic force microscopy. The potential cytotoxic effects of microchemistry on cell-surface interactions also are considered in studies of cell spreading and orientation over 9-day periods. These studies show that cells on microgrooved Ti6Al4V geometries that are 8 or 12 microm deep undergo contact guidance and limited cell spreading. Similar contact guidance is observed on the surfaces of diamond-polished surfaces on which nanoscale grooves are formed due to the scratching that occurs during polishing. In contrast, random cell orientations are observed on alumina-blasted Ti6Al4V surfaces. The possible effects of surface topography are discussed for scar-tissue formation and improved cell-surface integration.

Alloys↗

Human stefin B readily forms amyloid fibrils in vitro.

Human stefin B (cystatin B) is an intracellular cysteine proteinase inhibitor broadly distributed in different tissues. Here, we show that recombinant human stefin B readily forms amyloid fibrils in vitro. It dimerises and further oligomerises, starting from the native-like acid intermediate, I(N), populated at pH 5. On standing at room temperature it produces regular (over 4 microm long) fibrils over a period of several months. These have been visualised by transmission electron microscopy and atomic force microscopy. Their cross-sectional diameter is about 14 nm and blocks of 27 nm repeat longitudinally. The fibrils are smooth, of unbranched surface, consistent with findings of other amyloid fibrils. Thioflavin T fluorescence spectra as a function of time were recorded and Congo red dye binding to the fibrils was demonstrated. Adding 10% (v/v) trifluoroethanol resulted in an increased rate of fibrillation with a typical lag phase. The finding that human stefin B, in contrast to the homologue stefin A, forms amyloid fibrils rather easily should promote further studies of the protein's behaviour in vivo, and/or as a model system for fibrillogenesis.

Amyloid beta-Peptides↗

Silica-coated CdTe quantum dots functionalized with thiols for bioconjugation to IgG proteins.

Quantum dots (QDs) have been increasingly used in biolabeling recently as their advantages over molecular fluorophores have become clear. For bioapplications QDs must be water-soluble and buffer stable, making their synthesis challenging and time-consuming. A simple aqueous synthesis of silica-capped, highly fluorescent CdTe quantum dots has been developed. CdTe QDs are advantageous as the emission can be tuned to the near-infrared where tissue absorption is at a minimum, while the silica shell can prevent the leakage of toxic Cd(2+) and provide a surface for easy conjugation to biomolecules such as proteins. The presence of a silica shell of 2-5 nm in thickness has been confirmed by transmission electron microscopy and atomic force microscopy measurements. Photoluminescence studies show that the silica shell results in greatly increased photostability in Tris-borate-ethylenediaminetetraacetate and phosphate-buffered saline buffers. To further improve their biocompatibility, the silica-capped QDs have been functionalized with poly(ethylene glycol) and thiol-terminated biolinkers. Through the use of these linkers, antibody proteins were successfully conjugated as confirmed by agarose gel electrophoresis. Streptavidin-maleimide and biotinylated polystyrene microbeads confirmed the bioactivity and conjugation specificity of the thiolated QDs. These functionalized, silica-capped QDs are ideal labels, easily synthesized, robust, safe, and readily conjugated to biomolecules while maintaining bioactivity. They are potentially useful for a number of applications in biolabeling and imaging.

Biotinylation↗

Morphology and structure of photosensitive dye J-aggregates adsorbed on AgBr microcrystals grown in gelatin.

Though the cyanine dye J-aggregates carry the role to sense the exposing light in the silver halide photographic system, little research on the morphology of the aggregates in adsorption has been made with modern surface analytical methods. In this paper, we describe the size, epitaxy, multi-layered array formation, nucleation and preferential adsorption, and irregular distribution of population between particles and the segregation on a particle, of J-aggregates adsorbed on AgBr grown in gelatin. We employed cathodoluminescence microscopy, low energy high resolution scanning electron microscopy, and atomic force microscopy. Dye molecules aggregate together near the surface of AgBr and adsorb on the surface. The growth of adsorbed aggregates is controlled by the diffusion of dye molecules from the surrounding solution. The population of J-aggregates adsorbed on an AgBr particle varies from almost none to full coverage. Each aggregate is about (20-30) x (30-50) nm in size and is 2.1 nm thick for thiacarbocyanine with sodium ion, 1.04 nm for thiacarbocyanine with tosyl ion, and 0.5 nm for an oxacarbocyanine. The aggregates connect their longer edges to each other to form arrays, and the arrays build up multi-layered stacks. The arrays align parallel and segregate to form terraces. The longer edges of J-aggregates align along [210] on AgBr (100) or [632] on AgBr (111).

Bromides↗

Targeting of biotinylated compounds to its target tissue using a low-density lipoprotein receptor-avidin fusion protein.

The very high binding affinity of avidin to biotin is one of the highest to occur in nature. We constructed a fusion protein composed of avidin and the endocytotic LDL receptor in order to target biotinylated molecules to cells of the desired tissues. In addition to the native avidin, charge-mutated and nonglycosylated avidins were utilized as part of the fusion proteins, in order to modify its properties. All of the fusion protein versions retained the biotin-binding capacity. Although the specificity was not increased, however, fusion proteins composed of natural avidin and nonglycosylated avidin bound most efficiently to the biotinylated ligands. Fluorescence microscopy and atomic force microscopy studies revealed the expression of the fusion protein on cell membranes, and demonstrated specific and high-affinity binding of biotin to the low-density lipoprotein receptor (LDLR)-avidin fusion protein in vitro. Additionally, systemically administered biotinylated ligand targeted with high specificity the intracerebral tumors of rats that were expressing fusion protein after the virus-mediated gene transfer. These results suggest that local gene transfer of the fusion protein to target tissues may offer a novel tool for the delivery of biotinylated molecules in vitro and in vivo for therapeutic and imaging purposes.

Animals↗

[Experimental research on the effect of nanophase ceramics on osteoblasts functions].

In order to study the cytocompatibility of nanophase hydroxyapatite ceramic in vitro, we prepared hydroxyapatite by use of the wet chemistry techniques. The grain size of hydroxyapatite of interest to the present study was determined by scanning electron microscopy and atomic force microscopy with image analysis software. Primary culture of osteoblast from rat calvaria was established. Protein content, synthesis of alkaline phosphatase and deposition of calcium-containing mineral by osteoblasts cultured on nanophase hydroxyapatite ceramics and on conventional hydroxyapatite ceramics for 7, 14, 21 and 28 days were examined. The results showed that the average surface grain size of the nanophase and that of the conventional HA compact formulations was 55 (nanophase) and 780 (conventional) nm, respectively. More importantly, compared to the synthesis of alkaline phosphatase and deposition of calcium-containing mineral by osteoblasts cultured on nanophase was significantly greater than that on conventional ceramics after 21 and 28 days. The cytocompatibility was significantly greater on nanophase HA than on conventional formulations of the same ceramic.

Alkaline Phosphatase↗

Structural studies of soluble oligomers of the Alzheimer beta-amyloid peptide.

Recent studies have suggested that non-fibrillar soluble forms of Abeta peptides possess neurotoxic properties and may therefore play a role in the molecular pathogenesis of Alzheimer's disease. We have identified solution conditions under which two types of soluble oligomers of Abeta40 could be trapped and stabilized for an extended period of time. The first type of oligomers comprises a mixture of dimers/tetramers which are stable at neutral pH and low micromolar concentration, for a period of at least four weeks. The second type of oligomer comprises a narrow distribution of particles that are spherical when examined by electron microscopy and atomic force microscopy. The number average molecular mass of this distribution of particles is 0.94 MDa, and they are are stable at pH 3 for at least four weeks. Circular dichroism studies indicate that the dimers/tetramers possess irregular secondary structure that is not alpha-helix or beta-structure, while the 0.94 MDa particles contain beta-structure. Fluorescence resonance energy transfer experiments indicate that Abeta40 moieties in amyloid fibrils or protofibrils are more similar in structure to those in the 0.94 MDa particles than those in the dimers/tetramers. These findings indicate that soluble oligomeric forms of Abeta peptides can be trapped for extended periods of time, enabling their study by high resolution techniques that would not otherwise be possible.

Alzheimer Disease↗

Effects of reuse and bleach/formaldehyde reprocessing on polysulfone and polyamide hemodialyzers.

The surface features, morphology, and blood interactions of fibers from pristine, bleach/formaldehyde reprocessed, and reused Fresenius Polysulfone High Flux (Hemoflow F80B) hemodialyzers and Gambro Polyflux 21S Polyamide hemodialyzers have been studied. SEM images of fibers from both hemodialyzer types revealed a dense skin layer on the inner surface and a relatively thick porous layer on the outer surface. The 21S polyamide support layer consisted of interconnected highly porous structures. Environmental scanning electron microscopy and atomic force microscopy images of both membrane types showed alterations in morphology due to reprocessing and reuse; however the changes were more marked for the 21S polyamide dialyzers. Fluorescence microscopy images showed only minimal fluorescence associated with the fibers after patient use and reprocessing, suggesting that blood derived deposits were removed by processing. The protein layers formed on pristine and reused hemodialyzer membranes during clinical use were studied using SDS-PAGE and immunoblotting. Before bleach/formaldehyde treatment, protein layers of considerable amount and complexity were found on the blood side of singly and multiply used dialyzers. Proteins adsorbed on the dialysate side were predominantly in the molecular mass region below 30 kDa. However, some higher molecular mass proteins were detected on the dialysate side of the 21 S polyamide dialyzers. Very little protein was detected on dialyzers that were treated with bleach/formaldehyde after dialysis, regardless of whether they had been used/reprocessed once or 12 times.

Electrophoresis, Polyacrylamide Gel↗

Polyelectrolyte multilayer films as substrates for photoreceptor cells.

Reconstruction of extracellular matrix substrates for delivery of functional photoreceptors is crucial in pathologies such as retinal degeneration and age-related macular degeneration. In this study, we assembled polyelectrolyte films using the layer-by-layer deposition method. The buildup of three different films composed of poly(L-lysine)/chondroitin sulfate (PLL/CSA), poly(L-lysine)/poly(styrenesulfonate) (PLL/PSS), or poly(L-lysine)/hyaluronic acid (PLL/HA) was followed by means of quartz crystal microbalance measurements, optical waveguide light mode spectroscopy, confocal microscopy, and atomic force microscopy. The exponential growth regime and the diffusion of PLL chains from the bulk through the PLL/CSA, PLL/PSS, and PLL/HA films was examined. Evaluation of photoreceptor cell viability was optimal on one layer of PLL (PLL(1)), followed by 10 bilayers of PLL/HA [(PLL/HA)(10)] and 10 bilayers of PLL/CSA [(PLL/CSA)(10)]. The number of bilayers and the type of terminating layer also had a significant influence on the number of photoreceptor cells attached. Functionalized polyelectrolyte multilayer films were obtained by adsorbing basic fibroblastic factor (bFGF) or the insoluble fraction of interphotoreceptor matrix (IPM) on or within polyelectrolyte multilayers. bFGF and IPM adsorption on top of the (PLL/CSA)(10)/PLL polyelectrolyte films increased the number of photoreceptor cells attached and maintained the differentiation of rod and cone cells.

Animals↗

Characterization and surface reactivity of ferrihydrite nanoparticles assembled in ferritin.

Ferrihydrite nanoparticles with nominal sizes of 3 and 6 nm were assembled within ferritin, an iron storage protein. The crystallinity and structure of the nanoparticles (after removal of the protein shell) were evaluated by high-resolution transmission electron microscopy (HRTEM), atomic force microscopy (AFM), and scanning tunneling microscopy (STM). HRTEM showed that amorphous and crystalline nanoparticles were copresent, and the degree of crystallinity improved with increasing size of the particles. The dominant phase of the crystalline nanoparticles was ferrihydrite. Morphology and electronic structure of the nanoparticles were characterized by AFM and STM. Scanning tunneling spectroscopy (STS) measurements suggested that the band gap associated with the 6 nm particles was larger than the band gap associated with the 3 nm particles. Interaction of SO2(g) with the nanoparticles was investigated by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, and results were interpreted with the aid of molecular orbital/density functional theory (MO/DFT) frequency calculations. Reaction of SO2(g) with the nanoparticles resulted primarily in SO(3)2- surface species. The concentration of SO3(2-) appeared to be dependent on the ferrihydrite particle size (or differences in structural properties).

Ferric Compounds↗

Effects of heat/citric acid reprocessing on high-flux polysulfone dialyzers.

The surface features, morphology, and tensile properties of fibers obtained from pristine, reprocessed, and reused Fresenius Polysulfone High-Flux (Hemoflow F80A) hemodialyzers have been studied. Scanning electron microscopy of the dialyzer fibers revealed a dense skin layer on the inner surface of the membrane and a relatively thick porous layer on the outer surface. Transmission electron microscopy and atomic force microscopy showed an alteration in membrane morphology due to reprocessing and reuse, or to a deposition of blood-borne material on the membrane that is not removed with reprocessing. Fluorescent microscopy images also showed that a fluorescent material not removed by heat/citric acid reprocessing builds up with continued use of the dialyzers. The tensile properties of the dialyzer fibers were not affected by the heat/citric acid reprocessing procedure. The protein layers formed on pristine and reused hemodialyzer membranes during clinical use were also studied using sodium dodecyl sulfate polyacrylamide gel electrophoresis and immunoblotting. A considerable amount of protein was found on the blood side of single and multiple use dialyzers. Proteins adsorbed on the dialysate side of the membrane were predominantly in the molecular weight region below 30 kDa. Little protein was detected on the membranes of reprocessed hemodialyzers.

Anticoagulants↗

Structural analysis of wheat wax (Triticum aestivum, c.v. 'Naturastar' L.): from the molecular level to three dimensional crystals.

In order to elucidate the self assembly process of plant epicuticular waxes, and the molecular arrangement within the crystals, re-crystallisation of wax platelets was studied on biological and non-biological surfaces. Wax platelets were extracted from the leaf blades of wheat (Triticum aestivum L., c.v. 'Naturastar', Poaceae). Waxes were analysed by gas chromatography (GC) and mass spectrometry (MS). Octacosan-1-ol was found to be the most abundant chemical component of the wax mixture (66 m%) and also the determining compound for the shape of the wax platelets. The electron diffraction pattern showed that both the wax mixture and pure octacosan-1-ol are crystalline. The re-crystallisation of the natural wax mixture and the pure octacosan-1-ol were studied by scanning tunnelling microscopy (STM), atomic force microscopy (AFM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Crystallisation of wheat waxes and pure octacosano-1-ol on the non polar highly ordered pyrolytic graphite (HOPG) led to the formation of platelet structures similar to those found on the plant surface. In contrast, irregular wax morphologies and flat lying plates were formed on glass, silicon, salt crystals (NaCl) and mica surfaces. Movement of wheat wax through isolated Convallaria majalis cuticles led to typical wax platelets of wheat, arranged in the complex patterns typical for C. majalis. STM of pure octacosan-1-ol monolayers on HOPG showed that the arrangement of the molecules strictly followed the hexagonal structure of the substrate crystal. Re-crystallisation of wheat waxes on non-polar crystalline HOPG substrate showed that technical surfaces could be used to generate microstructured, biomimetic surfaces. AFM and SEM studies proved that a template effect of the substrate determined the orientation of the re-grown crystals. These effects of the structure and polarity of the substrate on the morphology of the epicuticular waxes are relevant for understanding interactions between biological as well as technical surfaces and waxes.

Crystallization↗

Crystallization behavior and thermal properties of melt-crystallized poly[(R)-3-hydroxybutyric acid-co-6-hydroxyhexanoic acid] films.

Melt-crystallized films of poly[(R)-3-hydroxybutyric acid-co-10mol% 6-hydroxyhexanoic acid] (P[(R)-3HB-co-6HH]) were prepared by isothermal crystallization at various temperatures for 3 days, and subsequently stored at room temperature after the films formed well-developed and volume-filled spherulites. The lamellar morphologies and properties of melt-crystallized films were characterized by means of wide-angle X-ray diffraction, small-angle X-ray scattering, differential scanning calorimetry, transmission electron microscopy, and atomic force microscopy. The melting endotherm of P[(R)-3HB-co-6HH] films was composed of a broad peak starting around room temperature and of a sharper peak starting above the isothermal crystallization temperature. The stacking of flat-on lamellae with lamellar periodicity of 8-10 nm was detected on the surface of P[(R)-3HB-co6HH] films after the primary crystallization at 110 degrees C. On storage at room temperature above the Tg (-5 degrees C) of copolyester, thin crystals of 1-4 nm thickness appeared on the surface of P[(R)-3HB-co-6HH] films crystallized at 110 degrees C. These results suggest that long sequences of (R)-3HB units in a random copolyester form relatively thick P[(R)-3HB] crystalline lamellae during the primary crystallization process at a given crystallization temperature, while shorter sequences of (R)-3HB units, which are incapable of crystallizing at a given crystallization temperature, form relatively thin crystalline lamellae during the subsequent crystallization process at room temperature.

Calorimetry, Differential Scanning↗

P-glycoprotein inserted in planar lipid bilayers formed by liposomes opened on amorphous carbon and Langmuir-Blodgett monolayer.

The insertion of proteins into planar lipid layers is of outstanding interest as the resulting films are suitable for the investigation of protein structure and aggregation in a lipid environment and/or the development of biotechnological applications as biosensors. In this study, purified P-glycoprotein (P-gp), a membrane drug pump, was incorporated in model membranes deposited on solid supports according to the method by Puu and Gustafson, Biochim. Biophys. Acta 1327 (1997) 149-161. The models were formed by a double lipid layer obtained by opening P-gp-containing liposomes onto two hydrophobic supports: amorphous carbon films and Langmuir-Blodgett (L-B) lipid monolayers, which were then observed by transmission electron microscopy and atomic force microscopy, respectively. Before the opening of liposomes, the P-gp structure and functionality were verified by circular dichroism spectroscopy and enzymatic assay. Our micrographs showed that liposomes containing P-gp fuse to the substrates more easily than plain liposomes, which keep their rounded shape. This suggests that the protein plays an essential role in the fusion of liposomes. To localize P-gp, the immunogold labeling of two externally exposed protein epitopes was carried out. Both imaging techniques confirmed that P-gp was successfully incorporated in the model membranes and that the two epitopes preserved the reactivity with specific mAbs, after sample preparation. Model membranes obtained on L-B monolayer incorporated few molecules with respect to those incorporated in the model membrane deposited onto amorphous carbon, probably because of the different mechanism of proteoliposome opening. Finally, all particles appeared as isolated units, suggesting that P-gp molecules were present as monomers.

1,2-Dipalmitoylphosphatidylcholine↗

Crystal orientation mapping of NiO grown on cube textured Ni tapes.

Samples of cube textured Ni tapes were oxidized in flowing oxygen at different temperatures. Crystal orientation maps (COMs) of the resulting oxide layers were produced by electron backscatter diffraction. The oxide layers were also analysed by X-ray diffraction (XRD), scanning electron microscopy and atomic force microscopy (AFM). The oxide grain size of a sample oxidized at 600 degrees C was similar to that of the substrate and the oxide was highly textured, both indicating epitaxial growth. The orientation relationship between the substrate and the oxide was directly observed from XRD to be (111)NiO//(001)Ni, [101]NiO//[110]Ni with four, equivalent, in-plane variants. In each variant, the oxide has both <110>- and <211>-type directions parallel to the Ni <110> directions. Differences in oxide thickness and surface roughness on neighbouring grains were revealed by AFM and these were attributed to the existence of a range of oxide growth conditions resulting from small differences in the orientation of each substrate grain. Similar macrotexture and microstructure were observed on a sample oxidized at 1300 degrees C, but additional, facetted oxide crystals had formed at the oxide grain boundaries. COMs showed that these crystals were either cube or 45 degrees rotated cube orientated, a texture different to that of the large oxide grains. The grain boundary crystals were thought to form by inward diffusion of oxygen at defects in the growing oxide scale.

Crystallography↗

Ligand substitution of receptor targeted DNA complexes affects gene transfer into hepatoma cells.

We have targeted the serpin enzyme complex receptor for gene transfer in human hepatoma cell lines using peptides < 30 amino acids in length which contain the five amino acid recognition sequence for this receptor, coupled to poly K of average chain length 100 K, using the heterobifunctional coupling reagent sulfo-LC SPDP. The number of sulfo-LC SPDP modified poly-L-lysine residues, as well as the degree of peptide substitution was assessed by nuclear magnetic resonance spectroscopy. Conjugates were prepared in which 3.5%, 7.8% or 26% of the lysine residues contained the sulfo-LC SPDP moiety. Each of these conjugates was then coupled with ligand peptides so that one in 370, one in 1039, or one in 5882 lysines were substituted with receptor ligand. Electron microscopy and atomic force microscopy were used to assess complex structure and size. HuH7 human hepatoma cells were transfected with complexes of these conjugates with the plasmid pGL3 and luciferase expression measured 2 to 16 days after treatment. All the protein conjugates in which 26% of the K residues were modified with sulfo-LC SPDP were poor gene transfer reagents. Complexes containing less substituted poly K, averaged 17 +/- 0.5 nm in diameter and gave peak transgene expression of 3-4 x 10(6) ILU/mg which persisted (> 7 x 10(5) ILU) at 16 days. Of these, more substituted polymers condensed DNA into complexes averaging 20 +/- 0.7 nm in diameter and gave five-fold less luciferase than complexes containing less substituted conjugates. As few as eight to 11 ligands per complex are optimal for DNA delivery via the SEC receptor. The extent of substitution of receptor-mediated gene transfer complexes affects the size of the complexes, as well as the intensity and duration of transgene expression. These observations may permit tailoring of complex construction for the usage required.

Animals↗

Cholesterol rules: direct observation of the coexistence of two fluid phases in native pulmonary surfactant membranes at physiological temperatures.

Pulmonary surfactant, the lipid-protein material that stabilizes the respiratory surface of the lungs, contains approximately equimolar amounts of saturated and unsaturated phospholipid species and significant proportions of cholesterol. Such lipid composition suggests that the membranes taking part in the surfactant structures could be organized heterogeneously in the form of inplane domains, originating from particular distributions of specific proteins and lipids. Here we report novel results concerning the lateral organization of bilayer membranes made of native pulmonary surfactant where the coexistence of two distinct micrometer sized fluid phases (fluid ordered and fluid disordered-like phases) is observed at physiological temperatures by using fluorescence microscopy and atomic force microscopy. Additional experiments using fluorescent-labeled proteins SP-B and SP-C show that at physiological temperatures these hydrophobic proteins are located exclusively in the fluid disordered-like phase. Most interestingly, the microscopic coexistence of fluid phases is maintained up to 37.5 degrees C, where most fluid ordered phases melt. This observation suggests that the particular composition of this material is naturally designed to be at the "edge" of a lateral structure transition under physiological conditions, likely providing particular structural and dynamic properties for its mechanical function. The observed lateral structure in native pulmonary surfactant membranes is dramatically affected by the extraction of cholesterol, an effect not observed upon extraction of the surfactant proteins. Furthermore, the spreading properties of the native surfactant material at the air-liquid interface were also greatly affected by cholesterol extraction, suggesting a connection between the observed lateral structure and a physiologically relevant function of the material. We suggest that the particular lipid composition of surfactant could be finely tuned to provide, under physiological conditions, a structural scaffold for surfactant proteins to act at appropriate local densities and lipid composition.

Calorimetry, Differential Scanning↗