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The original version of this Article omitted the author Kuan Wang, who is from the 'College of Biomedical Engineering, Taipei Medical University, Taipei 11031, Taiwan' and 'Nanyang Environment & Water Research Institute, Nanyang Technological University, Singapore 637141, Singapore.'Also, the author S.H. Lim was incorrectly given as L.S. Hoi and A. Larsson was incorrectly given as A. Larson.The "Author contributions" was amended to reflect the authorship changes. It previously read 'Y.Z.S., C.-W.Q., and A.Q.L. jointly conceived the idea. Y.Z.S., S.X., Y.Z., J.B.Z., W.S., J.H.W., T.N.C., Z.C.Y., Y.L.H., B.L., P.H.Y., D.P.T., and C.-W.Q. performed the numerical simulations and theoretical analysis. Y.Z.S., S.X., and L.K.C. did the fabrication and experiments of particle hopping, biomolecule binding and flow cytometry. A.L. and L.S.H. did the SPR experiments. S.X., Y.Z.S., Y.Z., C.-W.Q., Y.-Y.C., L.K.C., T.H.Z., and A.Q.L. prepared the manuscript. S.X., Y.Z., C.-W.Q., and A.Q.L. supervised and coordinated all the work. All authors commented on the manuscript.' The correct version states 'B.L., K. W., P.H.Y.' instead of 'B.L., P.H.Y.' and 'S.H.L.' in place of 'L.S.H.'This has been corrected in both the PDF and HTML versions of the Article.
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Particle trapping and binding in optical potential wells provide a versatile platform for various biomedical applications. However, implementation systems to study multi-particle contact interactions in an optical lattice remain rare. By configuring an optofluidic lattice, we demonstrate the precise control of particle interactions and functions such as controlling aggregation and multi-hopping. The mean residence time of a single particle is found considerably reduced from 7 s, as predicted by Kramer's theory, to 0.6 s, owing to the mechanical interactions among aggregated particles. The optofluidic lattice also enables single-bacteria-level screening of biological binding agents such as antibodies through particle-enabled bacteria hopping. The binding efficiency of antibodies could be determined directly, selectively, quantitatively and efficiently. This work enriches the fundamental mechanisms of particle kinetics and offers new possibilities for probing and utilising unprecedented biomolecule interactions at single-bacteria level.
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Bactérias , Nanopartículas/química , Cinética , Técnicas Analíticas Microfluídicas/métodosRESUMO
Surface plasmon field-enhanced fluorescence is reported for the readout of a heterogeneous assay that utilizes low affinity split aptamer ligands. Weak affinity ligands that reversibly interact with target analytes hold potential for facile implementation in continuous monitoring biosensor systems. This functionality is not possible without the regeneration of more commonly used assays relying on high affinity ligands and end-point measurement. In fluorescence-based sensors, the use of low affinity ligands allows avoiding this step but it imposes a challenge associated with the weak optical response to the specific capture of the target analyte which is also often masked by a strong background. The coupling of fluorophore labels with a confined field of surface plasmons is reported for strong amplification of the fluorescence signal emitted from the sensor surface and its efficient discrimination from the background. This optical scheme is demonstrated for time-resolved analysis of chosen model analytes - adenoside and adenosine triphosphate - with a split aptamer that exhibits an equilibrium affinity binding constant between 0.73 and 1.35 mM. The developed biosensor enables rapid and specific discrimination of target analyte concentration changes from low µM to mM in buffer as well as in 10% serum.
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Aptâmeros de Nucleotídeos/química , Técnicas Biossensoriais , Ressonância de Plasmônio de Superfície , Corantes FluorescentesRESUMO
An optofluidic chip is demonstrated in experiments for high-resolution and multi-range particle separation through the optically-induced microscopic vibration effect, where nanoparticles are trapped in loosely overdamped optical potential wells created with combined optical and fluidic constraints. It is the first demonstration of separating single nanoparticles with diameters ranging from 60 to 100 nm with a resolution of 10 nm. Nanoparticles vibrate with an amplitude of 3-7 µm in the loosely overdamped potential wells in the microchannel. The proposed optofluidic device is capable of high-resolution particle separation at both nanoscale and microscale without reconfiguring the device. The separation of bacteria from other larger cells is accomplished using the same chip and operation conditions. The unique trapping mechanism and the superb performance in high-resolution and multi-range particle separation of the proposed optofluidic chip promise great potential for a diverse range of biomedical applications.
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Correction for 'Optofluidic lens with low spherical and low field curvature aberrations' by H. T. Zhao et al., Lab Chip, 2016, 16, 1617-1624.
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This paper reports an optofluidic lens with low spherical and low field curvature aberrations through the desired refractive index profile by precisely controlling the mixing between ethylene glycol and deionized water in an optofluidic chip. The experimental results demonstrate that the spherical aberration is reduced to 19.5 µm and the full width at half maximum of the focal point is 7.8 µm with a wide divergence angle of 35 degrees. In addition, the optofluidic lens can focus light at different off-axis positions on the focal plane with Δx' < 6.8 µm and at opposite transverse positions with |Δy - Δy'| < 5.7 µm. This is the first demonstration of a special optofluidic lens that significantly reduces both the spherical and field curvature aberrations, which enhances the focusing power and facilitates multiple light source illumination using a single lens. It is anticipated to have high potential for applications such as on-chip light manipulation, sample illumination and multiplexed detection.
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Cell refractive index is a key biophysical parameter, which has been extensively studied. It is correlated with other cell biophysical properties including mechanical, electrical and optical properties, and not only represents the intracellular mass and concentration of a cell, but also provides important insight for various biological models. Measurement techniques developed earlier only measure the effective refractive index of a cell or a cell suspension, providing only limited information on cell refractive index and hence hindering its in-depth analysis and correlation. Recently, the emergence of microfluidic, photonic and imaging technologies has enabled the manipulation of a single cell and the 3D refractive index of a single cell down to sub-micron resolution, providing powerful tools to study cells based on refractive index. In this review, we provide an overview of cell refractive index models and measurement techniques including microfluidic chip-based techniques for the last 50 years, present the applications and significance of cell refractive index in cell biology, hematology, and pathology, and discuss future research trends in the field, including 3D imaging methods, integration with microfluidics and potential applications in new and breakthrough research areas.
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Biologia Celular , Doença , Refratometria/métodos , Animais , Hematologia , HumanosRESUMO
In laboratory-based biofouling assays, the influence of physico-chemical surface characteristics on barnacle settlement has been tested most frequently using the model organism Balanus amphitrite (= Amphibalanus amphitrite). Very few studies have addressed the settlement preferences of other barnacle species, such as Balanus improvisus (= Amphibalanus improvisus). This study aimed to unravel the effects of surface physico-chemical cues, in particular surface-free energy (SFE) and surface charge, on the settlement of cyprids of B. improvisus. The use of well-defined surfaces under controlled conditions further facilitates comparison of the results with recent similar data for B. amphitrite. Zero-day-old cyprids of B. improvisus were exposed to a series of model surfaces, namely self-assembled monolayers (SAMs) of alkanethiols with varying end-groups, homogenously applied to gold-coated polystyrene (PS) Petri dishes. As with B. amphitrite, settlement of cyprids of B. improvisus was influenced by both SFE and charge, with higher settlement on low-energy (hydrophobic) surfaces and negatively charged SAMs. Positively charged SAMs resulted in low settlement, with intermediate settlement on neutral SAMs of similar SFE. In conclusion, it is demonstrated that despite previous suggestions to the contrary, these two species of barnacle show similar preferences in response to SFE; they also respond similarly to charge. These findings have positive implications for the development of novel antifouling (AF) coatings and support the importance of consistency in substratum choice for assays designed to compare surface preferences of fouling organisms.
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Incrustação Biológica/prevenção & controle , Thoracica/fisiologia , Animais , Comportamento Animal , Fenômenos Eletromagnéticos , Propriedades de SuperfícieRESUMO
A facile route for sensitive label-free detection of bio-toxins using aligned single walled carbon nanotubes is described. This approach involves patterning of a catalyst on the surface of a quartz substrate using a sub-100 µm stripe-patterned polydimethylsiloxane stamp for aligned carbon nanotube generation followed by fabrication of field effect transistor (FET). Atomic force microscopy, field emission scanning electron microscopy and Raman spectroscopy are employed to characterize the synthesized nanotubes. Unlike previous reports, the adopted approach enables direct electronic detection of bio-toxins with sensitivities comparable to ELISA. As a proof of concept, the fabricated FET responds to nM concentration levels (with a LOD of â¼2 nM) of epsilon toxin produced by Clostridium perfringens and a prominent food toxin. This facile approach could be customized to detect other classes of toxins and biomarkers upon appropriate functionalization of the aligned carbon nanotubes. Finally, we demonstrate the use of the FET-platform for detection of toxin in more complex matrices such as orange juice.
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Toxinas Bacterianas/análise , Condutometria/instrumentação , Análise de Alimentos/instrumentação , Contaminação de Alimentos/análise , Nanotubos de Carbono/química , Nanotubos de Carbono/ultraestrutura , Transistores Eletrônicos , Técnicas Biossensoriais/instrumentação , Desenho de Equipamento , Análise de Falha de Equipamento , Coloração e RotulagemRESUMO
A facile and high performance biosensing platform using aligned carbon nanotubes on quartz substrate is reported in this communication. Single walled carbon nanotubes are grown on quartz substrates by a chemical vapor deposition process and are characterized with field emission scanning electron microscopy and atomic force microscopy in order to verify the quality of the material. The quartz substrate is then directly used as a biosensor in a field effect transistor configuration. In order to demonstrate the sensing capabilities of the fabricated sensor devices, electronic detection of prostate specific antigen, a potential cancer biomarker, is carried out by adopting liquid gated configuration. A conductivity change due to the specific binding of target antigen with the immobilized receptor antibody demonstrates the sensing capabilities of the fabricated device. Sub-nM detection sensitivities have been obtained using the adopted direct immunoassay approach, which shows that the device responds to clinically relevant concentration regimes.
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Técnicas Biossensoriais/instrumentação , Análise de Injeção de Fluxo/instrumentação , Imunoensaio/instrumentação , Técnicas Analíticas Microfluídicas/instrumentação , Nanotubos de Carbono/química , Antígeno Prostático Específico/análise , Quartzo/química , Desenho de Equipamento , Análise de Falha de Equipamento , Nanotubos de Carbono/ultraestrutura , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Transistores EletrônicosRESUMO
We recently reported on the strong interactions of zoospores of the green alga, Ulva linza with an arginine-rich oligopeptide self-assembled monolayer (SAM) [Biofouling 2008, 24, 303-312], where the arginine-rich peptide induced not only high spore settlement, but also a form of abnormal settlement, or "pseudo-settlement", whereby a proportion of spores do not go through the normal process of surface exploration, adhesive exocytosis, and loss of flagella. Further, it was demonstrated that both the total number of settled spores and the fraction of pseudosettled spores were related to the surface density of the arginine-rich peptide. Here we present a further investigation of the interactions of zoospores of Ulva with a set of oligomeric, de novo designed, arginine-rich peptides, specifically aimed to test the effect of peptide primary structure on the interaction. Via variations in the peptide length and by permutations in the amino acid sequences, we gain further insight into the spore-surface interactions. The interpretation of the biological assays is supported by physicochemical characterization of the SAMs using infrared spectroscopy, ellipsometry, and contact angle measurements. Results confirm the importance of arginine residues for the anomalous pseudosettlement, and we found that settlement is modulated by variations in both the total length and peptide primary structure. To elucidate the causes of the anomalous settlement and the possible relation to peptide-membrane interactions, we also compared the settlement of the "naked" zoospores of Ulva (which present a lipoprotein membrane to the exterior without a discrete polysaccharide cell wall), with the settlement of diatoms (unicellular algae that are surrounded by a silica cell wall), onto the peptide SAMs. Cationic SAMs do not notably affect settlement (attachment), adhesion strength, or viability of diatom cells, suggesting that the effect of the peptides on zoospores of Ulva is mediated via specific peptide-membrane interactions.
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Arginina/química , Oligopeptídeos/química , Esporos/química , Ulva/química , Sequência de Aminoácidos , Arginina/metabolismo , Adesão Celular , Dados de Sequência Molecular , Estrutura Molecular , Oligopeptídeos/metabolismo , Esporos/metabolismo , Propriedades de SuperfícieRESUMO
Identification of settlement cues for marine fouling organisms opens up new strategies and methods for biofouling prevention, and enables the development of more effective antifouling materials. To this end, the settlement behaviour of zoospores of the green alga Ulva linza onto cationic oligopeptide self-assembled monolayers (SAMs) has been investigated. The spores interact strongly with lysine- and arginine-rich SAMs, and their settlement appears to be stimulated by these surfaces. Of particular interest is an arginine-rich oligopeptide, which is effective in attracting spores to the surface, but in a way which leaves a large fraction of the settled spores attached to the surface in an anomalous fashion. These 'pseudo-settled' spores are relatively easily detached from the surface and do not undergo the full range of cellular responses associated with normal commitment to settlement. This is a hitherto undocumented mode of settlement, and surface dilution of the arginine-rich peptide with a neutral triglycine peptide demonstrates that both normal and anomalous settlement is proportional to the surface density of the arginine-rich peptide. The settlement experiments are complemented with physical studies of the oligopeptide SAMs, before and after extended immersion in artificial seawater, using infrared spectroscopy, null ellipsometry and contact angle measurements.
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Cátions/química , Oligopeptídeos/química , Ulva/fisiologia , Conformação Proteica , Água do Mar , Esporos/fisiologia , Propriedades de SuperfícieRESUMO
This contribution describes how de novo designed synthetic helix-loop-helix polypeptides are utilized to control the assembly of gold nanoparticles and as scaffolds for biosensing. The synthetic polypeptides are designed to fold into a four-helix bundle upon dimerization. When immobilized on gold nanoparticles, dimerization and folding occur between peptides on neighbouring particles as an effect of particle aggregation and the folded polypeptides are rigid enough to keep the particles separated at a distance corresponding to the size of the four-helix bundle. Moreover, peptide dimerization offers a convenient route to assemble nanoparticles into hybrid multilayers on planar substrates. The drastic change in the resonance conditions of the localized nanoparticle surface plasmon upon particle aggregation is shown to be useful for optical detection of biomolecular interactions.
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Técnicas Biossensoriais , Nanopartículas/química , Peptídeos/síntese química , Anidrase Carbônica II/análise , Materiais Revestidos Biocompatíveis , Dimerização , Desenho de Fármacos , Ouro , Humanos , Microscopia Eletrônica , Modelos Moleculares , Complexos Multiproteicos , Nanopartículas/ultraestrutura , Nanotecnologia , Peptídeos/química , Dobramento de Proteína , Estrutura Quaternária de ProteínaRESUMO
Designed, synthetic polypeptides that assemble into four-helix bundles upon dimerization in solution were studied with respect to folding on planar gold surfaces. A model system with controllable dimerization properties was employed, consisting of negatively and positively charged peptides. Circular dichroism spectroscopy and surface plasmon resonance based measurements showed that at neutral pH, the peptides were able to form heterodimers in solution, but unfavorable electrostatic interactions prevented the formation of homodimers. The dimerization propensity was found to be both pH- and buffer-dependent. A series of infrared absorption-reflection spectroscopy experiments of the polypeptides attached to planar gold surfaces revealed that if the negatively charged peptide was immobilized from a loading solution where it was folded, its structure was retained on the surface provided it had a cysteine residue available for anchoring to gold. If it was immobilized as random coil, it remained unstructured on the surface but was able to fold through heterodimerization if subsequently exposed to a positively charged polypeptide. When the positively charged peptide was immobilized as random coil, heterodimerization could not be induced, probably because of high-affinity interactions between the charged primary amine groups and the gold surface. These observations are intended to pave the way for future engineering of functional surfaces based on polypeptide scaffolds where folding is known to be crucial for function.
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Ouro/química , Peptídeos/química , Sequência de Aminoácidos , Dicroísmo Circular , Dimerização , Dados de Sequência Molecular , Conformação Proteica , Ressonância de Plasmônio de SuperfícieRESUMO
The structural properties of self-assembled monolayers (SAMs) of oligo(ethylene glycol) (OEG)-terminated and amide-containing alkanethiols (HS(CH(2))(15)CONH(CH(2)CH(2)O)(6)H and related molecules with shorter alkyl or OEG portions) on gold are addressed. Optimized geometry of the molecular constituents, characteristic vibration frequencies, and transition dipole moments are obtained using density-functional theory methods with gradient corrections. These data are used to simulate IR reflection-absorption (RA) spectra associated with different OEG conformations. It is shown that the positions and relative intensities of all characteristic peaks in the fingerprint region are accurately reproduced by the model spectra within a narrow range of the tilt and rotation angles of the alkyl plane, which turns out to be nearly the same for the helical and all-trans OEG conformations. In contrast, the tilt of the OEG axis changes considerably under conformational transition from helical to all-trans OEG. By means of ab initio modeling, we also clarify other details of the molecular structure and orientation, including lateral hydrogen bonding, the latter of which is readily possessed by the SAMs in focus. These results are crucial for understanding phase and folding characteristics of OEG SAMs and other complex molecular assemblies. They are also expected to contribute to an improved understanding of the interaction with water, ions, and ultimately biological macromolecules.
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Etilenoglicol/química , Ouro/química , Modelos MolecularesRESUMO
The aim of the present study was to evaluate mastication, food selection and nutritional aspects in two groups of persons restored with fixed (FPD, N=44) and removable (RPD, N=40) partial dentures respectively. The subjects were part of a cohort study of 67-68-year-old men living in Malmö, Sweden. The two groups were very similar regarding social factors and the inclusion criteria were chosen so that the groups were very equal regarding oral factors, apart from the difference in fixed and removable partial dentures. The number of natural teeth, number of replaced teeth and occlusal contacts did not differ significantly between the two groups, nor did the distribution of maxillary and mandibular dentures. A comprehensive examination of several general health factors included a home interview of dietary habits. A clinical examination included a 20-minute oral examination with registration of number of teeth, FPDs, RPDs, and occlusal contacts. It also included masticatory tests: chewing gum colour mixing, chewing gum bolus shaping, and swallowing threshold (number of strokes to the first swallow of an almond). The consumption of hard and soft foods was revealed by the dietary interview as well as the intake of energy and some nutrients. There was a significant difference between the groups regarding the capacity to mix the two-coloured chewing gum, to shape the chewing gum bolus and in the consumption of hard foods. There was no difference in the swallowing threshold and the consumption of soft foods. The intake of energy and nutrients did not differ significantly between the groups. The differences in masticatory capacity found thus seem to have little, if any, effect on the factors of importance for general health. A reasonable explanation for the differences found is that artificial teeth that are well retained, such as FPDs, make more active chewing possible than do removable, and often somewhat loose-fitting partial dentures.
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Prótese Parcial Fixa , Prótese Parcial Removível , Mastigação , Estado Nutricional , Idoso , Goma de Mascar , Estudos de Coortes , Deglutição , Demografia , Análise do Estresse Dentário , Ingestão de Energia , Preferências Alimentares , Humanos , Masculino , Nozes , Prunus , Estatísticas não ParamétricasRESUMO
The development of commercial biosensors based on surface plasmon resonance has made possible careful characterization of biomolecular interactions. Here, a set of destabilized human carbonic anhydrase II (HCA II) mutants was investigated with respect to their interaction kinetics with two different immobilized benzenesulfonamide inhibitors. Point mutations were located distantly from the active site, and the destabilization energies were up to 23 kJ/mol. The dissociation rate of wild-type HCA II, as determined from the binding to the inhibitor with higher affinity, was 0.019 s(-1). For the mutants, dissociation rates were faster (0.022-0.025 s(-1)), and a correlation between faster dissociation and a high degree of destabilization was observed. We interpreted these results in terms of increased dynamics of the tertiary structures of the mutants. This interpretation was supported by entropy determinations, showing that the entropy of the native structure significantly increased upon destabilization of the protein molecule. Our findings demonstrate the applicability of modern biosensor technology in the study of subtle details in molecular interaction mechanisms, such as the long-range effect of point mutations on interaction kinetics.
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Técnicas Biossensoriais/métodos , Anidrase Carbônica II/análise , Ressonância de Plasmônio de Superfície/métodos , Anidrase Carbônica II/genética , Humanos , Cinética , Modelos Moleculares , Mutação , Sulfonamidas/química , BenzenossulfonamidasRESUMO
Low protein adsorption is believed advantageous for blood-contacting materials and ethylene glycols (EG)-based polymeric compounds are often attached to surfaces for this purpose. In the present study, the adsorption of fibrinogen, serum, and plasma were studied by ellipsometry on a series of well-defined oligo(EG) terminated alkane-thiols self-assembled on gold. The layers were prepared with compounds of the general structure HS-(CH2)15-CONH-EGn, where n = 2, 4, and 6. Methoxy-terminated tri(EG) undecanethiol and hydroxyl-terminated hexadecanethiol self-assembled monolayers (SAMs) were used as references. The results clearly demonstrate that the adsorption depends on the experimental conditions with small amounts of fibrinogen adsorbing from a single protein solution, but larger amounts of proteins from serum and plasma. The adsorption of fibrinogen and blood plasma decreased with an increasing number of EG repeats and was temperature-dependent. Significantly less serum adsorbed to methoxy tri(EG) than to hexa(EG) and more proteins remained on the latter surface after incubation in a sodium dodecyl sulfate (SDS) solution, indicating a looser protein binding to the methoxy-terminated surface. All surfaces adsorbed complement factor 3 (C3) from serum and plasma, although no surface-mediated complement activation was observed. The present study points to the importance of a careful choice of the protein model system before general statements regarding the protein repellant properties of potential surfaces can be made.
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Etilenoglicol/química , Fibrinogênio/metabolismo , Plasma/citologia , Adsorção , Etilenoglicol/metabolismo , Ouro , Heparina/química , Humanos , Modelos Químicos , Dodecilsulfato de Sódio/química , Temperatura , Fatores de TempoRESUMO
An approach based on electrochemistry to differentiate between phosphorylated and nonphosphorylated amino acid analogues adsorbed on gold is presented. Analogues of serine, threonine, and tyrosine, containing thiohexadecyl headgroups, were synthesized and assembled on gold, and the surface capacitance was evaluated using electrochemical impedance spectroscopy. A procedure for deprotection of tert-butyl phosphate protecting groups, on the monolayer, is also described. Characterizations of the assembled analogues by cyclic voltammetry, infrared spectroscopy, and ellipsometry are used to confirm the insulating properties of the monolayers and the outcome of surface modifications. The results from cyclic voltammetry show good insulating properties for the monolayers even after phosphate deprotection. The infrared measurements reveal well-ordered monolayers, and the thickness from ellipsometry is in good agreement with expectations from molecular modeling. The impedance experiments show a capacitance increase up to 0.6 microF/cm2 as phosphate groups are introduced. The results in this study indicate the possibility of using a surface chemical and impedance spectroscopy approach to detect the kinase/phosphatase activity and kinetics involved in phosphorylation reactions.
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Aminoácidos/análise , Serina/análogos & derivados , Treonina/análogos & derivados , Tirosina/análogos & derivados , Eletroquímica , Fosforilação , Espectrofotometria Infravermelho/métodosRESUMO
A strategy for the synthesis of a series of closely related oligo(ethylene glycol)-terminated alkanethiol amides (principally HS(CH(2))(m)CONH(CH(2)CH(2)O)(n)H; m = 2, 5, 11, 15, n = 1, 2, 4, 6, 8, 10, 12) and analogous esters has been developed. These compounds were made to study the structure and stability of self-assembled monolayers (SAMs) on gold in the prospect of designing new biosensing interfaces. For this purpose, monodisperse heterofunctional oligo(ethylene glycols) with up to 12 units were prepared. Selective monoacylation of the symmetrical tetra- and hexa(ethylene glycol) diols as their mesylates with the use of silver(I) oxide was performed. The synthetic approach was based on carbodiimide couplings of various oligo(ethylene glycol) derivatives to omega-(acetylthio) carboxylic acids via a terminal amino or hydroxyl function. SAM structures on gold were studied with respect to thickness, wettability (water contact angles approximately 30 degrees ), and conformation. A good fit was obtained for the relation between monolayer thickness (d) and the number of units in the oligo(ethylene glycol) chain (n): d = 2.8n + 21.8 (A). Interestingly, the corresponding infrared spectroscopy analysis showed a dramatic change in conformation of the oligomeric chains from all-trans (n = 4) to helical (n > or = 6) conformation. A crystalline helical structure was observed in the SAMs for n > 6.