One-dimensional and two-dimensional nanomaterials for the detection of multiple biomolecules

By utilizing nanomaterials including one-dimensional materials (1DMs) and two-dimensional materials (2DMs), the recent development for the determination of multiple biomolecules has been focused.     

One-dimensional and two-dimensional nanomaterials for the detection of multiple biomolecules

The complexity of biological samples determines that the detection of a single biomolecule is unable to satisfy actual needs. Moreover, the “false positives” results caused by a single biomolecule detections easily leads to erroneous clinical diagnosis and treatment. Thus, it is important for the homogenous quantification of multiple biomolecules in not only basic research but also practical application. As a consequent, a large number of literatures have been exploited to monitor multiple biomolecules in homogenous solution, enabling facilitating the development of the disease diagnosis, treatment as well as drug discovery. One-dimensional nanomaterials and two-dimensional nanomaterials have special physical and chemical properties, such as good electrochemical properties, stable structure, large specific surface area, and biocompatibility, which are widely used in electrochemical and fluorescent detection of biomolecules. This tutorial review highlights the recent development for the detection of multiple biomolecules by using nanomaterials including one-dimensional materials (1DMs) as well as two-dimensional materials (2DMs).     

Protein micropatterning using surfaces modified by self-assembled polystyrene microspheres

A technique for micropatterning of proteins on a nonplanar surface to improve the coverage and functionality of biomolecules is demonstrated. A nonplanar microstructure is created by the self-assembly of polystyrene microspheres into an array of microwells on a silicon wafer to allow the integration of a nonplanar spot on a planar chip. After the microspheres were deposited into the microwells, they were conjugated with proteins. The curve surfaces of the microspheres present more surface area for attaching biomolecules which will increase the density of biomolecules and, hence, the sensitivity for detection. Moreover, proteins immobilized on a curved surface can retain their native structures and function better than on a planar surface because of a smaller area of interaction between the protein and the substrate. Patterning of biomolecules was tested with two model fluorescent proteins. The results show that precise patterning of biomolecules on a nonplanar spot can be achieved with this technique.     

Development of multivalent macromolecular ligands for enhanced detection of biological targets

Macromolecular conjugates enable simultaneous binding of multiple ligands on one biological entity and these polyvalent interactions can be collectively stronger than the corresponding monovalent ligands. We have synthesized macromolecules and conjugated them with a lectin (Helix Pomatia lectin, HPA), and an antibody, both with shown affinities to certain bacteria. The binding ability was studied by flow cytometry and the results showed that the affinity of the biomolecules was greatly enhanced due to the polyvalent effect.     

Ultra-high resolution 17O solid-state NMR spectroscopy of biomolecules: a comprehensive spectral analysis of monosodium L-glutamate·monohydrate

Monosodium L-glutamate monohydrate, a multiple oxygen site (eight) compound, is used to demonstrate that a combination of high-resolution solid-state NMR spectroscopic techniques opens up new possibilities for (17)O as a nuclear probe of biomolecules. Eight oxygen sites have been resolved by double rotation (DOR) and multiple quantum (MQ) NMR experiments, despite the (17)O chemical shifts lying within a narrow shift range of <50 ppm. (17)O DOR NMR not only provides high sensitivity and spectral resolution, but also allows a complete set of the NMR parameters (chemical shift anisotropy and electric-field gradient) to be determined from the DOR spinning-sideband manifold. These (17)O NMR parameters provide an important multi-parameter comparison with the results from the quantum chemical NMR calculations, and enable unambiguous oxygen-site assignment and allow the hydrogen positions to be refined in the crystal lattice. The difference in sensitivity between DOR and MQ NMR experiments of oxygen in bio/organic molecules is also discussed. The data presented here clearly illustrates that a high resolution (17)O solid-state NMR methodology is now available for the study of biomolecules, offering new opportunities for resolving structural information and hence new molecular insights.     

The facile synthesis of an aldehyde-containing graft copolymer membrane for covalent protein capture with retention of protein functionality

The immobilization of biomolecules onto an insoluble carrier surface has always been a subject of great interest to enhance their resistance to pH and temperature, which aids in an increased activity lifespan as well as easy reuse of the said biomolecules. However, traditional methods are only able to provide single-layer biomolecular binding and require multiple chemical reactions to prepare the final substrate before the immobilization can be carried out properly. Here we report a facile one-step chemical synthesis of a new aldehyde-bearing graft copolymer via atom transfer radical polymerization (ATRP) for covalent protein capture in a multilayered approach to covalently capture bovine serum albumin (BSA) onto a polymeric membrane. The resultant protein-bound membrane illustrated the retention of BSA's stereoselective discrimination ability by binding to an excess of 2 mol of tryptophan/mol of BSA and demonstrated an enantioresolution of a 0.184 mM racemic tryptophan mixture with a time-averaged-separation factor of 2.9.     

Nanostructures for magnetically triggered release of drugs and biomolecules

Specific targeting and controlled release are crucial factors in the administration of drugs and therapeutic biomolecules. It has been shown that drug delivery systems can significantly benefit of the introduction of superparamagnetic nanoparticles in terms of both targeting and controlled release. Magnetic gradients can be used to target therapeutics to specific regions, while alternating magnetic fields produce frequency-dependent effects at the nanoparticle level. This review reports on the latest developments of multifunctional systems based on magnetic nanoparticles where the release of drugs and/or biomolecules is triggered by the application of an external magnetic field. The potentials of these systems are presented through examples in the fields of surface functionalized magnetic nanoparticles, magnetic polymer nanocomposites and magnetoliposomes. Recent results suggest the importance of integrating multiple functions within a single nanostructured device in order to successfully transport, localize and release drugs and biomolecules.     

Quantitative Single-Molecule Electrochemiluminescence Bioassay

A single-molecule electrochemiluminescence bioassay is developed here which allows imaging and direct quantification of single biomolecules. Imaging single biomolecules is realized by localizing the electrochemiluminescence events of the labeled molecules. Such an imaging system allows mapping the spatial distribution of biomolecules with electrochemiluminescence and contains quantitative single-molecule insights. We further quantify biomolecules by spatiotemporally merging the repeated reactions at one molecule site and then counting the clustered molecules. The proposed single-molecule electrochemiluminescence bioassay is used to detect carcinoembryonic antigen, showing a limit of detection of 67 attomole concentration which is 10 000 times better than conventional electrochemiluminescence bioassays. This spatial resolution and sensitivity enable single-molecule electrochemiluminescence bioassay a new toolbox for both specific bioimaging and ultrasensitive quantitative analysis.     

Temperature-responsive chromatography for the separation of biomolecules

Temperature-responsive chromatography for the separation of biomolecules utilizing poly(N-isopropylacrylamide) (PNIPAAm) and its copolymer-modified stationary phase is performed with an aqueous mobile phase without using organic solvent. The surface properties and function of the stationary phase are controlled by external temperature changes without changing the mobile-phase composition. This analytical system is based on nonspecific adsorption by the reversible transition of a hydrophilic-hydrophobic PNIPAAm-grafted surface. The driving force for retention is hydrophobic interaction between the solute molecules and the hydrophobized polymer chains on the stationary phase surface. The separation of the biomolecules, such as nucleotides and proteins was achieved by a dual temperature- and pH-responsive chromatography system. The electrostatic and hydrophobic interactions could be modulated simultaneously with the temperature in an aqueous mobile phase, thus the separation system would have potential applications in the separation of biomolecules. Additionally, chromatographic matrices prepared by a surface-initiated atom transfer radical polymerization (ATRP) exhibit a strong interaction with analytes, because the polymerization procedure forms a densely packed polymer, called a polymer brush, on the surfaces. The copolymer brush grafted surfaces prepared by ATRP was an effective tool for separating basic biomolecules by modulating the electrostatic and hydrophobic interactions. Applications of thermally responsive columns for the separations of biomolecules are reviewed here.     

Ultraviolet photoelectron spectroscopic study on the interface electronic structure of the L-cysteine on Pd surface

L-cysteine is one of the most versatile biomolecules with a unique metal-binding ability. L-cysteine has an outstanding role in the bioelectronics field as a linker between proteins of biomolecules and metal electrodes of the inorganic metals through multiple functional groups. The interface electronic structures between L-cysteine with metals deserve further investigation for applications in bioelectronics. However, the interface electronic structures of L-cysteine and metals have not been well understood. We have previously reported the existence of a new state between the highest occupied molecular orbital (HOMO) of L-cysteine and the Fermi level of the metals for L-cysteine/Au(111), L-cysteine/Ag(111), and L-cysteine/Cu(111) using photoemission spectroscopy and attributed the formation of the new state to an interaction of the d band with HOMO of L-cysteine. In this study, the electronic structure at the interfaces of L-cysteine on a Palladium (Pd) surface is investigated by ultraviolet photoemission spectroscopy (UPS) using synchrotron radiation including work function, secondary electron cutoff (SECO), and HOMO onset; the position of an interface state, charge injection barrier, and ionization energy are estimated. It is observed that thin-film spectra of L-cysteine on Pd surfaces in the valance top region are different from the L-cysteine thick films, and this can be attributed to an interaction between a sulfur-originated state of L-cysteine HOMO with Pd d orbitals. Also, a 0.6-eV SECO shift is estimated due to the charge transferring between L-cysteine and Pd. The results of SECO further confirm the weakening of the Pd–sulfur bond with increasing L-cysteine coverage on Pd.     

An Aqueous Electrolyte Gated Artificial Synapse with Synaptic Plasticity Selectively Mediated by Biomolecules

The emulation of functions and behaviors of biological synapses using electronic devices has inspired the development of artificial neural networks (ANNs) in biomedical interfaces. Despite the achievements, artificial synapses that can be selectively responsive to non-electroactive biomolecules and directly operate in biological environments are still lacking. Herein, we report an artificial synapse based on organic electrochemical transistors and investigate the selective modulation of its synaptic plasticity by glucose. The enzymatic reaction between glucose and glucose oxidase results in long-term modulation of the channel conductance, mimicking selective binding of biomolecules to their receptors and consequent long-term modulation of the synaptic weight. Moreover, the device shows enhanced synaptic behaviors in the blood serum at a higher glucose concentration, which suggests its potential application in vivo as artificial neurons. This work provides a step towards the fabrication of ANNs with synaptic plasticity selectively mediated by biomolecules for neuro-prosthetics and human-machine interfaces.     

Biosurfactants: Formulations,properties, and applications

In this review, we attempt to give an overview on the recent progress made on biosurfactants, surface-active biomolecules produced by microorganisms, which are a sustainable alternative to synthetic surfactants. Different biosurfactants, their production techniques, and their physical and chemical properties are discussed. There is a focus on recent studies related to surface properties and rheology of biosurfactants, both being properties which affect their ability to take part in a stable formulation. Biosurfactants can have applications in multiple different industrial sectors, such as agriculture, medicine, personal care, food, petroleum, etc. The specific properties important for applications in these sectors are discussed in detail.     

Biosensors based on immobilization of biomolecules by electrogenerated polymer films

The concept and potentialities of electrochemical procedures of biomolecule immobilization are described. The entrapment of biomolecules within electropolymerized films consists of the application of an appropriate potential to an electrode soaked in an aqueous solution containing monomer and biomolecules. This method of biosensor construction is compared with a two-step procedure based on the adsorption of an aqueous amphiphilic pyrrole monomer-biomolecule mixture on an electrode followed by the electropolymerization of the adsorbed monomers. Another approach is based on the electrogeneration of polymer films functionalized by specific groups allowing subsequently the attachment of biomolecules. The immobilization of biomolecules on these films by covalent binding or noncovalent interactions is described.     

New applications of multiply charged ionic probes as cleavable cross-linker and polymerization reagent

We have designed new cleavable cross-linkers for biomolecules containing the 2,6-bis(oxazolinyl)pyridine (pybox) lanthanum complex. These species can effectively donate multiple charges to afford cross-linker ions for target biomolecules. The cross-linkers are cleaved easily by adding water. Moreover, we were able to detect chain structures of target molecules, including biomolecules and carbon clusters, such as fullerene C₆₀, by the addition of some MS ionic probes. Cold-spray ionization mass spectrometry demonstrated the applications of multiply charged ionic probes as cleavable cross-linkers and polymerization reagents.     

Interactions between bio(macro)molecules: Models and methods

Attempts are often made to describe interactions between biomolecules on the basis of models which do not adequately consider the fundamental features of bioprocesses. A model is proposed, consisting of two extensive biomolecules located in a box with a solvent. Intermediates occurring along a path connecting the initial and the final states are specified. The central role played by dynamic aspects of the process and the important role of the Helmholtz energy surface are pointed out.     

Bio‐Orthogonal Polymer Coatings for Co‐Presentation of Biomolecules

Controlled presentation of biomolecules on synthetic substrates is an important aspect for biomaterials development. If the immobilization of multiple biomolecules is required, highly efficient orthogonal surface chemistries are needed to ensure the precision of the immobilization. In this communication, chemical vapor deposition (CVD) copolymerization is used to fabricate polymer coatings with controlled ratio of alkyne and pentafluorophenyl ester (Pfp‐ester) groups. Cyclic argine‐glycine‐aspartic acid (cRGD) adhesion peptide and epidermal growth factor (EGF) are immobilized through alkyne–azide cycloaddtion (“click” chemistry) and active ester–amine reaction, respectively. Cell studies with human umbilical vein endothelial cells (HUVEC) and A431 cell lines demonstrate the biological activity of the coimmobilized biomolecules.     

Theoretical model for photochemical or thermally activated immobilization of macromolecules

The covalent immobilization of macromolecules on surfaces and within 3-dimensional networks is quantitatively described using a model based on Poisson statistics. This model determines the immobilized density or layer thickness as a function of molecular weight of the macromolecule or radiant exposure prior to and following the surface deposition of the macromolecule. Measurements of immobilized layer thickness provide first-order rate constants for decomposition of the surface-bound linker molecules and an estimate of the surface-bound linker density. The model predicts the relative density of immunocomplexed antibodies as a function of the irradiation time used to immobilize antigens. By providing the average number of bonds to the immobilized molecule, the model enables studies of the effect of multiple bonds on the activity of biomolecules. Experimental data by the authors and from the literature validate the model.     

Investigation of the interaction between arsenic species and thiols via electrospray ionization tandem mass spectrometry

Arsenic species have been known to participate in a number of chemical and biological reactions, including oxidation-reduction reactions, acid-base reactions, covalent interactions, and methylation-demethylation reactions because of the element's multiple and interconvertible oxidation states. Little is known about the structure or bonding behavior between arsenic species and thiolcontaining biomolecules. Therefore, a better understanding of the bonding behavior and detailed information on the molecular structure for arsenic-thiol complexes is needed. As a result, we have investigated the interaction between arsenic species (arsenate (As^V), arsenite (As^III), monomethylarsonic acid (MMA^V), and dimethylarsinic acid (DMA^V)) with biomolecules containing thiol groups (glutathione and cysteine) by electrospray ionization mass spectrometry (ESI-MS). These compounds were dissolved in methanol/water solution and introduced into the MS instrument in order to elucidate the direct bonding behavior of thiol group of biomolecules with arsenic species. In addition, further detailed structural information on this complex was obtained by collision-induced dissociation (CID) measurements.In each mass spectrum for mixture solutions between arsenic species and thiol compounds, various peaks such as protonated arsenic-thiol complexes, protonated noncomplexed thiol compounds, sodium bound cluster ions, and proton bound cluster ions were observed. In these mass spectra, the arsenic complexes were formed by interaction with thiol groups on the cysteine residues. These arsenic-thiol complexes produced a variety of fragment ions by cleavage of chemical bonds, and by interaction of other binding site on thiol compounds in tandem mass spectrometry experiments.     

Nano-bio interface study between Fe content TiO2 nanoparticles and adenosine triphosphate biomolecules

The advent of nano-biotechnology has inspired the interface interaction study between engineered nanoparticles (NPs) and biomolecules. The interaction between Fe content titanium dioxide (TiO₂) NPs and adenosine triphosphate (ATP) biomolecules has been envisioned. The effect of Fe content in TiO₂ matrix was studied using X-ray diffraction (XRD) and transmission electron microscopy (TEM). The increase in Fe content caused a decrease in particle size with change in morphology from spherical to one-dimensional rod structure. The Fe incorporation in the TiO₂ matrix reduced the transition temperature from anatase to rutile (A-R) phase along with formation of haematite phase of iron oxide at 400°C. The interaction of Fe content TiO₂ NPs with ATP molecule has been studied using spectroscopic method of Raman scattering and infrared vibration spectrum along with TEM. Fe content in TiO₂ has enhanced the interaction efficiency of the NPs with ATP biomolecules. Raman spectroscopy confirms that the NPs interact strongly with nitrogen (N₇) site in the adenine ring of ATP biomolecule. Engineering of Fe content TiO₂ NP could successfully tune the coordination between metal oxide NPs with biomolecules, which could help in designing devices for biomedical applications.     

Rapid multitarget immunomagnetic separation through programmable DNA linker displacement

Immunomagnetic separation has become an essential tool for high-throughput and low-cost isolation of biomolecules and cells from heterogeneous samples. However, as magnetic selection is essentially a "black-and-white" assay, its application has been largely restricted to single-target and single-parameter studies. To address this issue, we have developed an immunomagnetic separation technology that can quickly sort multiple targets in high yield and purity using selectively displaceable DNA linkers. We envision that this technology will be readily adopted for experiments requiring high-throughput selection of multiple targets or further adapted for selection of a single target based on multiple surface epitopes.