A protein-polymer hybrid mediated by DNA

Protein-polymer hybrids (PPHs) represent an important and rapidly expanding class of biomaterials. Typically in these hybrids the linkage between the protein and the polymer is covalent. Here we describe a straightforward approach to a noncovalent PPH that is mediated by DNA. Although noncovalent, the DNA-mediated approach affords the highly specific pairing and assembly properties of DNA. To obtain the protein-DNA conjugate for assembly of the PPH, we report here the first direct copper catalyzed azide-alkyne cycloaddition-based protein-DNA conjugation. This significantly simplifies access to protein-DNA conjugates. The protein-DNA conjugate and partner polymer-DNA conjugate are readily assembled through annealing of the cDNA strands to obtain the PPH, the assembly of which was confirmed via dynamic light scattering and fluorescence spectroscopy.     

Preferential oxidative addition in palladium(0)-catalyzed suzuki cross-coupling reactions of dihaloarenes with arylboronic acids

The study of Pd(0)-/t-Bu3P system as a powerful catalyst for the cross-coupling of n,m-dihaloarenes with 1 equiv of arylboronic acids is described. Our work demonstrated that the fate of the regenerated Pd(0) catalyst can be controlled when the appropriate ligand is employed. The results described here may lead to the development of new, efficient processes to conjugate polymers with controlled length which are potentially useful in molecular electronics.     

A newly developed immunoassay method based on optical measurement for Protein A detection

We describe the development of an immunoassay using an antibody-silver nanoparticle (Ab-AgNP) conjugate as a catalyst for the silver enhancement reaction. The immuno-reaction signals that were magnified by silver metal precipitation were quantified using a commercial flatbed scanner. Protein A from Staphylococcus aureus (S. aureus), a common clinical pathogenic bacterium, was used in this research. The ease of infection of S. aureus necessitates the development of a fast detection method. The framework of the method described in this paper is based on the sandwich immunoassay and contains a 1st antibody (immunoglobulin G, IgG), an antigen (Protein A), and a 2nd antibody-colloidal silver conjugate (IgG-AgNPs). The silver enhancement reaction, a signal amplification method in which silver ions are reduced to metallic silver, is used to magnify the immuno-reaction signal. The change in signal, as visualized in grayscale, can be easily observed and analyzed by our optical scanning detection system. The relationship between antigen concentration and grayscale value is discussed. The detectable concentration limit for the antigen was found to be 1 ng/mL with 10 μg/mL of IgG and 300 μM of the IgG-AgNP conjugate. This immunoassay method provides the advantages of low cost, easy operation, and short detection time. Moreover, it has potential applications in clinical diagnoses.     

On the search for glycated lipoprotein ApoA-I in the plasma of diabetic and nephropathic patients

The analysis of plasma samples from healthy, diabetic and nephropathic subjects was carried out by 2D gel electrophoresis. This approach shows clear differences among the three classes of subjects. In the case of diabetic and nephropathic patients intense spots appear. Their enzymatic digestion followed by matrix assisted laser desorption ionization/mass spectrometry (MALDI/MS) analysis shows that an overexpression of unglycated and glycated ApoA-I is present in both pathological states. Interestingly, this trend is also observed for the retinol-binding protein (RBP). The data obtained can be relevant to assess possible risks associated either with the glycation level of ApoA-I or with the overexpression of RBP. In fact, in the former case possibly a different functionality of the glycated protein is to be expected, reflecting a different efficiency in cholesterol transport. In the latter case, the increase of RBP level can be related to the overweight of the diabetic subjects under investigation: it is known that obesity leads to RBP overexpression. In the case of nephropathic patients, the RBP level increases in parallel with serum creatinin.     

Supramolecular Nested Microbeads as Building Blocks for Macroscopic Self‐Healing Scaffolds

The ability to construct self‐healing scaffolds that are injectable and capable of forming a designed morphology offers the possibility to engineer sustainable materials. Herein, we introduce supramolecular nested microbeads that can be used as building blocks to construct macroscopic self‐healing scaffolds. The core–shell microbeads remain in an “inert” state owing to the isolation of a pair of complementary polymers in a form that can be stored as an aqueous suspension. An annealing process after injection effectively induces the re‐construction of the microbead units, leading to supramolecular gelation in a preconfigured shape. The resulting macroscopic scaffold is dynamically stable, displaying self‐recovery in a self‐healing electronic conductor. This strategy of using the supramolecular assembled nested microbeads as building blocks represents an alternative to injectable hydrogel systems, and shows promise in the field of structural biomaterials and flexible electronics.     

基于一维和二维纳米材料的神经界面构筑

Neural interfaces have contributed significantly to our understanding of brain functions as well as the development of neural prosthetics. An ideal neural interface should create a seamless and reliable link between the nervous system and external electronics for long periods of time. Implantable electronics that are capable of recording and stimulating neuronal activities have been widely applied for the study of neural circuits or the treatment of neurodegenerative diseases. However, the relatively large cross-sectional footprints of conventional electronics can cause acute tissue damage during implantation. In addition, the mechanical mismatch between conventional rigid electronics and soft brain tissue has been shown to induce chronic tissue inflammatory responses, leading to signal degradation during long-term studies. Thus, it is essential to develop new strategies to overcome these existing challenges and construct more stable neural interfaces. Owing to their unique physical and chemical properties, one-dimensional (1D) and two-dimensional (2D) nanomaterials constitute promising candidates for next-generation neural interfaces. In particular, novel electronics based on 1D and 2D nanomaterials, including carbon nanotubes (CNTs), silicon nanowires (SiNWs), and graphene (GR), have been demonstrated for neural interfaces with improved performance. This review discusses recent developments in neural interfaces enabled by 1D and 2D nanomaterials and their electronics. The ability of CNTs to promote neuronal growth and electrical activity has been proven, demonstrating the feasibility of using CNTs as conducting layers or as modifying layers for electronics. Owing to their good mechanical, electrical and biological properties, CNTs-based electronics have been demonstrated for neural recording and stimulation, neurotransmitter detection, and controlled drug release. Different from CNTs-based electronics, SiNWs-based field effect transistors (FETs) and microelectrode arrays have been successfully demonstrated for intracellular recording of action potentials through penetration into neural cells. Significantly, SiNWs FETs can detect neural activity at the level of individual axons and dendrites with a high signal-to-noise ratio. Their ability to record multiplexed intracellular signals renders SiNWs-based electronics superior to traditional intracellular recording techniques such as patch-clamp recording. Besides, SiNWs have been explored for optically controlled nongenetic neuromodulation due to their tunable electrical and optical properties. As the star of the 2D nanomaterials family, GR has been applied as biomimetic substrates for neural regeneration. Transparent GR-based electronics combining electrophysiological measurements, optogenetics, two-photon microscopy with multicellular calcium imaging have been applied for the construction of multimodal neural interfaces. Finally, we provide an overview of the challenges and future perspectives of nanomaterial-based neural interfaces.     

Assay of human transthyretin-bound holo-retinol-binding protein with reversed-phase high-performance liquid chromatography

We describe a reversed-phase high-performance liquid chromatographic method for the determination of vitamin A-transporting (holo) transthyretin-bound (TTR) retinol-binding protein (RBP) concentrations in serum or plasma. Holo-TTR-RBP and free retinol derived primarily from free RBP are consistently observed with this chromatographic method. Holo-TTR-RBP concentrations determined by this method are highly correlated to holo-TTR-RBP concentrations measured by chromatography. This method has the advantage of using less expensive columns and having peak areas which are more proportional to their true concentrations in plasma, as determined by comparison to purified protein spectrophotometry and radial immunodiffusion. The percentage of RBP circulating as holo-TTR-RBP decreased significantly as the total concentration of RBP or retinol increased. Because purified holo-TTR-RBP did not dissociate under these chromatographic conditions, this suggests that more vitamin A circulates as holo-free RBP or free retinol in the blood of people with high serum RBP.     

Peptide‐poly(tert‐butyl methacrylate) conjugate into composite micelles in organic solvents versus peptide‐poly(methacrylic acid) conjugate into spherical and worm‐like micelles in water: Synthesis and self‐assembly

Peptide–polymer conjugates are versatile class of biomaterials composed of a peptide block covalently linked with a synthetic polymer block. This report demonstrates the synthesis of peptide‐poly(tert‐butyl methacrylate) (Peptide‐P^tBMA) conjugates of varying molecular weights via a “grafting from” atom transfer radical polymerization (ATRP) technique using as‐synthesized peptide‐based initiator in toluene. Peptide‐P^tBMA conjugate is soluble in many organic solvents and undergoes self‐assembly into micro/nanospheres in DMF/THF as observed from both FESEM and DLS results. The conjugate micro/nanospheres are nothing but the composite micelles formed by the secondary aggregation of primary micelles generated initially in these organic solvents. The hydrolysis of tert‐butyl groups of Peptide‐P^tBMA conjugate leads to the formation of peptide‐poly(methacrylic acid) (Peptide‐PMA) conjugate. The circular dichroism (CD) analysis exhibits the presence of β‐sheet conformation of peptide moiety in synthesized conjugates. The formed Peptide‐PMA conjugate is soluble in water and owing to its amphiphilic character, the conjugate molecules self‐assemble into spherical micelles as well as worm‐like micelles upon increasing the concentration of conjugate in water. However, the sodium salt of Peptide‐PMA conjugates (Peptide‐PMAS) self‐assembles into only spherical swollen micelles in water at higher (pH ～10). The critical aggregation concentrations (CACs) of both Peptide‐PMA and Peptide‐PMAS micelles are measured by fluorescence spectroscopy. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016 , 54, 3019–3031     

An ITO Based Disposable Biosensor for Ultrasensitive Analysis of Retinol Binding Protein

A highly sensitive electrochemical biosensor based on anti‐RBP biorecognition capable to analyze concentrations of retinol binding protein (RBP) was developed. The construction of the biosensor interfaces was carefully characterized by techniques such as electrochemistry, EIS, and scanning electron microscopy. In order to characterize impedance data, Kramers‐Kronig Transform was performed on the experimental impedance data. Besides, for an immunosensor system the Single Frequency Impedance technique was firstly used for the characterization of the interaction between RBP and anti‐RBP. Finally, artificial serum samples spiked with RBP were analyzed by the proposed ITO based immunosensor to investigate the usefulness of the biosensor for early biomarker diagnosis.     

An Auto‐Inductive Cascade for the Optical Sensing of Thiols in Aqueous Media: Application in the Detection of a VX Nerve Agent Mimic

A new auto‐inductive protocol employs a Meldrum's‐acid‐based conjugate acceptor ( 1 ) as a latent source of thiol for signal amplification, as well as optical detection of thiols. The auto‐induction is initiated by a thiol‐disulfide exchange that leads to the generation of β‐mercaptoethanol, which in turn decouples the conjugate acceptor to release more thiols, resulting in a self‐propagating cycle that continues until all the conjugate acceptor is consumed. Using 1 in a two‐step integrated protocol yields a rapid, sensitive, and precise diagnostic assay for the ultratrace quantitation of a thiophosphate nerve agent surrogate.     

组织工程相关生物材料表面工程的研究进展

生物材料用作人工细胞外基质(ECM ) 在组织工程中占据重要位置。本文在分析细胞2生物材料表面相互作用的基础上, 从生物材料中的水、材料表面的形态、材料表面的特异性识别及生物材料诱发愈合等方面探讨了生物材料的复杂性。生物材料对细胞的影响是一个双向、动态过程, 起着调节细胞增殖和凋亡平衡的作用。基于生物材料对细胞生长的影响, 本文提出了生物材料表面生物仿生化以提高细胞亲和力,糖链团簇、糖脂质及材料表面蛋白质修饰以提高细胞特异性识别, 材料表面的自组装修饰以改善表面形态等观点。     

Recent Advances in Biomaterials-Based Therapies for Alleviation and Regeneration of Traumatic Brain Injury

Traumatic brain injury (TBI), a major public health problem accompanied with numerous complications, usually leads to serve disability and huge financial burden. The adverse and unfavorable pathological environment triggers a series of secondary injuries, resulting in serious loss of nerve function and huge obstacle of endogenous nerve regeneration. With the advances in adaptive tissue regeneration biomaterials, regulation of detrimental microenvironment to reduce the secondary injury and to promote the neurogenesis becomes possible. The adaptive biomaterials could respond and regulate biochemical, cellular, and physiological events in the secondary injury, including excitotoxicity, oxidative stress, and neuroinflammation, to rebuild circumstances suitable for regeneration. In this review, the development of pathology after TBI is discussed, followed by the introduction of adaptive biomaterials based on various pathological characteristics. The adaptive biomaterials carried with neurotrophic factors and stem cells for TBI treatment are then summarized. Finally, the current drawbacks and future perspective of biomaterials for TBI treatment are suggested.     

Mixing Biomimetic Heterodimers of Nucleopeptides to Generate Biocompatible and Biostable Supramolecular Hydrogels

As a new class of biomaterials, most supramolecular hydrogels formed by small peptides require the attachment of long alkyl chains, multiple aromatic groups, or strong electrostatic interactions. Based on the fact that the most abundant protein assemblies in nature are dimeric, we select short peptide sequences from the interface of a heterodimer of proteins with known crystal structure to conjugate with nucleobases to form nucleopeptides. Being driven mainly by hydrogen bonds, the nucleopeptides self‐assemble to form nanofibers, which results in supramolecular hydrogels upon simple mixing of two distinct nucleopeptides in water. Moreover, besides being biocompatible to mammalian cells, the heterodimer of the nucleopeptides exhibit excellent proteolytic resistance against proteinase K. This work illustrates a new and rational approach to create soft biomaterials by a supramolecular hydrogelation triggered by mixing heterodimeric nucleopeptides.     

Development of a phoswich detector system for radioxenon monitoring

Measurement of radioactive xenon in the atmosphere is one of several techniques to detect nuclear weapons testing. For high sensitivity, some existing systems use beta/gamma coincidence detection to suppress background, which is very effective, but increases complexity due to separate beta and gamma detectors that require careful calibration and gain matching. In this paper, we will describe the development and evaluation of a simpler detector system, named PhosWatch, consisting of a CsI(Tl)/BC-404 phoswich well detector, digital readout electronics, and pulse shape analysis algorithms implemented in a digital signal processor on the electronics, and compare its performance to existing multi-detector systems.     

Enhanced lateral flow assay with double conjugates for the detection of exosomes

Exosomes are promising biological biomarkers for monitoring a number of diseases, especially cancers. Here, we developed a double gold nanoparticles(GNPs) conjugates based lateral flow assay(D-LFA) for rapidly and sensitively detecting and molecular profiling of exosomes. Based on these two GNPs conjugates, the signal amplification can be achieved without any additional operation. The antibody on the 1 st GNPs conjugate could recognize exosomes and form a sandwich format on the test zone. The 2 nd GNPs conjugate was designed to bind to the 1 st GNPs conjugate to realize signal amplification. This biosensor enabled visual and quantitative detection of exosomes by the accumulation of GNPs on the test zone and showed a low detection limit of 1.3×10~3 particles/μL, which has been improved 13-fold compared with the normal lateral flow assay. The D-LFA exhibited good sensitivity and reproducibility and has been successfully used for the detection of exosomes in fetal bovine serum,which proved its potential application in practical diagnostics.     

Time-resolved fluorimetric detection of terbium-labelled deoxyribonucleic acid separated by gel electrophoresis

Deoxyribonucleic acid (DNA) was reacted with a strong chelating agent and labelled with terbium, yielding a highly fluorescent conjugate with a lifetime of 1.5 ms. When a pulsed source and gated detection electronics were employed, the long-lived decay allowed effective discrimination against background fluorescence and scattered excitation. Detection limits should therefore be significantly improved in comparison with covalent labels with fluorescence lifetimes in the nanosecond regime or stains such as ethidium bromide. The conjugate is very stable, remaining fluorescent on dilution and in an electric field at elevated temperatures (60 degrees C), conditions typically encountered during polyacrylamide gel electrophoresis. As an enhancement solution is not required to develop the fluorescence, this system could be utilised in situations where on-line detection is desirable. Although only DNA was labelled, the method is equally applicable to ribonucleic acid.     

Molecular design strategies for biomaterials that heal

Biomaterials are widely used in medical devices with good success. However, the surface chemistries of our present generation of biomaterials are not specifically recognized by living organisms. Thus, biological reactions to biomaterials in use today are primarily influenced by non-specific interactions occurring at their surfaces. This paper develops a hypothesis for the development of a future generation of biomaterials. A discussion of the meaning of biocompatibility is followed by a strategy for developing biomaterials that actively induce healing and reconstruction by turning on specific biologic pathways. Materials strategies for encouraging specific reactions and inhibiting non-specific bioreaction are presented.     

Matrix assisted laser desorption/ionization-time of flight-mass spectrometry analysis of proteins detected by anti-phosphotyrosine antibody on two-dimensional-gels of fibrolast cell lysates after tumor necrosis factor-alpha stimulation

We describe efficient methods for using functional proteomics analysis to study signal transduction pathways in murine fibroblast L929 cells following stimulation with tumor necrosis factor (TNF)-alpha. After stimulation with TNF-alpha, cellular proteins of L929 cells were extracted with a lysis buffer containing 0.3% sodium dodecyl sulfate (SDS) for 10-30 min time intervals, and were separated by two-dimensional (2-D) electrophoresis followed by immunoblot analysis with anti-phosphotyrosine antibody and alkaline phosphatase-anti IgG antibody conjugate. To improve detection sensitivity by immunoblot analysis we used a chemifluorescent substrate for alkaline phosphatase. One hundred protein spots were detected in the TNF-alpha stimulated L929 cell extract by immunoblot analysis. The use of chemifluorescence allowed us to quantitate immunoblotted spots with fluoroscanner so that we were able to detect time-dependent changes of a number of immunoblotted spots. Protein spots on a silver-stained 2-D gel corresponding to those detected by immunoblot analysis were subjected to in-gel trypsin digestion- matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF)-mass spectrometry analysis, respectively. Twenty-one proteins detected by immunoblot analysis were identified by MS-Fit database search analysis. Among them, the proteins that show time-dependent changes in staining intensity include vimentin, tubulin beta-chain, eukaryotic translation initiation factor 1A, chromatin assembly factor 1 (P48 subunit), probable protein disulfide isomerase P5, and several other proteins. Vimentin and tubulin beta-chain have been reported to be phosphorylated at tyrosine residues and involved in the signal transduction pathway induced by TNF-alpha. However, the other proteins have no previously known function in the signal transduction pathway. Thus, the methods used in this study seem to be suitable for the identification of time-dependent changes in many proteins that are involved in signal transduction. Usefulness of the method for comprehensive analysis of the proteins involved in signal transduction pathway and the limitations of the method are discussed.     

Development of a multi-component chemically reactive detection conjugate for the determination of Hg(II) in water samples

Mercury ions (Hg(II)) are considered highly toxic and hazardous element even at low levels. The contamination of Hg(II) is a global problem. To develop selective and sensitive technique for the detection of Hg(II) has attracted considerable attention. In this study, a multi-component chemically reactive detection conjugate for determination of Hg(II) has been synthesized and a competitive format assay was proposed. In the technique, the chemically reactive capture conjugate was coated on the plate. The reactive detection conjugate was then captured by the capture conjugate. TMB solution was added and catalyzed by HRP molecules immobilized on AuNPs. Finally, the developed enzymatic signal was measured at 450 nm. The linear range of the assay was 0.35–350 ppb with a detection limit of 0.1 ppb. The average recoveries of Hg(II) from mineral water, tap water and lake water were 100.03%, 103.13% and 102.03%, respectively. All coefficients of variation (CVs) were less than 10%. The results are closely correlated with those from inductively coupled plasma mass spectrometry (ICP-MS), which indicated that the developed technique is a reliable method for and sensitive detection of Hg(II) in water samples.     

Control of the Structure and Functions of Biomaterials by Light

Vision and other light-triggered biochemical transformations in plants and living organisms represent a sophisticated biological processes in which optical signals are recorded and transduced as (physico)chemical events. Photoswitchable biomaterials are a new class of substances in which optical signals generate discrete “On” and “Off” states of biological functions, resembling logic gates that flip between 0 and 1 states in response to the changes in electric currents in computers. The (photo)chemistry of photochromic materials has been extensively developed in the past four decades. These materials isomerize reversibly upon light absorption, and the discrete photoisomeric states exhibit distinct spectral and chemical features. Integration of photoisomerizable (or photochromic) units into biomaterials allow their secondary functions such as biocatalysis, binding, and electron transfer to be tailored so that they can be switched on or off. This can be accomplished by chemical modification of the biomaterial by photoisomerizable units and by integration of biomaterials in photoisomerizable microenvironments such as monolayers or polymers. The photoswitchable properties of chemically modified biomaterials originate from the light-induced generation or perturbation of the biologically active site, whereas in photoisomerizable matrices they depend upon the regulation of the physical or chemical features of the photoisomerizable assemblies of polymers, monolayers, or membranes. Light-triggered activation of catalytic biomaterials provides a means of amplifying the recorded optical signal by biochemical transformations, and photostimulated biochemical redox switches allow its electrochemical transduction and amplification. The field of photoswitches based on biomaterials has developed extensively in the past few years within the general context of molecular switching devices and micromachinery. The extensive knowledge on the manipulation of biomaterials through genetic engineering and the fabrication of surfaces modified by biologically active materials enables us to prepare biomaterials with improved optical-switching features. Their application in optoelectronic or bioelectronic devices has been transformed from fantasy to reality. The use of photoswitchable biomaterials in information storage and processing devices (biocomputers), sensors, reversible immunosensors, and biological amplifiers of optical signals has already been demonstrated, but still leaves important future challenges.