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PubMed2002

The scanning probe microscopy of metalloproteins and metalloenzymes.

In recent years, the concept of microscopy and the ability to study processes at a truly molecular level have been revolutionised by the development of a family of instruments based on acquiring data through the scanning of a proximal probe across a surface. Scanning Probe Microscopes (SPMs) enable surface-confined structures to be resolved at ångstrom-resolution, in real time, and under a variety of controllable conditions. Despite initial difficulties, much progress has been in the application of this technology to the high-resolution analysis of biological systems; these have varied from complex cellular systems to molecular biopolymers. Studying the interactions of protein with surfaces has been intrinsic to the development of our understanding of blood coagulation, fibrinolysis, thrombus formation and the synthesis of biocompatible materials. The specific interactions of metalloproteins and enzymes with electrode surfaces remains central to the understanding of the bioelectrochemical processes and to the development of biosensing devices. Though ellipsometry, Raman, microcalorimetry, surface plasmon resonance, and other spectroscopic methods, can provide much information on these interfaces, the acquired data are averaged over a large number of molecular species with a low spatial resolution. Proximal probe methods have much to offer in this regard and have revolutionized our ability to monitor such interactions.

Animals↗
PubMed2003

Enzyme-dispersed carbon-nanotube electrodes: a needle microsensor for monitoring glucose.

The preparation of an enzyme-dispersed carbon-nanotube (CNT) electrode, based on mixing glucose oxidase (GOx) within CNT, is described. The new binderless biocomposite was packed within a 21-gauge needle and used for amperometric monitoring of glucose. The resulting microsensor offers a low-potential highly selective and sensitive detection of glucose. The high sensitivity and selectivity are coupled to a wide linear range, prolonged lifetime and oxygen independence. About 80% of the GOx activity is retained during a 24 h thermal stress at 90 degrees C, reflecting the enzyme-stabilization action of CNT. The marked electrocatalytic action towards hydrogen peroxide allows highly selective detection of the glucose substrate at -0.1 V (vs. Ag/AgCl) with no interferences from coexisting ascorbic acid, acetaminophen or uric acid. Linearity prevails up to 40 mM glucose (with analytically useful signals observed up to 0.1 M). Factors affecting the performance of the CNT-based glucose biosensor were assessed and optimized. The attractive performance of the new needle electrode offers great promise for continuous monitoring of glucose in connection to the management of diabetes, and for the biosensing of other metabolites.

Electrochemistry↗
PubMed2005

Exploring the electrocatalytic sites of carbon nanotubes for NADH detection: an edge plane pyrolytic graphite electrode study.

The electrocatalytic properties of multi-walled carbon nanotube modified electrodes toward the oxidation of NADH are critically evaluated. Carbon nanotube modified electrodes are examined and compared with boron-doped diamond and glassy carbon electrodes, and most importantly, edge plane and basal pyrolytic graphite electrodes. It is found that CNT modified electrodes are no more reactive than edge plane pyrolytic graphite electrodes with the comparison with edge plane and basal plane pyrolytic graphite electrodes allowing the electroactive sites for the electrochemical oxidation of NADH to be unambiguously determined as due to edge plane sites. Using these highly reactive edge plane sites, edge plane pyrolytic graphite electrodes are examined with cyclic voltammetry and amperometry for the electroanalytical determination of NADH. It is demonstrated that a detection limit of 5 microM is possible with cyclic voltammetry or 0.3 microM using amperometry suggesting that edge plane pyrolytic graphite electrodes can conveniently replace carbon nanotube modified glassy carbon electrodes for biosensing applications with the relative advantages of reactivity, cost and simplicity of preparation. We advocate the routine use of edge plane and basal plane pyrolytic graphite electrodes in studies utilising carbon nanotubes particularly if 'electrocatalytic' properties are claimed for the latter.

Animals↗
PubMed2005

Fabrication of nanopatterned films of bovine serum albumin and staphylococcal protein A using latex particle lithography.

Arrays of protein nanostructures can be formed on surfaces such as mica(0001) and Au(111) using lithography with polystyrene latex particles. To create arrays of protein nanostructures, monodisperse latex spheres are mixed with the desired protein (e.g. BSA, protein A or IgG) and deposited onto substrates. Protein-coated nanospheres self-assemble into organized crystalline layers when dried on flat surfaces. After rinsing with water, dried latex spheres are displaced to expose periodic arrays of uncovered circular cavities. The immobilized proteins remain attached to the surface and form nanopatterns over broad areas (microns) corresponding to the thickness of a single layer of proteins. The nanostructures of immobilized proteins maintain the order and periodicity of the latex scaffold. The morphology and diameter of the protein nanostructures are tuneable by selecting the ratios of protein-to-latex and the diameters of latex spheres. Well-defined nanostructured surfaces of proteins supply a tool for fundamental investigations of protein binding interactions in biological systems at the nanoscale and have potential applications in biochip and biosensing systems.

Animals↗
PubMed2026

Colorimetric gold nanosensors for monitoring protein aggregation: implications for Alzheimer's disease.

Alzheimer's disease (AD) is the leading cause of dementia worldwide. It remains a major public health challenge due to the lack of early diagnostic tools and effective disease-modifying therapies. Molecularly, AD is characterized by extracellular amyloid-β (Aβ) plaques and intracellular Tau tangles, as well as soluble oligomers that are likely the neurotoxic species. However, the transient and heterogeneous nature of these oligomers makes them difficult to detect using conventional biosensing approaches. Nanomaterial-based colorimetric biosensors have emerged as promising platforms for detecting protein aggregates and discovering aggregation inhibitors. Specifically, the localized surface plasmon resonance properties of metallic nanomaterials can enable rapid, label-free, and visually detectable colorimetric sensing of molecular interactions. These features can be leveraged to monitor protein aggregation processes in real time and achieve high-throughput screening of aggregation inhibitors, which may collectively enable early detection and timely intervention of AD progression. This Review Article presents the design and engineering of gold-nanomaterial-based colorimetric biosensors for monitoring protein aggregation and highlights the current challenges and emerging opportunities for applying these nanosensors to combat AD.

Journal Article↗
PubMed2003

Use of electrogram characteristics during sinus rhythm to delineate the endocardial scar in a porcine model of healed myocardial infarction.

INTRODUCTION: Substrate-based catheter ablation of postmyocardial infarction (post-MI) ventricular tachycardia necessitates electroanatomic definition of the scarred endocardium. We sought to determine whether electrogram criteria during sinus rhythm could identify the location and extent of the myocardial scar by electroanatomic mapping. METHODS AND RESULTS: A porcine model of healed MI was generated by injecting agarose microspheres into the mid left anterior descending coronary artery. At least 4 weeks post-MI, the animals (n = 24) underwent detailed left ventricular endocardial electroanatomic mapping using a 4-mm-tip catheter (BioSense-Webster, Inc.). Based upon mapping data in normal animals, infarcted tissue was defined as bipolar electrogram amplitude < 1.5 mV and electrogram duration > or = 50 msec. Radiofrequency ablation lesions (2-10 per animal) were placed to tag the endocardial borders of the electroanatomic mapping-defined scar. The area of the scar defined by abnormal voltage amplitude was 25.9 +/- 15.4 cm2 (range 6.9-60.5). This area correlated well with that defined as scar by the electrogram duration criteria (26.4 +/- 16 cm2). Of those points remote from the infarct with falsely low voltage amplitude resulting from presumed poor catheter-tissue contact, 94% were correctly identified as normal when using the electrogram duration criteria. Late potentials were observed predominantly along the borders of the infarcted myocardium. The radiofrequency lesions placed to tag the scar borders were located along the scar periphery during gross pathologic examination. CONCLUSION: During normal sinus rhythm, both bipolar electrogram voltage amplitude and electrogram duration criteria are able to help differentiate normal from scarred myocardial tissue. Using these criteria, a detailed reconstruction of the endocardial scar can be rendered by electroanatomic mapping of the heart.

Animals↗
PubMed2005

Mechanisms of right atrial tachycardia occurring late after surgical closure of atrial septal defects.

UNLABELLED: Postatriotomy atrial tachycardia ablation. INTRODUCTION: In patients without structural heart disease, the most frequently occurring AT is the common atrial flutter. In patients with repaired congenital heart disease other mechanisms of AT may occur, due to the presence of an atriotomy that can provide a substrate for reentry. The aim of the present study was to identify the mechanisms of atrial tachycardia (AT) occurring late after atrial septum defect (ASD) repair, with the help of a three-dimensional electroanatomical mapping system. METHODS AND RESULTS: Twenty-two consecutive patients presenting with AT underwent complete electroanatomic mapping (CARTO, Biosense Webster, Diamond Bar, CA) of spontaneously occurring and inducible right ATs. Complete maps of 26 ATs were obtained. Three tachycardia mechanisms were identified: single-loop macroreentrant atrial tachycardia (MAT) (n=7), double-loop MAT (n=18), and focal AT (n=1). In all MATs, protected isthmuses were identified as the electrophysiological substrate of the arrhythmia, most frequently the cavotricuspid isthmus (CTI) (n=24), and a gap between the inferior vena cava and a line of double potentials (n=11). A mean number of 13.5+/-2.1 radiofrequency applications were delivered to transect these critical parts of the circuit. During a follow-up of 25+/-16 months the RF ablation was acutely successful in all patients. Thirteen patients (59%) had an early recurrence of MAT and needed an additional ablation procedure. One of those patients needed two additional ablation procedures. CONCLUSIONS: Three-dimensional electroanatomic mapping is useful to identify postsurgical AT mechanisms; the CTI isthmus is involved in 92% MAT, and if the right atrial free wall (RAFW) abnormal tissue related to surgical scar is present this substrate contributes to the MAT circuit.

Adult↗
PubMed2005

Catheter ablation of recipient right ventricular tachycardia after heterotopic heart transplantation.

Ventricular tachycardia after heart transplantation. A case is reported of ventricular tachycardia (VT) in a 62-year-old male after heterotopic heart transplantation, who occasionally had attacks of palpitation. Surface electrocardiogram suggested VT arising from the recipient heart. Intracardiac electrograms and entrainment mapping confirmed macroreentrant VT located in the recipient right ventricle. Radiofrequency ablation using an electroanatomical mapping system (CARTO, Biosense Webster, Diamond Bar, CA, USA) successfully eliminated VT.

Catheter Ablation↗
PubMed2003

Validation of R wave voltage endomyocardial mapping to assess myocardial fibrosis: comparison with thallium and dobutamine echocardiography in a swine model.

The first aim of this study was to validate R wave voltage values measured with a catheter-based three-dimensional mapping system (NOGA, Biosense) against myocardial fibrosis in a swine model of ameroid placement. The second aim was to correlate the UPV maps with thallium uptake and low-dose dobutamine echocardiography. The electromechanical catheter mapping system can be used to guide endomyocardial laser therapy and direct gene delivery to the myocardium. The accuracy of R wave voltage in identifying myocardial fibrosis and its comparison with other imaging technologies against histopathology has not been fully investigated in an animal model. Ameroid constrictors were placed on 24 vessels in 14 swine. Wall motion abnormalities by echocardiography were detected in distribution of the constrictors at 22 +/- 10 days. Animals underwent rest and delayed thallium imaging, low-dose dobutamine echocardiography, and R wave voltage mapping. Animals were sacrificed, hearts were removed, 4-micron slices were stained, and fibrosis was quantified. Five control animals were studied to obtain segment normalization. There was significant agreement between each imaging modality and fibrosis by chi-square analysis. The correlation for segment normalized thallium uptake and segment normalized UPV score versus fibrosis were both significant (P < 0.001) with an r2 value of 0.362 vs. thallium, and r2 = 0.445 vs. UPV. The correlation was improved using log of UPV vs. fibrosis (r = 0.532). These results help validate R wave voltage mapping as an imaging technique to identify myocardial fibrosis.

Animals↗
PubMed2002

Invasive electrophysiology in children: state of the art.

While noninvasive techniques, including esophageal recording and pacing, clearly have advantages in the pediatric population; they also have significant limitations. Invasive electrophysiology (EP) in children now encompasses the use of many advanced engineering applications, which contribute to the efficacy of such procedures. This is particularly true with respect to the performance of radiofrequency catheter ablation. First, microcatheters (eg, Cardima) offer advantages as diagnostic catheters in small hearts, and allow pacing and recording from both atrium and ventricle in even the smallest infants using limited venous access. In addition, there is the possibility of mapping the distal coronary sinus, and the right atrioventricular groove via the right coronary artery. Second, electroanatomic mapping using the CARTO system (Biosense Webster) allows complex maps to be constructed in patients with congenital heart disease and a history of cardiac surgery. Although somewhat laborious to construct, such maps offer great detail to guide ablation, and also provide voltage information to guide identification of patches and scars. Similarly, non-contact mapping systems (eg, EnSite, Endocardial Solutions) are available which record far-field potentials and employ solutions to the inverse problem to reconstruct endocardial potentials. Three-dimensional mapping based on a single beat is possible, and the success of ablation in creating conduction block in unique channels can be evaluated. Both of these three-dimensional mapping systems have the potential to improve outcomes in patients with complex disease undergoing ablation. Finally, patients are now undergoing combined procedures with interventional catheterization (eg, device closure, stenting) along with EP procedures (eg, ablation, device implant) in the same laboratory session. The state of the art in invasive pediatric EP increasingly involves the use of advanced technology adapted for use in pediatric and congenital heart disease applications.

Adolescent↗
PubMed1999

Image-guided surgery of the skull base using a novel miniature position sensor.

Image-guided navigational systems have been a useful adjunct for minimally invasive surgery of the skull base. A novel miniature position sensor has been developed that uses a low magnetic field for real-time tracking of surgical instruments. The 1.7-mm-diameter sensor attached to the position and orientation system (Magellan(R), Biosense, Inc., Johnson and Johnson Co., Baldwin Park, CA) was deployed through various surgical instruments or used in a hand-held fashion with a malleable shaft probe. We report on our experience using this electromagnetic system in a series of lesions of the sella and clivus. After patient/image registration, the system was consistently accurate to within 2 mm. We have found this system to be particularly advantageous in endoscopic surgery of cystic lesions of the skull base, where access is limited and anatomy may be distorted. In three patients, this device obviated the need for an extensive external surgical approach. Case histories are presented, which illustrate the specific advantages this miniature system provides during skull base surgery.

Journal Article↗
PubMed2000

Evaluation of left ventricular volumes and ejection fraction with a nonfluoroscopic endoventricular three-dimensional mapping technique.

BACKGROUND: Recently, a novel nonfluoroscopic 3-dimensional electromechanical mapping technique was introduced in the clinical arena. Although initial in vitro and in vivo studies suggested the reliability of the system in volumetric and hemodynamic evaluation of the left ventricle, no validation in human beings has been performed. METHODS: A nonfluoroscopic electromechanical mapping (NOGA, Biosense-Webster) procedure was performed in 44 patients. All patients received a contrast left ventriculogram during the same session. Volumetric (end-diastolic [EDV] and end-systolic volumes [ESV]) and hemodynamic (left ventricular ejection fraction [LVEF] and stroke volume) parameters of both systems were compared. RESULTS: Two uncomplicated pericardial effusions occurred with the first-generation mapping catheters. No procedural complications were noted with the new-generation mapping catheters. Significant correlations were found between mapping-derived and ventriculography-based measurements for both ESV (r = 0.67, P <.001) and LVEF (r = 0.78, P <.001). Absolute volumes, however, were only comparable for ESV (46.6 +/- 25.3 mL vs 48.8 +/- 37.0 mL, respectively; P =.13) but differed greatly for LVEF (35% +/- 13% vs 65% +/- 19%, respectively; P <.001), EDV (69.1 +/- 28.6 mL vs 125.9 +/- 53.4 mL, respectively; P <.001) and stroke volume (22.4 +/- 9.9 mL vs 77.1 +/- 33.7 respirations; P <.001). Moreover, Bland-Altman analysis showed the clinical noninterchangeability between these techniques for the measurement of hemodynamic parameters. CONCLUSION: Measurement of hemodynamic parameters with nonfluoroscopic mapping of the left ventricle is feasible and safe. The system provides data that strongly correlate but that are in clinical disagreement with angiographic data. Therefore the interchangeability of these techniques may be questioned.

Algorithms↗
PubMed2004

Triggered amplification by hybridization chain reaction.

We introduce the concept of hybridization chain reaction (HCR), in which stable DNA monomers assemble only upon exposure to a target DNA fragment. In the simplest version of this process, two stable species of DNA hairpins coexist in solution until the introduction of initiator strands triggers a cascade of hybridization events that yields nicked double helices analogous to alternating copolymers. The average molecular weight of the HCR products varies inversely with initiator concentration. Amplification of more diverse recognition events can be achieved by coupling HCR to aptamer triggers. This functionality allows DNA to act as an amplifying transducer for biosensing applications.

Base Sequence↗
PubMed2006

Collection, focusing, and metering of DNA in microchannels using addressable electrode arrays for portable low-power bioanalysis.

Although advances in microfluidic technology have enabled increasingly sophisticated biosensing and bioassay operations to be performed at the microscale, many of these applications employ such small amounts of charged biomolecules (DNA, proteins, and peptides) that they must first be preconcentrated to a detectable level. Efficient strategies for precisely handling minute quantities of biomolecules in microchannel geometries are critically needed; however, it has proven challenging to achieve simultaneous concentration, focusing, and metering capabilities with current-generation sample-injection technology. By using microfluidic chips incorporating arrays of individually addressable microfabricated electrodes, we demonstrate that DNA can be sequentially concentrated, focused into a narrow zone, metered, and injected into an analysis channel. This technique transports charged biomolecules between active electrodes upon application of a small potential difference (1 V) and is capable of achieving orders of magnitude concentration increases within a small device footprint. The collected samples are highly focused, with sample zone size and shape defined solely by electrode geometry.

DNA↗
PubMed2026

Ultrasound-driven mechanophore activation in living plants.

This study presents a biocompatible, ultrasound-responsive platform for remotely activating mechanochemical reactions within live plant tissue. Fluorogenic Mechanophore-embedded silica NanoParticles (FMNPs) that are thermally stable were engineered to emit blue fluorescence at 440 nm upon mechanical activation. In Solanum lycopersicum (tomato) leaves, activation was achieved through the synergistic combination of gas vesicles (GVs) and high-frequency focused ultrasound (FUS, 550 kHz), enabling spatially localized and minimally invasive stimulation. Low-frequency ultrasound (25 kHz) triggered activation but caused extensive tissue damage, while high-frequency FUS alone was biocompatible yet insufficient to activate FMNPs. Incorporation of GVs as a cavitation amplifier significantly boosted activation efficiency under mild acoustic conditions without observable tissue disruption. In planta fluorescence imaging confirmed that FMNPs retained their functionality after injection into leaf vasculature, and only the combination of GV and FUS produced a statistically significant fluorescence increase, indicating successful mechanochemical activation. This represents a demonstration of noninvasive and biocompatible ultrasound-induced mechanophore activation in live plants. This modular and noninvasive strategy opens possibilities for programmable release of regulatory and metabolic chemicals, biosensing, and synthetic molecular control in plant systems.

Plant Leaves↗
PubMed1995

Molecular organization of histidine-tagged biomolecules at self-assembled lipid interfaces using a novel class of chelator lipids.

In molecular biology, the expression of fusion proteins is a very useful and well-established technique for the identification and one-step purification of gene products. Even a short fused sequence of five or six histidines enables proteins to bind to an immobilized metal ion chelate complex. By synthesis of a class of chelator lipids, we have transferred this approach to the concept of self-assembly. The specific interaction and lateral organization of a fluorescent fusion molecule containing a C-terminal oligohistidine sequence was studied by film balance techniques in combination with epifluorescence microscopy. Due to the phase behavior of the various lipid mixtures used, the chelator lipids can be laterally structured, generating two-dimensional arrays of histidine-tagged biomolecules. Because of the large variety of fusion proteins already available, this concept represents a powerful technique for orientation and organization of proteins at lipid interfaces with applications in biosensing, biofunctionalization of nanostructured interfaces, two-dimensional crystallization, and studies of lipid-anchored proteins.

Amines↗