Baking Powder Actuated Centrifugo-Pneumatic Valving for Automation of Multi-Step Bioassays.

We report a new flow control method for centrifugal microfluidic systems; CO₂ is released from on-board stored baking powder upon contact with an ancillary liquid. The elevated pressure generated drives the sample into a dead-end pneumatic chamber sealed by a dissolvable film (DF). This liquid incursion wets and dissolves the DF, thus opening the valve.

Xurography actuated valving for centrifugal flow control.

We introduce a novel instrument controlled valving scheme for centrifugal platforms which is based upon xurography. In a first approach, which is akin to previously presented event-triggered flow control, the valves are composed of a pneumatic chamber sealed by a dissolvable film (DF) and by a pierceable membrane. Liquid is initially prevented from wetting the DF by the counter pressure of a trapped gas.

Eosinophil peroxidase activates cells by HER2 receptor engagement and β1-integrin clustering with downstream MAPK cell signaling.

Eosinophils account for 1-3% of peripheral blood leukocytes and accumulate at sites of allergic inflammation, where they play a pathogenic role. Studies have shown that treatment with mepolizumab (an anti-IL-5 monoclonal antibody) is beneficial to patients with severe eosinophilic asthma, however, the mechanism of precisely how eosinophils mediate these pathogenic effects is uncertain. Eosinophils contain several cationic granule proteins, including Eosinophil Peroxidase (EPO).

Comprehensive N-Glycan Profiling of Avian Immunoglobulin Y.

Recent exploitation of the avian immune system has highlighted its suitability for the generation of high-quality, high-affinity antibodies to a wide range of antigens for a number of therapeutic and biotechnological applications. The glycosylation profile of potential immunoglobulin therapeutics is species specific and is heavily influenced by the cell-line/culture conditions used for production. Hence, knowledge of the carbohydrate moieties present on immunoglobulins is essential as certain glycan structures can adversely impact their physicochemical and biological properties.

Use of biosensors for the detection of marine toxins.

Increasing occurrences of harmful algal blooms (HABs) in the ocean are a major concern for countries around the globe, and with strong links between HABs and climate change and eutrophication, the occurrences are only set to increase. Of particular concern with regard to HABs is the presence of toxin-producing algae. Six major marine biotoxin groups are associated with HABs.

Antibodies and antibody-derived analytical biosensors.

The rapid diagnosis of many diseases and timely initiation of appropriate treatment are critical determinants that promote optimal clinical outcomes and general public health. Biosensors are now being applied for rapid diagnostics due to their capacity for point-of-care use with minimum need for operator input. Antibody-based biosensors or immunosensors have revolutionized diagnostics for the detection of a plethora of analytes such as disease markers, food and environmental contaminants, biological warfare agents and illicit drugs.

Development of internally controlled duplex real-time NASBA diagnostics assays for the detection of microorganisms associated with bacterial meningitis.

Three duplex molecular beacon based real-time Nucleic Acid Sequence Based Amplification (NASBA) assays have been designed and experimentally validated targeting RNA transcripts for the detection and identification of Haemophilus influenzae, Neisseria meningitidis and Streptococcus pneumoniae respectively. Each real-time NASBA diagnostics assay includes an endogenous non-competitive Internal Amplification Control (IAC) to amplify the splice variant 1 mRNA of the Homo sapiens TBP gene from human total RNA. All three duplex real-time NASBA diagnostics assays were determined to be 100% specific for the target species tested for.

Measurement of the viscoelastic properties of blood plasma clot formation in response to tissue factor concentration-dependent activation.

The coagulation of blood plasma in response to activation with a range of tissue factor (TF) concentrations was studied with a quartz crystal microbalance (QCM), where frequency and half width at half maximum (bandwidth) values measured from the conductance spectrum near resonant frequency were used. Continuous measurement of bandwidth along with the frequency allows for an understanding of the dissipative nature of the forming viscoelastic clot, thus providing information on the complex kinetics of the viscoelastic changes occurring during the clot formation process. Using a mathematical model, these changes in frequency and bandwidth have been used to derive novel QCM parameters of effective elasticity, effective mass density and rigidity factor of the viscoelastic layer.

Advances in ovarian cancer diagnosis: A journey from immunoassays to immunosensors.

This review focuses on the technological advancements, challenges and trends in immunoassay technologies for ovarian cancer diagnosis. Emphasis is placed on the principles of the technologies, their merits and limitations and on the evolution from laboratory-based methods to point-of-care devices. While the current market is predominantly associated with clinical immunoassay kits, over the last decade a major thrust in development of immunosensors is evident due to their potential in point-of-care devices.

A new approach to design an efficient micropost array for enhanced direct-current insulator-based dielectrophoretic trapping.

Direct-current insulator-based dielectrophoresis (DC-iDEP) is a well-known technique that benefits from the electric field gradients generated by an array of insulating posts to separate or trap biological particles. The aim of this study is to provide a first geometrical relationship of the post array that independent of the particles and/or medium, maximizes the trapping. A novel figure of merit is proposed to maximize the particle trapping in the post array while minimizing the required voltage, with a similar footprint and channel thickness.

Density-Gradient Mediated Band Extraction of Leukocytes from Whole Blood Using Centrifugo-Pneumatic Siphon Valving on Centrifugal Microfluidic Discs.

Here we present retrieval of Peripheral Blood Mononuclear Cells by density-gradient medium based centrifugation for subsequent analysis of the leukocytes on an integrated microfluidic "Lab-on-a-Disc" cartridge. Isolation of white blood cells constitutes a critical sample preparation step for many bioassays. Centrifugo-pneumatic siphon valves are particularly suited for blood processing as they function without need of surface treatment and are 'low-pass', i.

Phase-selective graphene oxide membranes for advanced microfluidic flow control.

For the first time, we harness the unique phase-selectivity of chip-integrated graphene oxide (GO) membranes to significantly enhance flow control on centrifugal microfluidic platforms. In this paper, we present novel processes for the assembly of these GO membranes into polymeric microfluidic systems and demonstrate that multilayer GO membranes allow the passage of water while blocking pressurized air and organic solutions.

Galectin-1 and Galectin-3 Constitute Novel-Binding Partners for Factor VIII.

Objective: Recent studies have demonstrated that galectin-1 (Gal-1) and galectin-3 (Gal-3) can bind von Willebrand factor and directly modulate von Willebrand factor-dependent early thrombus formation in vivo. Because the glycans expressed on human factor VIII (FVIII) are similar to those of von Willebrand factor, we investigated whether galectins might also bind and modulate the activity of FVIII.

Approach And Results: Immunosorbant assays and surface plasmon resonance analysis confirmed that Gal-1 and Gal-3 bound purified FVIII with high affinity.

Self-Powered Microfluidic Device for Rapid Assay of Antiplatelet Drugs.

We report the development of a microfluidic device for the rapid assay in whole blood of interfacial platelet-protein interactions indicative of the efficacy of antiplatelet drugs, for example, aspirin and Plavix, two of the world's most widely used drugs, in patients with cardiovascular disease (CVD). Because platelet adhesion to surface-confined protein matrices is an interfacial phenomenon modulated by fluid shear rates at the blood/protein interface, and because such binding is a better indicator of platelet function than platelet self-aggregation, we designed, fabricated, and characterized the performance of a family of disposable, self-powered microfluidic chips with well-defined flow and interfacial shear rates suitable for small blood volumes (≤200 μL). This work demonstrates that accurate quantification of cell adhesion to protein matrices, an important interfacial biological phenomenon, can be used as a powerful diagnostic tool in those with CVD, the world's leading cause of death.

Pulsed-Plasma Physical Vapor Deposition Approach Toward the Facile Synthesis of Multilayer and Monolayer Graphene for Anticoagulation Applications.

We demonstrate the growth of multilayer and single-layer graphene on copper foil using bipolar pulsed direct current (DC) magnetron sputtering of a graphite target in pure argon atmosphere. Single-layer graphene (SG) and few-layer graphene (FLG) films are deposited at temperatures ranging from 700 °C to 920 °C within <30 min. We find that the deposition and post-deposition annealing temperatures influence the layer thickness and quality of the graphene films formed.

Referencing cross-reactivity of detection antibodies for protein array experiments.

Protein arrays are frequently used to profile antibody repertoires in humans and animals. High-throughput protein array characterisation of complex antibody repertoires necessitates the use of extensively validated secondary detection antibodies. This article details the validation of an affinity-isolated anti-chicken IgY antibody produced in rabbit and a goat anti-rabbit IgG antibody conjugated with alkaline phosphatase using protein arrays consisting of 7,390 distinct human proteins.

Differential internalin A levels in biofilms of Listeria monocytogenes grown on different surfaces and nutrient conditions.

Listeria monoctyogenes is a foodborne pathogen containing the surface protein, internalin A (InlA). The expression of this protein permits the invasion of L. monocytogenes into intestinal epithelial cells expressing the receptor E-cadherin, thus crossing the intestinal barrier and resulting in listerosis.

Development of a rapid recombinase polymerase amplification assay for the detection of Streptococcus pneumoniae in whole blood.

Background: Streptococcus pneumoniae is an important cause of microbial disease in humans. The introduction of multivalent vaccines has coincided with a dramatic decrease in the number of pneumococcal-related deaths. In spite of this, at a global level, pneumococcal infection remains an important cause of death among children under 5 years of age and in adults 65 years of age or older.

The Purification of Natural and Recombinant Peptide Antibodies by Affinity Chromatographic Strategies.

The purification of peptide antibodies (e.g., IgG, IgY, scFv, and Fab) are described in this chapter.

The Structure of Natural and Recombinant Antibodies.

Immunoglobulins (Ig) isotypes A, D, E, G, and M are glycoproteins which are mainly composed of a "Y"-shaped Ig monomer (~150 kDa), consisting of two light and two heavy chains. Both light and heavy chains contain variable (N-terminal) and constant regions (C-terminal). Each light chain consists of one variable domain and one constant domain, whereas each heavy chain has one variable domain and three constant domains.

The need for effective pancreatic cancer detection and management: a biomarker-based strategy.

Pancreatic cancer (Pa) is generally a very aggressive disease, with few effective approaches available for early diagnosis or therapy. These factors, combined with the aggressiveness and chemoresistance of Pa, results in a bleak outcome post-diagnosis. Cancer-related biomarkers have established capabilities for diagnosis, prognosis and screening and can be exploited to aid in earlier less-invasive diagnosis and optimization of targeted therapies.

Cross Platform Standardisation of an Experimental Pipeline for Use in the Identification of Dysregulated Human Circulating MiRNAs.

Introduction: Micro RNAs (miRNAs) are a class of highly conserved small non-coding RNAs that play an important part in the post-transcriptional regulation of gene expression. A substantial number of miRNAs have been proposed as biomarkers for diseases. While reverse transcriptase Real-time PCR (RT-qPCR) is considered the gold standard for the evaluation and validation of miRNA biomarkers, small RNA sequencing is now routinely being adopted for the identification of dysregulated miRNAs.

Aspirin and P2Y12 inhibition attenuate platelet-induced ovarian cancer cell invasion.

Background: Platelet-cancer cell interactions play a key role in successful haematogenous metastasis. Disseminated malignancy is the leading cause of death among ovarian cancer patients. It is unknown why different ovarian cancers have different metastatic phenotypes.

Rapid, culture-independent, optical diagnostics of centrifugally captured bacteria from urine samples.

This work presents a polymeric centrifugal microfluidic platform for the rapid and sensitive identification of bacteria directly from urine, thus eliminating time-consuming cultivation steps. This "Lab-on-a-Disc" platform utilizes the rotationally induced centrifugal field to efficiently capture bacteria directly from suspension within a glass-polymer hybrid chip. Once trapped in an array of small V-shaped structures, the bacteria are readily available for spectroscopic characterization, such as Raman spectroscopic fingerprinting, providing valuable information on the characteristics of the captured bacteria.

Multivalent linkers for improved covalent binding of oligonucleotides to dye-doped silica nanoparticles.

This paper describes the fabrication of oligonucleotide-coated Cy5-doped silica nanoparticles using a combination of multivalent linkers and their use in surface-based DNA sandwich hybridization assays. Dipodal silane is introduced as a means to fabricate amine-coated silica nanoparticles and its advantages compared to monopodal silanes are discussed. The use of dipodal silane in conjunction with three different polymer linkers (oxidized dextran, linear and 8-arm polyethylene glycol (PEG)) to immobilize single-stranded DNA to Cy5-doped nanoparticles is investigated and dynamic light scattering measurements and Fourier transform infrared spectroscopy are used to follow the progression of the functionalization of the nanoparticles.