Christina G. Siontorou

Nanobodies as novel agents for disease diagnosis and therapy

The discovery of naturally occurring, heavy-chain only antibodies in Camelidae, and their further development into small recombinant nanobodies, presents attractive alternatives in drug delivery and imaging. Easily expressed in microorganisms and amenable to engineering, nanobody derivatives are soluble, stable, versatile, and have unique refolding capacities, reduced aggregation tendencies, and high-target binding capabilities. This review outlines the current state of the art in nanobodies, focusing on their structural features and properties, production, technology, and the potential for modulating immune functions and for targeting tumors, toxins, and microbes.

Innovation in biotechnology: moving from academic research to product development: the case of biosensors

The fast pace of technological change in the biotechnology industry and the market demands require continuous innovation, which, owing to the science base of the sector, derives from academic research through a transformation process that converts science-oriented knowledge to marketable products. There appear to be some inherent difficulties in transforming directly the knowledge output of academic research to industrial use. The purpose of this article is to examine certain transition mechanisms from monodisciplinary academic isolation (curiosity-driven and internal-worth innovation) to university-industry alliances (market-driven and public-worth innovation) through inter-organizational multidisciplinary collaboration and contextualize the analysis with the case of biosensors. While the majority of literature on the subject studies the channels of knowledge transfer as determinants of alliance success (transferor/transferee interactions), either from the university side (science base) or the industry side (market base), this article focuses on the transferable (technology base) and how it can be strategically modeled and managed by the industry to promote innovation. Based on the valuable lessons learnt from the biosensor paradigm, the authors argue that strategic industry choices deal primarily with the best stage/point to intersect and seize the university output, implanting the required element of marketability that will transform an idea to a viable application. The authors present a methodological approach for accelerating the knowledge transfer from the university to industry aiming at the effective transition of science to products through a business model reconfiguration.

Flow Injection Monitoring of Aflatoxin M1 in Cheese Using Filter-Supported Bilayer Lipid Membranes with Incorporated DNA

This work describes a technique for the rapid and sensitive electrochemical flow injection monitoring of aflatoxin M1 (AFM1) in cheese samples. Stabilized filter-supported bilayer lipid membranes (BLMs) were used as detectors Single stranded DNA oligomers terminated with alkyl chains (dT20–C16) were incorporated into the membranes to control surface electrostatic properties. The incorporation of dT20–C16 in BLMs lowered the detection limit for the detection of this toxin by one to four orders of magnitude as compared with the detection limit obtained in the absence of DNA. Therefore, it is now possible to continuously monitor this toxin at concentrations that approached those that could be of interest as set by the U.S. Food and Drug Administration and most European countries. The work described herein takes a significant step towards development of a detector of greater practical potential by demonstrating that the incorporation of C16-ssDNA into lipid membranes results in a combination of properties that provides for a much more sensitive and robust detection system. Injections of AFM1 were made into flowing streams of a 0.1 M KCl electrolyte solution, and a transient current signal with duration of seconds reproducibly appeared in about 12 s after exposure of the detector element to the toxin. The magnitude of this signal was linearly related to the concentration of AFM1 with detection limits at subnanomolar level. The effect of interferents such as proteins and lipids was investigated. It was determined that interferences from proteins could be eliminated by adjustment of the flow rate of the carrier electrolyte solution. The technique was applied for the rapid flow injection determination of aflatoxin M1 in cheese samples. AFM1 could be determined in continuous flowing systems with a rate of at least 4 samples min–1. Repetitive cycles of injection of AFM1 have shown no signal degradation during each cycle for experiments that attempted over 30 cycles of detection.

A Knowledge-Based Approach to Online Fault Diagnosis of FET Biosensors

Real-time diagnosis of insulator-semiconductor field-effect transistor (ISFET)-based biosensor systems aims at promptly correcting errors caused by insufficient function; insufficiency is judged by the operational behavior of the sensor, i.e., the data that it produces. Ultimately, a complete failure of the system (i.e., a “dead” sensor) should easily be recognized. Much more difficult is the recognition of a gradual malfunction of this complex system, which may be attributed to faults or failures in one or more of its subsystems. Evidently, the identification of the possible fault modes and their symptoms requires in-depth knowledge of sensors design and operation, both from the biochemical and electrical/electronic points of view, along with tackling uncertain, incomplete, or imprecise information. In this paper, a novel real-time diagnostic expert scheme for field-effect transistor (FET)-based biosensing is proposed. This paper 1) investigates the causes of sensor misfunction by means of fault tree analysis (FTA) relying on fuzzy reasoning to account for uncertainty and 2) proposes a computer-aided method for diagnosing biosensor failure during operation through an algorithmic procedure that is based on a nested loop mechanism. The tree (dendritic) structure (built using the information provided by the biosensor components and their intrarelations/interrelations on a surface- and a deep-knowledge level) serves as the knowledge base (KB), and the fuzzy-rules-based decision mechanism is the inference engine for fault detection and isolation.

Creating a specific domain ontology for supporting R&D in the science-based sector - The case of biosensors

In science-based and technology-intensive projects, knowledge management challenges require a tentative and cautious review of the technological domains, as well as, venues to monitor and assess the way those domains evolve, emerge, mature, and decline. Ontologies play a crucial role in conceptualizing/formalizing domain knowledge, yet any ontological platform that is constructed for supporting R&D throughout the knowledge creation process, must explicitly address the interplay between exploitation and exploration of knowledge at deep and surface levels. Focusing on the product per se and its downstream and upstream knowledge evolution complex system, ontology engineering adopts herein a process-driven view for capturing a continuously changing environment. The authors present a methodological framework for creating specific domain ontologies by means of a cybernetic infrastructure built on a modification of the Nonakas SECI process. This rationale is exemplified on biosensors, a class of devices strongly attached to multidisciplinary basic and applied science, bearing along many levels of input and output knowledge. The proposed ontological representation, expresses and defines a target product as a metamodel. Combined with knowledge about the scientific background of the product, an aspect model at physical concept level is generated from the metamodel and is further converted into a design model. This scheme enables knowledge to be used not only for representation but also for reasoning at functional level. The research logic followed herein does not bring yet another ontology building methodology through a project-management context, but rather contributes to an ontological approach for exploring the diverse knowledge inputs that a product requires through a specific domain-derived and domain-oriented context, which relies on a collaborative model building methodology and a systemic modeling formalism by using 2nd order cybernetics in order to include human intervention.

DNA Biosensor Based on Self-Assembled Bilayer Lipid Membranes for the Detection of Hydrazines

This article reports a new DNA biosensor strategy for detection of chemicals. This strategy is based on electrochemical monitoring of changes of ion current through a lipid membrane with immobilized DNA probes caused by interaction of these modified BLMs with hydrazine compounds. A self-assembled metal supported bilayer lipid membrane (s-BLM) composed of egg phosphatidyl choline was prepared on a silver metal electrode. The oligomers used were single stranded deoxyribonucleic acids: thymidylic acid icosanucleotide terminated with a C-16 alkyl chain to assist incorporation into s-BLMs (dT20-C16), and deoxyadenylic acid icosanucleotide (dA20). These s-BLMs with incorporated DNA interact with hydrazines, and the s-BLMs display an analytically useful tool for ppb detection levels of hydrazine compounds (i.e., hydrazine, methylhydrazine, dimethylhydrazine and phenylhydrazine). This BLM/DNA biosensor offers a highly sensitive, rapid, and portable device for monitoring these environmentally and toxicologically significant compounds.

A methodological combined framework for roadmapping biosensor research: a fault tree analysis approach within a strategic technology evaluation frame

Abstract Biosensor technology began in the 1960s to revolutionize instrumentation and measurement. Despite the glucose sensor market success that revolutionized medical diagnostics, and artificial pancreas promise currently the approval stage, the industry is reluctant to capitalize on other relevant university-produced knowledge and innovation. On the other hand, the scientific literature is extensive and persisting, while the number of university-hosted biosensor groups is growing. Considering the limited marketability of biosensors compared to the available research output, the biosensor field has been used by the present authors as a suitable paradigm for developing a methodological combined framework for “roadmapping” university research output in this discipline. This framework adopts the basic principles of the Analytic Hierarchy Process (AHP), replacing the lower level of technology alternatives with internal barriers (drawbacks, limitations, disadvantages), modeled through fault tree analysis (FTA) relying on fuzzy reasoning to count for uncertainty. The proposed methodology is validated retrospectively using ion selective field effect transistor (ISFET) – based biosensors as a case example, and then implemented prospectively membrane biosensors, putting an emphasis on the manufacturability issues. The analysis performed the trajectory of membrane platforms differently than the available market roadmaps that, considering the vast industrial experience in tailoring and handling crystallic forms, suggest the technology path of biomimetic and synthetic materials. The results presented herein indicate that future trajectories lie along with nanotechnology, and especially nanofabrication and nano-bioinformatics, and focused, more on the science-path, that is, on controlling the natural process of self-assembly and the thermodynamics of bioelement-lipid interaction. This retained the nature-derived sensitivity of the biosensor platform, pointing out the differences between the scope of academic research and the market viewpoint.

Artificial Lipid Membranes: Past, Present, and Future

The multifaceted role of biological membranes prompted early the development of artificial lipid-based models with a primary view of reconstituting the natural functions in vitro so as to study and exploit chemoreception for sensor engineering. Over the years, a fair amount of knowledge on the artificial lipid membranes, as both, suspended or supported lipid films and liposomes, has been disseminated and has helped to diversify and expand initial scopes. Artificial lipid membranes can be constructed by several methods, stabilized by various means, functionalized in a variety of ways, experimented upon intensively, and broadly utilized in sensor development, drug testing, drug discovery or as molecular tools and research probes for elucidating the mechanics and the mechanisms of biological membranes. This paper reviews the state-of-the-art, discusses the diversity of applications, and presents future perspectives. The newly-introduced field of artificial cells further broadens the applicability of artificial membranes in studying the evolution of life.

Stabilized filter-supported bilayer lipid membranes (BLMs) for automated flow monitoring of compounds of clinical, pharmaceutical, environmental and industrial interest

This paper describes the results of analytical applications of electrochemical biosensors based on bilayer lipid membranes (BLMs) for the automated rapid and sensitive flow monitoring of substrates of hydrolytic enzymes, antigens and triazine herbicides. BLMs, composed of mixtures of egg phosphatidylcholine (egg PC) and dipalmitoylphosphatidic acid (DPPA), were supported on ultrafiltration membranes (glass microfibre or polycarbonate filters) which were found to enhance their stability for flow experiments. The proteins (enzymes, antibodies) were incorporated into a floating lipid matrix at an air-electrolyte interface, and then a casting procedure was used to deliver the lipid onto the filter supports for BLM formation. Injections of the analyte were made into flowing streams of the carrier electrolyte solution and a current transient signal was obtained with a magnitude related to the analyte concentration. Substrates of hydrolytic enzyme reactions (acetylcholine, urea and penicillin) could be determined at the micromolar level with a maximum rate of 220 samples/h, whereas antigens (thyroxin) and triazine herbicides (simazine, atrazine and propazine) could be monitored at the nanomolar level in less than 2 min. The time of appearance of the transient response obtained for herbicides was increased to the order of simazine, atrazine and propazine which has permitted analysis of these triazines in mixtures.

Lipid Membrane Nanosensors for Environmental Monitoring: The Art, the Opportunities, and the Challenges

The advent of nanotechnology has brought along new materials, techniques, and concepts, readily adaptable to lipid membrane-based biosensing. The transition from micro-sensors to nano-sensors is neither straightforward nor effortless, yet it leads to devices with superior analytical characteristics: ultra-low detectability, small sample volumes, better capabilities for integration, and more available bioelements and processes. Environmental monitoring remains a complicated field dealing with a large variety of pollutants, several decomposition products, or secondary chemicals produced ad hoc in the short- or medium term, many sub-systems affected variously, and many processes largely unknown. The new generation of lipid membranes, i.e., nanosensors, has the potential for developing monitors with site-specific analytical performance and operational stability, as well as analyte-tailored types of responses. This review presents the state-of-the art, the opportunities for niche applicability, and the challenges that lie ahead.