Methods Collections
Protein-Protein Interaction Methods
Guest Editors: Alastair Barr, Michael Overduin
The study of protein-protein interactions is key to understanding the network of molecular interactions underlying biological effects, and defining how individual proteins perform their functional role within a cell. Detailed understanding of t… Show Morehese interactions often provides insight into aberrant signaling in disease. This collection brings together the wide range of techniques that have been developed to study interactions using in vitro, cell-based and in vivo approaches. Each technique has its own strengths, for example enabling systematic interrogation of the complete ‘interactome’, providing detailed atomic level information about a single interaction or insight into the sub-cellular localization of interacting proteins. Some techniques are easily adaptable to high-throughput screening of libraries or pulling out novel interacting partners while others have a more limited capacity. Readouts such as microscopy and mass-spectrometry are employed in protein-protein interaction techniques and some provide quantitative information on the affinity of an interaction.
Examples of methods to be included in this collection are: co-immunoprecipitation and antibody interference; pull-down assays using fusion proteins or modular protein domains; proximity-dependent labelling techniques (e.g. BioID); far western and receptor affinity probes; affinity purification coupled with mass-spectrometry; two-hybrid technologies (yeast and mammalian); fluorescence microscopy (e.g. immunofluoresence, FRET, super-resolution microscopy); protein complementation assays (e.g. BiFC); biophysical approaches (X-ray crystallography, NMR, AUC, SPR) and microarray based techniques (i.e. tissue and protein arrays). Show Less
Examples of methods to be included in this collection are: co-immunoprecipitation and antibody interference; pull-down assays using fusion proteins or modular protein domains; proximity-dependent labelling techniques (e.g. BioID); far western and receptor affinity probes; affinity purification coupled with mass-spectrometry; two-hybrid technologies (yeast and mammalian); fluorescence microscopy (e.g. immunofluoresence, FRET, super-resolution microscopy); protein complementation assays (e.g. BiFC); biophysical approaches (X-ray crystallography, NMR, AUC, SPR) and microarray based techniques (i.e. tissue and protein arrays). Show Less
Current Research Methods in Rabies Diagnosis, Prevention, Treatment, and Control
Guest Editor: Charles E. Rupprecht, VMD MS PhD
Rabies is an acute progressive encephalitis caused by a lyssavirus. The case fatality of this zoonosis is the highest for any infectious disease. Although laboratory techniques on rabies have progressed greatly over the last century, typically such i… Show Morenformation falls in disparate locations and is often unavailable for easy access by those with the greatest need. Hence, the opportunity for a visual, methodological catalog is critical for basic research in pathobiology, immunology and molecular biology, as well for applied diagnostics, prophylaxis, therapy and intervention. The time is opportune because a global program is underway for the elimination of human rabies transmitted by dogs through mass canine vaccination, requiring multiple tests for risk mitigation and measurement of programmatic success. Advances have occurred in pathogen discovery, characterization, novel biologics and anti-virals. Based on in vitro and animal models, hope has arisen not only for immunization after viral exposure but for potential treatment after illness. Besides direct application to humans and domestic animals, efficacious vaccines may also be distributed to free-ranging wildlife en masse via vaccine-laden baits. The objective of this collection is to: share standardized protocols for detection, characterization and response; promote broader global access to existing methods; produce open networks for research collaborations anew; and ultimately create a pathway for an evidence-based approach for additional knowledge on this neglected disease. We trust this approach will encourage a myriad of scientists to participate in this global endeavor in a trans-disciplinary manner, towards renewed biomedical progress in human and veterinary science and conservation biology. Show Less
Comprehensive Guide to Inflammation Detection Methodologies
Guest Editor: Payel Sil
Inflammation is the natural response of the body to harmful stimuli and inflammatory symptoms are routinely used to diagnose ailments. However, acute and chronic inflammation display differing characteristics, which requires deeper exploration for ac… Show Morecurate identification. The tools that are currently available are often lacking in this aspect. With the rapid advancement in the field of immunotherapies, there has been a surge in interest in developing effective methodologies to gain a deeper insight into the state of inflammation.
Swelling, redness and pain are the primary indicators of inflammation. Various detection strategies utilize surface markers, cytokine release and migration patterns of cells as inflammation indicators. The effectiveness of these methods in both immune and non-immune cells is limited by the subtleties of cellular functioning. The signaling pathways and associated mechanisms that launch inflammatory responses are hard to correlate with physiological outcomes. Therefore, it is essential to discover as well as document novel techniques to measure and quantify markers of inflammation in a disease context. This methods collection aims to highlight the unique inflammatory characteristics of multiple cell types relevant to different diseases such as cancer, autoimmune disorders and infections. The collection is intended to be a repository of cutting edge methods developed by researchers to encourage knowledge transfer and innovation, promote cross-pollination of ideas between research teams and ultimately enhance the accuracy of disease diagnosis. Show Less
Swelling, redness and pain are the primary indicators of inflammation. Various detection strategies utilize surface markers, cytokine release and migration patterns of cells as inflammation indicators. The effectiveness of these methods in both immune and non-immune cells is limited by the subtleties of cellular functioning. The signaling pathways and associated mechanisms that launch inflammatory responses are hard to correlate with physiological outcomes. Therefore, it is essential to discover as well as document novel techniques to measure and quantify markers of inflammation in a disease context. This methods collection aims to highlight the unique inflammatory characteristics of multiple cell types relevant to different diseases such as cancer, autoimmune disorders and infections. The collection is intended to be a repository of cutting edge methods developed by researchers to encourage knowledge transfer and innovation, promote cross-pollination of ideas between research teams and ultimately enhance the accuracy of disease diagnosis. Show Less
Mass Spectrometry Imaging, From Tissues to Single Cells
Guest Editors: Matthew O'Rourke, Matthew Padula, Greg Sutherland
This collection is focused on the techniques used for preparing samples for the different forms of mass spectrometry imaging (MSI) as well as any novel data processing pipelines. The aim is to demonstrate workflows that are not well known and that ar… Show Moree platform neutral as well as exploring the range of samples that can be analysed and the different types of analytes that can be detected in those samples. By bringing these methods together into a single accessible collection, there will be more connection between members of a currently fragmented scientific community using MSI. Since the community is quite small and spread around out, new developments in the field are slow and new innovations are either not communicated at all or limited to the primary literature (often in highly specialised journals). By creating this collection, we will provide a central repository for the most significant advances in the field thereby eliminating the need for novices in the field to start from scratch. This will speed up development of MSI techniques as a whole and open up lines of communication between novices and experts in the field, thereby creating a more cohesive community. This means that development avenues in areas such as diagnostics will be supported by a community working towards a central goal as opposed to individual teams working in isolation. Show Less
Current methods in Astyanax mexicanus research
Guest Editors: Misty Rose Riddle, Clifford J. Tabin
The Mexican Tetra, Astyanax mexicanus is an excellent model organism to study the developmental and genetic basis of evolution. This species of fish consists of a river-dwelling (surface) population and a number of cave-dwelling po… Show Morepulations that evolved from surface fish trapped in caves. Cavefish have distinct morphology and physiology including: absence of eyes, reduced pigment, increased sensory structures, sensitive smell, reduced circadian rhythm, altered blood glucose regulation, and increased fat accumulation. Surface fish and all cavefish are interfertile allowing for genetic mapping studies that, in combination with a sequenced and annotated genome, have revealed the genetic changes leading to some of the unique cavefish traits. A. mexicanus have been bred in the lab for generations and numerous protocols exist for investigating their biology. The goal of this collection is to demonstrate standardized protocols for breeding, provide broader access to existing methods, and ultimately lower the threshold for researchers interested in working with this species.
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Studying lipid-protein interactions
Guest Editor: Balaji Olety
Peripheral and integral membrane proteins that interact with lipids make up nearly one third of the total proteome. Despite its abundance, the mechanisms of their interactions with lipid moieties are well understood only for a fraction of them. This … Show Moreis in part due to technical challenges associated with performing and analyzing these interactions. This is in part due to the diversity of lipid structures and the challenges in handling them. In addition to this, the lack of a compendium for these methods makes it challenging for a beginner interested in studying proteins that interact with lipids. In order to address these issues, an exclusive collection of methods used to study lipid-protein interactions was conceived. A wide gamut of methods is used to understand protein-lipid interactions. These methods can broadly fall in the realm of cell-biology, biochemistry or biophysics. Each of these techniques have their own strengths and limitations. For example, certain techniques are well suited to derive qualitative information as readout and while others yield quantitative information or both. Examples of techniques that are used to study such interactions include but are not limited to such as Surface Plasmon resonance, Fluorescence microscopy, membrane flotation assays, AFM, isothermal titration calorimetry, lipid-overlay or pulldown assays, lipid arrays, membrane preparations, quantitative methods etc. Show Less
Methods and Techniques for the Development of Drug Delivery Systems
Guest Editor: Wing-Fu Lai
As far as pharmaceutical interventions are concerned, one of the important factors to be considered is drug bioavailability, which can be easily jeopardized by various factors (particularly the physiological barriers and blood clearance) after drug a… Show Moredministration. This has limited the sustainability of the drug effect and has hampered the therapeutic efficacy. Over the years, different drug carriers (e.g., liposomes, polymers, emulsions, hydrogels, and metal nanoparticles) have emerged. Since the turn of the last century, the versatility and tunability of these carriers have been further enhanced by advances in microfabrication technologies. Many of
these technologies have shown tremendous promise in modulating the pharmacokinetic parameters of diverse medicinal entities, ranging from protein drugs to therapeutic genes. As a multidisciplinary undertaking, drug delivery research may involve experimental techniques not only in materials chemistry but also in biomedical sciences. Such a multidisciplinary nature of the discipline may turn out to be one of the challenges to researchers who wish to enter this rapidly advancing field of research. Regarding this fact, this collection describes techniques and methods to synthesize and characterize different materials systems, and to evaluate the performance of these systems as drug carriers. Its objective is to streamline the research process undertaken by beginning researchers, and to increase the consistency and comparability of data collected from different laboratories during the development, characterization and evaluation of drug delivery systems. Show Less
these technologies have shown tremendous promise in modulating the pharmacokinetic parameters of diverse medicinal entities, ranging from protein drugs to therapeutic genes. As a multidisciplinary undertaking, drug delivery research may involve experimental techniques not only in materials chemistry but also in biomedical sciences. Such a multidisciplinary nature of the discipline may turn out to be one of the challenges to researchers who wish to enter this rapidly advancing field of research. Regarding this fact, this collection describes techniques and methods to synthesize and characterize different materials systems, and to evaluate the performance of these systems as drug carriers. Its objective is to streamline the research process undertaken by beginning researchers, and to increase the consistency and comparability of data collected from different laboratories during the development, characterization and evaluation of drug delivery systems. Show Less
Analysis of Plant Acyl Lipids
Guest Editors: Jenny Mortimer, Toshiki Ishikawa
Plant acyl lipids have many different functions which include compartmentalization, energy storage, signaling, drought tolerance and pathogen resistance. Classes of acyl lipids include the sphingolipids, phospholipids, and glycerolipids, and a high d… Show Moreegree of structural variation exists within each class. Because of this, specialized isolation and analysis methods are required. Recently, there has been a renewed interest in understanding and engineering plant lipids for bioenergy and human nutrition purposes. The explosion of synthetic biology and gene editing, as well as ‘omics data, has increased the pace at which this has occurred. However, there is still much to learn. Here, we will assemble a collection of acyl lipid analysis methods which spans from high-tech, high-throughput lipidomics to more traditional lipid biochemistry. We hope that this collection will encourage plant scientists from other disciplines to explore the lipid chemistry of their plants. Show Less
Mechanical Stimulation Techniques
Guest Editor: Pushkar Lele
A significant body of research suggests that extracellular mechanical cues influence important biological processes, including intracellular signaling, genetic regulation, division and motility. Such mechanical effects are prominent in eukaryotes as … Show Morewell as prokaryotes. To probe the mechanisms of mechanical regulation of cellular functions, several techniques have been developed that enable precise application of mechanical stimuli to cells. Broadly, these approaches can be classified into two groups, global-stimulation methods that stimulate the entire cellular structure, and local-stimulation methods that stimulate a part of the cell structure. The former approach enables global, high-throughput measurements of mechano-responses. Analysis of response is typically based on the assumption of homogeneity in whole-cell properties. The latter approach does not assume homogeneity and offers the advantages of high spatio-temporal resolution in force-application and the ability to measure single-organelle response-dynamics. The two approaches together have facilitated fundamental advances in cell mechanics and mechanobiology.
Owing to the similarities between soft matter and biological matter, several techniques that are commonly employed in soft matter research are also useful in probing biological systems. Of interest in this collection are novel applications of popular techniques, which allow local as well as global force-application, to interesting problems in biocolloid manipulation and cell mechanics. These include popular tools such as optical tweezers, magnetic traps, electric fields, micropipette aspiration, bulk and micro-rheology methods. Other novel implementations for mechanical-stimulation of biological systems such as those that rely on hydrodynamic flows, 3D-printing and ultra-sonication are also within the scope of this collection. Show Less
Owing to the similarities between soft matter and biological matter, several techniques that are commonly employed in soft matter research are also useful in probing biological systems. Of interest in this collection are novel applications of popular techniques, which allow local as well as global force-application, to interesting problems in biocolloid manipulation and cell mechanics. These include popular tools such as optical tweezers, magnetic traps, electric fields, micropipette aspiration, bulk and micro-rheology methods. Other novel implementations for mechanical-stimulation of biological systems such as those that rely on hydrodynamic flows, 3D-printing and ultra-sonication are also within the scope of this collection. Show Less
Methods and Techniques for Micro and Electro-Propulsion for Cubesats and Smart Satellites
Guest Editors: Igor Levchenko, Angelo Cervone, Stéphane Mazouffre, Mitchel Walker, Shuyan Xu
Rapid development of information technologies and material science has formed a unique base for the space exploration renaissance. Miniaturization of electronic and energy generating devices, along with novel materials and innovative paradigms in roc… Show Moreket technology, have evoked a dawn of miniaturized spaceship capable of performing all and even more functions intrinsic to oldfashioned large satellites. A task of extensive exploration of a whole Solar system is now faced, and appropriate efforts already undertaken by setting a global satellite-based communication system and sending the scientific orbiters and landers to planets including Venus and Mars.
Like smartphones in our pockets, smart satellites replace the obsolete heavy equipment at orbit. On the other hand, one persistent problem hinders the further progress in modern, smart spaceship technology; namely, miniaturized satellites require miniature but efficient engines, which are a challenge now. Powerful chemical rocket engines are inherently inefficient upon miniaturization; thus, only electric propulsion technique which does not have any physical limitations on the exhaust velocity could be a base for efficient yet small thrusters capable of driving modern, miniaturized, smart satellites between orbits, make them interference with each other and so on.
In this collection we will see various methods and techniques for designing, testing and space application of quite various electric propulsion techniques, with a stress to those suitable for smart satellites. Hall-type thrusters, ion gridded thrusters and several other perspective types will be represented by their creators, thus forming a whole picture describing various types of electric and plasma types that different labs and research groups specialize on. On the other hand, each presentation will describe some specific type of thrusters and thus could be selfconsistent in some extent. Significant attention will be devoted to research groups already tested their systems in space. Show Less
Like smartphones in our pockets, smart satellites replace the obsolete heavy equipment at orbit. On the other hand, one persistent problem hinders the further progress in modern, smart spaceship technology; namely, miniaturized satellites require miniature but efficient engines, which are a challenge now. Powerful chemical rocket engines are inherently inefficient upon miniaturization; thus, only electric propulsion technique which does not have any physical limitations on the exhaust velocity could be a base for efficient yet small thrusters capable of driving modern, miniaturized, smart satellites between orbits, make them interference with each other and so on.
In this collection we will see various methods and techniques for designing, testing and space application of quite various electric propulsion techniques, with a stress to those suitable for smart satellites. Hall-type thrusters, ion gridded thrusters and several other perspective types will be represented by their creators, thus forming a whole picture describing various types of electric and plasma types that different labs and research groups specialize on. On the other hand, each presentation will describe some specific type of thrusters and thus could be selfconsistent in some extent. Significant attention will be devoted to research groups already tested their systems in space. Show Less
Chloroplast research methods: probing the localization, topology, targeting and interactions of chloroplast proteins
Guest Editor: Paul Jarvis
Chloroplasts are the organelles that define plants. Along with many other metabolic, developmental and signalling functions, chloroplasts are responsible for photosynthesis – the process whereby sunlight energy is harnessed to power the cellula… Show Morer activities of life. Consequently, chloroplasts are essential, not only for plants but also for the myriad ecosystems that depend on plants, and for agriculture. Chloroplasts are composed of thousands of different proteins, most of which are nucleus-encoded and imported from the cytosol before being routed internally to one of six, clearly-distinct intraorganellar compartments. To achieve a more complete understanding of chloroplast development and functions, and to enable strategies through which chloroplasts may be manipulated to address global challenges linked to food security or bioenergy, it will be essential to determine the localization, topology, targeting and interactions of important chloroplast proteins. The methods presented here – which focus on the widely-used model plant Arabidopsis thaliana (thale cress), but may also be adapted and applied to other organisms – will be critically important as we pursue these goals. Show Less
Methods for the Assessment of Mitochondrial Bioenergetics and Functions.
Guest Editor: Ryan J. Mailloux, PhD
Mitochondria are the main source of ATP in most, if not all, mammalian cells. However, these dynamic organelles fulfill a myriad of other cell functions including providing antioxidant support and reactive oxygen species to the cell, serving as an im… Show Moreportant site for the biosynthesis of various biomolecules, and have been documented to serve as a signaling platform, regulating a range of cellular functions in response to different physiological cues. As a consequence, mitochondria are integral for modulating a myriad of physiological functions ranging from locomotion to immune cell function and memory. Defects in mitochondrial function have also been linked to a number of human pathologies transforming this double-membraned organelle into a valuable target for gene or pharmacological therapy. It is, therefore, of the utmost importance that cutting-edge tools and techniques are used to understand the relationship between basic mitochondrial functions and their role in physiology and disease. The goal of this collection is to bring together a rich panoply of methods developed by experts to study mitochondrial functions using in vitro, cell-based, and in vivo approaches. Of interest in this collection are the novel protocols used to measure mitochondrial bioenergetics and functions and how findings can be applied to understand the role of defective mitochondria in the pathogenesis of human diseases. Show Less
