Metal-enhanced fluorescence: an emerging tool in biotechnology

Over the past 15 years, fluorescence has become the dominant detection/sensing technology in medical diagnostics and biotechnology. Although fluorescence is a highly sensitive technique, where single molecules can readily be detected, there is still a drive for reduced detection limits. The detection of a fluorophore is usually limited by its quantum yield, autofluorescence of the samples and/or the photostability of the fluorophores; however, there has been a recent explosion in the use of metallic nanostructures to favorably modify the spectral properties of fluorophores and to alleviate some of these fluorophore photophysical constraints. The use of fluorophore-metal interactions has been termed radiative decay engineering, metal-enhanced fluorescence or surface-enhanced fluorescence.     

POLYVIEW: a flexible visualization tool for structural and functional annotations of proteins

The POLYVIEW visualization server can be used to generate protein sequence annotations, including secondary structures, relative solvent accessibilities, functional motifs and polymorphic sites. Two-dimensional graphical representations in a customizable format may be generated for both known protein structures and predictions obtained using protein structure prediction servers. POLYVIEW may be used for automated generation of pictures with structural and functional annotations for publications and proteomic on-line resources. AVAILABILITY: http://polyview.cchmc.org.     

Hard X-ray micro(spectro)scopy: a powerful tool for the geomicrobiologists

During the past few decades, the use of electron microscopy approaches – many developed by Terry Beveridge – to probe the physiology of microorganisms has become a mainstay in fields including microbiology, human health, and geomicrobiology. Recent developments of third-generation synchrotron X-ray sources and X-ray-based microscopy approaches for studying microbial systems have proved their utility as complements to the very powerful approaches regularly employed by electron microscopists. In addition, in recent geomicrobiological studies, researchers have begun to take advantage of the strengths of each technique by using the superior spatial resolution of the electron microscope (relative to the X-ray microscope) and the superior elemental sensitivity of the X-ray microscope (relative to the electron microscope), along with the ability of the X-ray microscope to spatially probe the chemical speciation of elements. The benefits of integrating these two nanoprobes for investigating the same microenvironments within a geomicrobial system are far superior to those of independent studies separately employing each probe.     

Small angle X-ray scattering as a complementary tool for high-throughput structural studies

Structural crystallography and nuclear magnetic resonance (NMR) spectroscopy are the predominant techniques for understanding the biological world on a molecular level. Crystallography is constrained by the ability to form a crystal that diffracts well and NMR is constrained to smaller proteins. Although powerful techniques, they leave many soluble, purified structurally uncharacterized protein samples. Small angle X-ray scattering (SAXS) is a solution technique that provides data on the size and multiple conformations of a sample, and can be used to reconstruct a low-resolution molecular envelope of a macromolecule. In this study, SAXS has been used in a high-throughput manner on a subset of 28 proteins, where structural information is available from crystallographic and/or NMR techniques. These crystallographic and NMR structures were used to validate the accuracy of molecular envelopes reconstructed from SAXS data on a statistical level, to compare and highlight complementary structural information that SAXS provides, and to leverage biological information derived by crystallographers and spectroscopists from their structures. All the ab initio molecular envelopes calculated from the SAXS data agree well with the available structural information. SAXS is a powerful albeit low-resolution technique that can provide additional structural information in a high-throughput and complementary manner to improve the functional interpretation of high-resolution structures.     

Advances in visualization of copper in mammalian systems using X-ray fluorescence microscopy

Synchrotron-based X-ray fluorescence microscopy (XFM) has become an important imaging technique to investigate elemental concentrations and distributions in biological specimens. Advances in technology now permit imaging at resolutions rivaling that of electron microscopy, and researchers can now visualize elemental concentrations in subcellular organelles when using appropriate correlative methods. XFM is an especially valuable tool to determine the distribution of endogenous trace metals that are involved in neurodegenerative diseases. Here, we discuss the latest research on the unusual copper (Cu) storage vesicles that were originally identified in mouse brains and the involvement of Cu in Alzheimer's disease. Finally, we provide an outlook of how future improvements to XFM will drive current trace element research forward.     

VISTAL--a new 2D visualization tool of protein 3D structural alignments

We offer a tool, denoted VISTAL, for two-dimensional visualization of protein structural alignments. VISTAL describes aligned structures as a series of matched secondary structure elements, colored according to the three-dimensional distance of their Calpha atoms. AVAILABILITY: VISTAL can be downloaded from http://trantor.bioc.columbia.edu/~kolodny/software.html.     

MotViz: a tool for sequence motif prediction in parallel to structural visualization and analyses

Linking similar proteins structurally is a challenging task that may help in finding the novel members of a protein family. In this respect, identification of conserved sequence can facilitate understanding and classifying the exact role of proteins. However, the exact role of these conserved elements cannot be elucidated without structural and physiochemical information. In this work, we present a novel desktop application MotViz designed for searching and analyzing the conserved sequence segments within protein structure. With MotViz, the user can extract a complete list of sequence motifs from loaded 3D structures, annotate the motifs structurally and analyze their physiochemical properties. The conservation value calculated for an individual motif can be visualized graphically. To check the efficiency, predicted motifs from the data sets of 9 protein families were analyzed and MotViz algorithm was more efficient in comparison to other online motif prediction tools. Furthermore, a database was also integrated for storing, retrieving and performing the detailed functional annotation studies. In summary, MotViz effectively predicts motifs with high sensitivity and simultaneously visualizes them into 3D strucures. Moreover, MotViz is user-friendly with optimized graphical parameters and better processing speed due to the inclusion of a database at the back end. MotViz is available at http://www.fi-pk.com/motviz.html.     

BicOverlapper: a tool for bicluster visualization

Summary: BicOverlapper is a tool to visualize biclusters fromgene-expression matrices in a way that helps to compare biclusteringmethods, to unravel trends and to highlight relevant genes andconditions. A visual approach can complement biological andstatistical analysis and reduce the time spent by specialistsinterpreting the results of biclustering algorithms. The techniqueis based on a force-directed graph where biclusters are representedas flexible overlapped groups of genes and conditions. Availability: The BicOverlapper software and supplementary materialare available at http://vis.usal.es/bicoverlapper Contact: rodri{at}usal.es Associate Editor: John Quackenbush The first two authors should be reported as joint first authors.     

Stem cell-derived exosomes-an emerging tool for myocardial regeneration

Cardiovascular diseases(CVDs) continue to represent the number one cause of death and disability in industrialized countries. The most severe form of CVD is acute myocardial infarction(AMI), a devastating disease associated with high mortality and disability. In a substantial proportion of patients who survive AMI, loss of functional cardiomyocytes as a result of ischaemic injury leads to ventricular failure, resulting in significant alteration to quality of life and increased mortality. Therefore, many attempts have been made in recent years to identify new tools for the regeneration of functional cardiomyocytes. Regenerative therapy currently represents the ultimate goal for restoring the function of damaged myocardium by stimulating the regeneration of the infarcted tissue or by providing cellsthat can generate new myocardial tissue to replace the damaged tissue. Stem cells(SCs) have been proposed as a viable therapy option in these cases. However, despite the great enthusiasm at the beginning of the SC era, justified by promising initial results, this therapy has failed to demonstrate a significant benefit in large clinical trials. One interesting finding of SC studies is that exosomes released by mesenchymal SCs(MSCs) are able to enhance the viability of cardiomyocytes after ischaemia/reperfusion injury, suggesting that the beneficial effects of MSCs in the recovery of functional myocardium could be related to their capacity to secrete exosomes. Ten years ago, it was discovered that exosomes have the unique property of transferring miRNA between cells, acting as miRNA nanocarriers. Therefore, exosomebased therapy has recently been proposed as an emerging tool for cardiac regeneration as an alternative to SC therapy in the post-infarction period. This review aims to discuss the emerging role of exosomes in developing innovative therapies for cardiac regeneration as well as their potential role as candidate biomarkers or for developing new diagnostic tools.     

Comparison of X-ray fluorescence and optical fluorescence measured behind scattering layers as a basis for X-ray fluorescence endoscopy.

Recent developments in the technology of capillary-fiber optics suitable for X-rays in the range of approximately 4-10keV point to the possible realization of endoscopes applicable in X-ray fluorescence analysis. A general problem is the determination of scattering and absorption processes with consideration to tissue optics, X-ray scattering and X-ray absorption in a diagnostic partial volume. Therefore comparative investigations were performed in order to answer these questions. Zinc-oxide nanoparticles configured as single particles and ZnO clusters provided the fluorescence source in cell layers. An artificial scattering material was employed, which closely approximated the tissue optical conditions and the X-ray optical application conditions in possible diagnostic situations. As a result imaging of spatially resolved X-ray contrasts was better than adequate optical fluorescence imaging by approximately one magnitude. Hence a very important precondition for realizing X-ray fluorescence endoscopy is fulfilled.     

BiVisu: software tool for bicluster detection and visualization

BiVisu is an open-source software tool for detecting and visualizing biclusters embedded in a gene expression matrix. Through the use of appropriate coherence relations, BiVisu can detect constant, constant-row, constant-column, additive-related as well as multiplicative-related biclusters. The biclustering results are then visualized under a 2D setting for easy inspection. In particular, parallel coordinate (PC) plots for each bicluster are displayed, from which objective and subjective cluster quality evaluation can be performed. Availability: BiVisu has been developed in Matlab and is available at http://www.eie.polyu.edu.hk/~nflaw/Biclustering/.     

GenomeComp: a visualization tool for microbial genome comparison

We have developed a software tool, GenomeComp, for summarizing, parsing and visualizing the genome sequences comparison results derived from voluminous BLAST textual output. With GenomeComp, the variation between genomes can be easily highlighted, such as repeat regions, insertions, deletions and rearrangements of genomic segments. This software provides a new visualizing tool for microbe comparative genomics.     

Energy-dispersive X-ray fluorescence spectrometry as a tool for zinc, iron and selenium analysis in whole grain wheat

Background and aims

Crop biofortification programs require fast, accurate and inexpensive methods of identifying nutrient dense genotypes. This study investigated energy-dispersive X-ray fluorescence spectrometry (EDXRF) for the measurement of zinc (Zn), iron (Fe) and selenium (Se) concentrations in whole grain wheat.

Methods

Grain samples were obtained from existing biofortification programs. Reference Zn, Fe and Se concentrations were obtained using inductively coupled plasma optical emission spectrometry (ICP-OES) and/or inductively coupled plasma mass spectrometry (ICP-MS). One set of 25 samples was used to calibrate for Zn (19–60?mg?kg^–1) and Fe (26–41?mg?kg^–1), with 25 further samples used to calibrate for Se (2–31?mg?kg^–1 ). Calibrations were validated using an additional 40–50 wheat samples.

Results

EDXRF limits of quantification (LOQ) were estimated as 7, 3 and 2?mg?kg^–1 for Zn, Fe, and Se, respectively. EDXRF results were highly correlated with ICP-OES or -MS values. Standard errors of EDXRF predictions were ±2.2?mg Zn kg^–1, ±2.6?mg Fe kg^–1, and ±1.5?mg Se kg^–1.

Conclusion

EDXRF offers a fast and economical method for the assessment of Zn, Fe and Se concentration in wheat biofortification programs.     

Fluorescence complementation: an emerging tool for biological research

Numerous technologies based on utilizing fluorescent proteins have been developed for biological research, and fluorescence complementation (FC) is a recent application for visualization of molecular events in living cells and organisms. Currently, ten fluorescent proteins have been demonstrated to support FC. Over the past five years, FC-based technologies have been developed to visualize a variety of molecular events, such as protein-protein interactions, post-translational modifications, protein folding, conformational changes, RNA-protein interactions, mRNA localization and DNA hybridization. In addition, FC has also been used for drug discovery. These applications are providing fascinating insights into many biological processes. Here, we review the principles and applications of FC technologies, discuss their current challenges and examine prospects for future advances.     

methGraph: A genome visualization tool for PCR-based methylation assays

Abnormalities in DNA methylation of CpG islands that play a role in gene regulation affect gene expression and hence play a role in disease, including cancer. Bisulfite-based DNA methylation analysis methods such as methylation-specific PCR (MSP) and bisulfite sequencing (BiSeq) are most commonly used to study gene-specific DNA methylation. Assessing specificity and visualizing the position of PCR primers in their genomic context is a laborious and tedious task, primarily due to the sequence changes induced during the bisulfite conversion. For this purpose, we developed methGraph, a web application for easy, fast and flexible visualization and accurate in silico quality evaluation of PCR-based methylation assays. The visualization process starts by submitting PCR primer sequences for specificity assessment and mapping on the genome using the BiSearch ePCR primer-search algorithm. The next step comprises the selection of relevant UCSC genome annotation tracks for display in the final graph. A custom track showing all individual CpG dinucleotides, representing their distribution in the CpG island is also provided. Finally, methGraph creates a BED file that is automatically uploaded to the UCSC genome browser, after which the resulting image files are extracted and made available for visualization and download. The generated high-quality figures can easily be customized and exported for use in publications or presentations. methGraph is available at http://mellfire.ugent.be/methgraph/.