Point-of-Care Testing for Infectious Diseases: Past, Present, and Future.

Point-of-care (POC) diagnostics provide rapid actionable information for patient care at the time and site of an encounter with the health care system. The usual platform has been the lateral flow immunoassay. Recently, emerging molecular diagnostics have met requirements for speed, low cost, and ease of use for POC applications. A major driver for POC development is the ability to diagnose infectious diseases at sites with a limited infrastructure. The potential use in both wealthy and resource-limited settings has fueled an intense effort to build on existing technologies and to generate new technologies for the diagnosis of a broad spectrum of infectious diseases.

[Progresses in screening active compounds from herbal medicine by affinity chromatography].

Affinity chromatography is a chromatographic method for separating molecules using the binding characteristics of the stationary phase with potential drug molecules. This method can be performed as a high throughput screening method and a chromatographic separation method to screen a variety of active drugs. This paper summarizes the history of affinity chromatography, screening technology of affinity chromatography, and application of affinity chromatography in screening bio-active compounds in herbal medicines, and then discusses its application prospects, in order to broaden applications of the affinity chromatography in drug screening.

New trends in fluorescence immunochromatography.

During the last decade, immunochromatographic techniques, in particular fluorescence labeled immunochromatography, have become more popular for the determination of low concentrations of analytes. It offers several potential advantages including highly sensitive detection and wide applications in clinical chemistry, bioanalysis, and environmental analysis. Currently, fluorescence labeled immunochromatography is not the exclusive preserve for a few specialists who are well experienced with antigen-antibody analysis, since the recent developments in fluorescence labeled immunochromatography have become widespread, from simple screening application to quantitative analysis of analytes. In this review, we assembled recent advances in the development and applications of fluorescence labeled immunochromatography techniques, considering their potential benefits in the future.

Techniques for phosphopeptide enrichment prior to analysis by mass spectrometry.

Mass spectrometry is the tool of choice to investigate protein phosphorylation, which plays a vital role in cell regulation and diseases such as cancer. However, low abundances of phosphopeptides and low degrees of phosphorylation typically necessitate isolation and concentration of phosphopeptides prior to MS analysis. This review discusses the enrichment of phosphopeptides with immobilized metal affinity chromatography, reversible covalent binding, and metal oxide affinity chromatography. Capture of phosphopeptides on TiO(2) seems especially promising in terms of selectivity and recovery, but the success of all methods depends on careful selection of binding, washing, and elution solutions. Enrichment techniques are complementary, such that a combination of methods greatly enhances the number of phosphopeptides isolated from complex samples. Development of a standard series of phosphopeptides in a highly complex mixture of digested proteins would greatly aid the comparison of different enrichment methods. Phosphopeptide binding to magnetic beads and on-plate isolation prior to MALDI-MS are emerging as convenient methods for purification of small (microL) samples. On-plate enrichment can yield >70% recoveries of phosphopeptides in mixtures of a few digested proteins and can avoid sample-handling steps, but this technique is likely limited to relatively simple samples such as immunoprecipitates. With recent advances in enrichment techniques in hand, MS analysis should provide important insights into phosphorylation pathways.

Recent Progress in the Methodologies to Identify Physiological Ligands of Siglecs.

Siglecs, a family of receptor-like lectins, recognize glycoproteins and/or glycolipids containing sialic acid in the extracellular space and transduce intracellular signaling. Recently, researchers uncovered significant contributions of Siglecs in cancer immunity, renewing interest in this family of proteins. Previous extensive studies have defined how Siglecs recognize glycan epitopes (glycotopes). Nevertheless, the biological role of these glycotopes has not been fully evaluated. Recent studies using live cells have begun unraveling the constituents of Siglec ligands. These studies demonstrated that glycoprotein scaffolds (counter-receptors) displaying glycotopes are sometimes just as important as the glycotope itself. These new insights may guide future efforts to develop therapeutic agents to target the Siglec - ligand axis.

Affinity chromatography: A review of trends and developments over the past 50 years.

The field of affinity chromatography, which employs a biologically-related agent as the stationary phase, has seen significant growth since the modern era of this method began in 1968. This review examines the major developments and trends that have occurred in this technique over the past five decades. The basic principles and history of this area are first discussed. This is followed by an overview of the various supports, immobilization strategies, and types of binding agents that have been used in this field. The general types of applications and fields of use that have appeared for affinity chromatography are also considered. A survey of the literature is used to identify major trends in these topics and important areas of use for affinity chromatography in the separation, analysis, or characterization of chemicals and biochemicals.

Affinity chromatography approaches to overcome the challenges of purifying plasmid DNA.

The diversity of biomolecules present in plasmid DNA (pDNA)-containing extracts and the structural and chemical similarities between pDNA and impurities are some of the main challenges of improving or establishing novel purification procedures. In view of the unequalled specificity of affinity purification, this technique has recently begun to be applied in downstream processing of plasmids. This paper discusses the progress and importance of affinity chromatography (AC) for the purification of pDNA-based therapeutic products. Several affinity approaches have already been successfully developed for a variety of applications, and we will focus here on highlighting their possible contributions to the pDNA purification challenge. Diverse affinity applications and their advantages and disadvantages are discussed, as well as the most significant results and improvements in the challenging task of purifying plasmids.

Affinity as a tool in life science.

The use of affinity-based tools has become invaluable as a platform for basic research and in the development of drugs and diagnostics. Applications include affinity chromatography and affinity tag fusions for efficient purification of proteins as well as methods to probe the protein network interactions on a whole-proteome level. A variety of selection systems has been described for in vitro evolution of affinity reagents using combinatorial libraries, which make it possible to create high-affinity reagents to virtually all biomolecules, as exemplified by generation of therapeutic antibodies and new protein scaffold binders. The strategies for high-throughput generation of affinity reagents have also opened up the possibility of generating specific protein probes on a whole-proteome level. Recently, such affinity proteomics have allowed the detailed analysis of human protein expression in a comprehensive manner both in normal and disease tissue using tissue microarrays and confocal microscopy.

The social network of a cell: recent advances in interactome mapping.

Proteins very rarely act in isolation. Biomolecular interactions are central to all biological functions. In human, for example, interference with biomolecular networks often lead to disease. Protein-protein and protein-metabolite interactions have traditionally been studied one by one. Recently, significant progresses have been made in adapting suitable tools for the global analysis of biomolecular interactions. Here we review this suite of powerful technologies that enable an exponentially growing number of large-scale interaction datasets. These new technologies have already contributed to a more comprehensive cartography of several pathways relevant to human pathologies, offering a broader choice for therapeutic targets. Genome-wide scale analyses in model organisms reveal general organizational principles of eukaryotic proteomes. We also review the biochemical approaches that have been used in the past on a smaller scale for the quantification of the binding constant and the thermodynamics parameters governing biomolecular interaction. The adaptation of these technologies to the large-scale measurement of biomolecular interactions in (semi-)quantitative terms represents an important challenge.

Affinity-based methodologies and ligands for antibody purification: advances and perspectives.

Many successful, recent therapies for life-threatening diseases such as cancer and rheumatoid arthritis are based on the recognition between native or genetically engineered antibodies and cell-surface receptors. Although naturally produced by the immune system, the need for antibodies with unique specificities and designed for single application, has encouraged the search for novel antibody purification strategies. The availability of these products to the end-consumer is strictly related to manufacture costs, particularly those attributed to downstream processing. Over the last decades, academia and industry have developed different types of interactions and separation techniques for antibody purification, affinity-based strategies being the most common and efficient methodologies. The affinity ligands utilized range from biological to synthetic designed molecules with enhanced resistance and stability. Despite the successes achieved, the purification "paradigm" still moves interests and efforts in the continuous demand for improved separation performances. This review will focus on recent advances and perspectives in antibody purification by affinity interactions using different techniques, with particular emphasis on affinity chromatography.

Future of antibody purification.

Antibody purification seems to be safely ensconced in a platform, now well-established by way of multiple commercialized antibody processes. However, natural evolution compels us to peer into the future. This is driven not only by a large, projected increase in the number of antibody therapies, but also by dramatic improvements in upstream productivity, and process economics. Although disruptive technologies have yet escaped downstream processes, evolution of the so-called platform is already evident in antibody processes in late-stage development. Here we perform a wide survey of technologies that are competing to be part of that platform, and provide our [inherently dangerous] assessment of those that have the most promise.

Diagnosis of opportunistic infections: HIV co-infections - tuberculosis.

PURPOSE OF REVIEW: Tuberculosis (TB) incidence has declined ∼1.5% annually since 2000, but continued to affect 10.4 million individuals in 2015, with 1/3 remaining undiagnosed or underreported. The diagnosis of TB among those co-infected with HIV is challenging as TB remains the leading cause of death in such individuals. Accurate and rapid diagnosis of active TB will avert mortality in both adults and children, reduce transmission, and assist in timeous decisions for antiretroviral therapy initiation. This review describes advances in diagnosing TB, especially among HIV co-infected individuals, highlights national program's uptake, and impact on patient care. RECENT FINDINGS: The TB diagnostic landscape has been transformed over the last 5 years. Molecular diagnostics such as Xpert MTB/RIF, which simultaneously detects Mycobacterium tuberculosis (MTB) resistance to rifampicin, has revolutionized TB control programs. WHO endorsed the use of Xpert MTB/RIF in 2010 for use in HIV/TB co-infected patients, and later in 2013 for use as the initial diagnostic test for all adults and children with signs and symptoms of pulmonary TB. Line probe assays (LPAs) are recommended for the detection of rifampicin and isoniazid resistance in sputum smear-positive specimens and mycobacterial cultures. A second-line line probe assay has been recommended for the diagnosis of extensively drug-resistant (XDR)-TB Assays such as the urine lateral flow (LF)-lipoarabinomannan (LAM), can be used at the point of care (POC) and have a niche role to supplement the diagnosis of TB in seriously ill HIV-infected, hospitalized patients with low CD4 cell counts of less than 100 cells/µl. Polyvalent platforms such as the m2000 (Abbott Molecular) and GeneXpert (Cepheid) offer potential for integration of HIV and TB testing services. While the Research and Development (R&D) pipeline appears to be rich at first glance, there are actually few leads for true POC tests that would allow for earlier TB diagnosis or rapid, comprehensive drug susceptibility testing, especially when considering the very high attrition rates observed between biomarker discovery and product market entry. SUMMARY: In this review, we describe diagnostic strategies specifically for HIV and TB co-infected individuals. Molecular diagnostics in particular within the past 5 years have revolutionized and 'disrupted' this field. They lend themselves to integration of services with platforms capable of polyvalent testing. Impact on patient care is, however, still debatable. What has been highlighted is the need for health system strengthening and for true POC testing that can be used in active case finding.

New concepts in the chromatography of peptides and proteins.

Although chromatography using a variety of novel bed configurations (e.g. fluidized beds, expanded beds, simulated moving beds, annular rotating beds, etc.) has been of recent interest, the majority of practical applications of analytical and preparative chromatography employ a stationary adsorbent bed into which a feed slug is charged periodically, similar to the technique first described by Mikhail Tswett over 100 years ago. However, new concepts in both the practice and theory of fixed-bed chromatography are continuing to expand the available range of applications for separating peptides and proteins.

Advances in gentle immunoaffinity chromatography.

Immunoaffinity chromatography is one of the most powerful fractionation steps available for protein purification; however, it is often difficult to elute bound protein without using harsh or denaturing elution conditions. The development of methods to identify monoclonal antibodies that bind antigens tightly, but release under gentle, non-denaturing conditions has made possible the immunoaffinity purification of labile, multisubunit enzyme complexes with high yield and high specific activity. This work has implications for emerging proteomic applications, allowing identification of new protein-protein interaction partners, retention of biological activity and the isolation of protein complexes more amenable to crystallization and structure determination.

Two-dimensional gel electrophoresis; better than a poke in the ICAT?

To date, the most widely used technology for conducting proteomic studies has been two-dimensional gel electrophoresis (2DGE), but this approach does have drawbacks. Isotope-coded affinity tagging (ICAT) is starting to challenge 2DGE as a new proteomic tool for the analysis of proteins in complex biological specimens. An appraisal of these two methodologies reveals that neither ICAT nor 2DGE provide comprehensive coverage on a proteome-wide scale.

Analytical methods for kinetic studies of biological interactions: A review.

The rates at which biological interactions occur can provide important information concerning the mechanism and behavior of these processes in living systems. This review discusses several analytical methods that can be used to examine the kinetics of biological interactions. These techniques include common or traditional methods such as stopped-flow analysis and surface plasmon resonance spectroscopy, as well as alternative methods based on affinity chromatography and capillary electrophoresis. The general principles and theory behind these approaches are examined, and it is shown how each technique can be utilized to provide information on the kinetics of biological interactions. Examples of applications are also given for each method. In addition, a discussion is provided on the relative advantages or potential limitations of each technique regarding its use in kinetic studies.

Current trends in molecular recognition and bioseparation.

Molecular recognition guides the selective interaction of macromolecules with each other in essentially all biological processes. Perhaps the most impactful use of biomolecular recognition in separation science has been in affinity chromatography. The results of the last 26 years, since Cuatrecases, Wilchek and Anfinsen first reported the purification of staphylococcal nuclease, have validated the power of biomolecular specificity for purification. This power has stimulated an explosion of solid-phase ligand designs and affinity chromatographic applications. An ongoing case in point is the purification of recombinant proteins, which has been aided by engineering the proteins to contain Affinity-Tag sequences, such as hexa-histidine for metal-chelate separation and epitope sequence for separation by an immobilized monoclonal antibody. Tag technology can be adapted for plate assays and other solid-phase techniques. The advance of affinity chromatography also has stimulated immobilized ligand-based methods to characterize macromolecular recognition, including both chromatographic and optical biosensor methods. And, new methods such as phage display and other diversity library approaches continue to emerge to identify new recognition molecules of potential use as affinity ligands. Overall, it is tantalizing to envision a continued evolution of new affinity technologies which use the selectivity built into biomolecular recognition as a vehicle for purification, analysis, screening and other applications in separation sciences.

New developments in affinity chromatography with potential application in the production of biopharmaceuticals.

Affinity chromatography is likely to play an increasingly important role in the purification of pharmaceutical proteins. This review describes new approaches to the design and synthesis of affinity ligands based on the ability to combine knowledge of X-ray crystallographic or NMR structures with defined or combinatorial chemical synthesis. The de novo design process is based on peptidal templates, complementarity to surface exposed residues and mimicking natural biological recognition. Examples of ligands designed to bind specifically to kallikrein, elastase, immunoglobulin G, insulin, alpha(1)-antitrypsin, clotting factor VII and glyco-proteins are given. The exceptional selectivity and stability of these synthetic ligands allows their use in harsh manufacturing environments.

Current trends in separation of plasmid DNA vaccines: a review.

Plasmid DNA (pDNA)-based vaccines offer more rapid avenues for development and production if compared to those of conventional virus-based vaccines. They do not rely on time- or labour-intensive cell culture processes and allow greater flexibility in shipping and storage. Stimulating antibodies and cell-mediated components of the immune system are considered as some of the major advantages associated with the use of pDNA vaccines. This review summarizes the current trends in the purification of pDNA vaccines for practical and analytical applications. Special attention is paid to chromatographic techniques aimed at reducing the steps of final purification, post primary isolation and intermediate recovery, in order to reduce the number of steps necessary to reach a purified end product from the crude plasmid.