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Chondrocytes derived from mesenchymal stromal cells and induced pluripotent cells of patients with familial osteochondritis dissecans exhibit an endoplasmic reticulum stress response and defective matrix assembly

Familial osteochondritis dissecans (FOCD) is an inherited skeletal defect characterized by the development of large cartilage lesions in multiple joints, short stature, and early onset of severe osteoarthritis. It is associated with a heterozygous mutation in the ACAN gene, resulting in a Val-Met replacement in the C-type lectin domain of aggrecan. To understand the cellular pathogenesis of this c

Elucidating the Molecular Composition of Cartilage by Proteomics

Articular cartilage consists of chondrocytes and two major components, a collagen-rich framework and highly abundant proteoglycans. Most prior studies defining the zonal distribution of cartilage have extracted proteins with guanidine-HCl. However, an unextracted collagen-rich residual is left after extraction. In addition, the high abundance of anionic polysaccharide molecules extracted from cart

Selective extraction of small proteins from biological samples using a novel restricted access column with cation exchange properties

The determination of proteins utilising a polymer-based restricted access support material with ion exchange properties (IERAM) is outlined. Solid phase extraction coupled on-line with a microbore reversed phase HPLC system for the quantitation of small marker proteins is demonstrated. The cation-exchange restricted access packings were characterised with respect to their adsorption and desorption

Development of fluorescence based flow immunoassays utilising restricted access columns

The development of high-speed flow immunoassay techniques is described. The principles are based on heterogeneous flow immunoassay interactions. High sample throughput can be used for screening small analytes in a number of biological matrices originating from samples of water from environmentally polluted areas, or biological fluids such as urine and plasma. The immunochemical detection principle

Integrated microanalytical technology enabling rapid and automated protein identification

Protein identification through peptide mass mapping by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has become a standard technique, used in many laboratories around the world. The traditional methodology often includes long incubations (6-24 h) and extensive manual steps. In an effort to address this, an integrated microanalytical platform has been d

Protein identification platform utilizing micro dispensing technology interfaced to matrix-assisted laser desorption ionization time-of-flight mass spectrometry

An integrated protein microcharacterization/identification platform has been developed. The system has been designed to allow a high flexibility in order to tackle challenging analytical problems. The platform comprises a cooled microautosampler, an integrated system for microcolumn HPLC, and a capillary reversed-phase column that is interfaced to matrix-assisted laser desorption ionization time-o

Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the ne

Picoliter sample preparation in MALDI-TOF MS using a micromachined silicon flow-through dispenser

This paper presents a picoliter sample preparation technique utilizing the flow-through principle, allowing on-line coupling of chromatographic systems to be made. The work was performed in order to investigate the characteristics and the physicochemical properties of the sample preparation using typical mobile phase conditions from μ-CLC (column liquid chromatography) separations. The device pres

Fluorescence polarisation for immunoreagent characterisation

Antibodies were characterised using fluorescence polarisation, a homogeneous assay technique in which all reagents are in solution. Kinetic studies on the association and dissociation of the immunocomplex were performed. A competitive assay was used and the sensitivities, operational linearities, as well as the specificities of the immunoassays were experimentally determined for various antibody p

A flow immunoassay for studies of human exposure and toxicity in biological samples

This paper describes a heterogeneous competitive flow immunoassay with a high sample throughput which can be used for the screening of smaller analytes in various samples. The method is based on off-line incubation of the analyte (Ag), a fluorescent labelled tracer (Ag*) and the corresponding antibody (Ab). The separation of bound (Ab-Ag*) and free tracer (Ag*) is based on a size exclusion and rev

High sample throughput flow immunoassay utilising restricted access columns for the separation of bound and free label

A flow immunodetection system with high sample throughput capacity is described for the screening of various analytes. The immunochemical detection principle is based on the chromatographic separation of the formed immunocomplex (AbAg or AbAg*) and the free antigen (Ag) by a restricted access (RA) column, utilising size-exclusion and reversed-phase mechanism. A fluorescein labelled analyte (Ag*) w

Performance of two immunoassays for the determination of atrazine in sea water samples as compared with on-line solid phase extraction-liquid chromatography-diode array detection

Two immunoassay formats, magnetic particles-based assay (Atrazine RaPID assay and Atrazine High-Sensitivity RaPID assay) and microtiter plate based assay (Department of Entomology and Environmental Toxicology, University of California in Davis) were evaluated for the determination of atrazine in sea water samples. The results obtained were compared and validated with those obtained by using on-lin

On-line solid-phase extraction in liquid chromatography using restricted access pre-columns for the analysis of s-triazines in humic-containing waters

A coupled-column liquid chromatographic system using solid phase extraction pre-columns for the seven chlorotriazines, atrazine, desethylatrazine, desisopropylatrazine (DIA), propazine, simazine, desethylterbuthylazine and terbuthylazine, using photodiode-array detection (PDAD) was developed. Restricted access (RA), was investigated as a pre- column material, making use of non-adsorptive size excl

Tyrosinase graphite-epoxy based composite electrodes for detection of phenols

The characterization and analytical performance of a tyrosinase graphite-epoxy electrode for the detection of phenolic compounds are described. The biocomposite configuration is based on the entrapment of commercially available tyrosinase in a graphite-epoxy matrix, and the mixing of the resulting conductive epoxy resin with a hardener. The enzyme electrode is mounted as a working electrode in an

Genetic signals of high-altitude adaptation in amphibians : A comparative transcriptome analysis

Background: High-altitude adaptation provides an excellent system for studying how organisms cope with multiple environmental stressors and interacting genetic modifications. To explore the genetic basis of high-altitude adaptation in poikilothermic animals, we acquired transcriptome sequences from a high-altitude population and a low-altitude population of the Asiatic toad (Bufo gargarizans). Tra

Infrastructures as analytical framework for mapping research evaluation landscapes and practices

This paper suggests an infrastructure perspective, as suggested by Star and Bowker (2006), as an analytical framework for studying the research evaluation landscape. An infrastructure is suggested to be understood, not as a concrete technology, but as a system of contextual factors including ‘Actors/Stakeholders’, ‘Technical systems’, and ‘Evaluation practices’. How the framework can be operationa

Infrastructures as an analytical framework for understanding research evaluation systems

When trying to map the bibliometrics based ‘research evaluation landscape’ in terms of for instance methods, technical systems and stakeholders; the complexities soon become obvious. Different methods are used at different national and local levels; there is a great variation of systems used for gathering, organizing, and analyzing data; and a multitude of stakeholders, many of which taking on a v

Historical change and drivers of insect pest abundances in red clover seed production

Plant-feeding insects reduce yields substantially in all major crops despite considerable crop protection efforts. A lack of long-term monitoring data and insufficient ecological knowledge of pest insects limit our possibilities to characterize drivers of change in their abundances. We demonstrate how a suite of complementary methods and data sources can be used to analyse historical shifts in pes

Scientific publications as boundary objects: theorising the intersection of classification and research evaluation

Introduction. When using bibliometrics for research evaluation, the classification of research fields is an issue of great importance. The purpose of this paper is to outline a brief theoretical framework for analysing the role of classification in research evaluation practices.Theory. Taking departure in the concept of ‘boundary objects’ we develop a theoretical framework for analyses of how scie