Specific label-free and real-time detection of oxidized low density lipoprotein (oxLDL) using an immunosensor with three monoclonal antibodies

Increased levels of plasma oxLDL, which is the oxidized fraction of Low Density Lipoprotein (LDL), are associated with atherosclerosis, an inflammatory disease, and the subsequent development of severe cardiovascular diseases that are today a major cause of death in modern countries. It is therefore important to find a reliable and fast assay to determine oxLDL in serum. A new immunosensor employing three monoclonal antibodies (mAbs) against oxLDL is proposed in this work as a quick and effective way to monitor oxLDL. The oxLDL was first employed to produce anti-oxLDL monoclonal antibodies by hybridoma cells that were previously obtained. The immunosensor was set-up by selfassembling cysteamine (Cyst) on a gold (Au) layer (4 mm diameter) of a disposable screen-printed electrode. Three mAbs were allowed to react with N-hydroxysuccinimide (NHS) and ethyl(dimethylaminopropyl)carbodiimide (EDAC), and subsequently incubated in the Au/Cys. Albumin from bovine serum (BSA) was immobilized further to ensure that other molecules apart from oxLDL could not bind to the electrode surface. All steps were followed by various characterization techniques such as electrochemical impedance spectroscopy (EIS) and square wave voltammetry (SWV). The analytical operation of the immunosensor was obtained by incubating the sensing layer of the device in oxLDL for 15 minutes, prior to EIS and SWV. This was done by using standard oxLDL solutions prepared in foetal calf serum, in order to simulate patient's plasma with circulating oxLDL. A sensitive response was observed from 0.5 to 18.0 mg mL 1 . The device was successfully applied to determine the oxLDL fraction in real serum, without prior dilution or necessary chemical treatment. The use of multiple monoclonal antibodies on a biosensing platform seemed to be a successful approach to produce a specific response towards a complex multi-analyte target, correlating well with the level of oxLDL within atherosclerosis disease, in a simple, fast and cheap way

Economic performance or electoral necessity? Evaluating the system of voluntary income to political parties

Whilst the public funding of political parties is the norm in western democracies, its comprehensive introduction has been resisted in Britain. Political and electoral arrangements in Britain require parties to function and campaign on a regular basis, whilst their income follows cycles largely related to general elections. This article shows that the best predictor of party income is the necessity of a well-funded general election campaign rather than party performance. As a result, income can only be controlled by parties to a limited degree, which jeopardises their ability to determine their own financial position and fulfil their functions as political parties

The Effect of Enzymatically Polymerised Polyphenols on CD4 Binding and Cytokine Production in Murine Splenocytes

High-molecular weight polymerised polyphenols have been shown to exhibit anti-influenza virus, anti-HIV, and anti-cancer activities. The purpose of this study was to evaluate the immunomodulating activities of enzymatically polymerised polyphenols, and to clarify the underlying mechanisms of their effects. The cytokine-inducing activity of the enzymatically polymerised polyphenols derived from caffeic acid (CA), ferulic acid (FA), and p-coumaric acid (CoA) was investigated using murine splenocytes. Polymerised polyphenols, but not non-polymerised polyphenols, induced cytokine synthesis in murine splenocytes. Polymerised polyphenols induced several cytokines in murine splenocytes, with interferon-γ (IFN-γ) and granulocyte-macrophage colony-stimulating factor (GM-CSF) being the most prominent. The underlying mechanisms of the effects of the polymerised polyphenols were then studied using neutralising antibodies and fluorescent-activated cell sorting (FACS) analysis. Our results show that polymerised polyphenols increased IFN-γ and GM-CSF production in splenocytes. In addition, the anti-CD4 neutralised monoclonal antibody (mAb) inhibited polymerised polyphenol-induced IFN-γ and GM-CSF secretion. Moreover, polymerised polyphenols bound directly to a recombinant CD4 protein, and FACS analysis confirmed that interaction occurs between polymerised polyphenols and CD4 molecules expressed on the cell surface. In this study, we clearly demonstrated that enzymatic polymerisation confers immunoactivating potential to phenylpropanoic acids, and CD4 plays a key role in their cytokine-inducing activity

Leishmania infantum heat shock protein 83 for the serodiagnosis of tegumentary leishmaniasis

Fil: Celeste, B. J. Instituto de Medicina Tropical de São Paulo. Laboratório de Soroepidemiologia e Imunobiologia. Departamentos de Dermatologia; Brasil.Fil: Angel, Sergio O. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Departamento de Parasitología; Argentina.Fil: Castro, G. M. Instituto de Medicina Tropical de São Paulo. Laboratório de Soroepidemiologia e Imunobiologia. Departamentos de Dermatologia; Brasil.Fil: Gidlund, M. Universidade de São Paulo. Instituto de Ciências Biomédicas. Departamento de Imunologia, Laboratório de Imunofisiologia; Brasil.Fil: Goto, H. Instituto de Medicina Tropical de São Paulo. Laboratório de Soroepidemiologia e Imunobiologia. Departamentos de Dermatologia; Brasil.The serologic assay is an important tool in the diagnosis of leishmaniasis. One of the most commonly used tests is enzyme-linked immunosorbent assay (ELISA). Since total Leishmania promastigotes are used as antigen in the routine assay, false-positive reactions are frequent due to cross-reaction with sera from other diseases, mainly Chagas' disease. Therefore, an antigen that determines less cross-reactivity has been pursued for the serodiagnosis of leishmaniasis. In the present study we analyzed the use of recombinant Leishmania infantum heat shock protein (Hsp) 83 in ELISA for the serodiagnosis of cutaneous (N = 12) and mucocutaneous leishmaniasis (N = 14) and we observed the presence of anti-L. infantum Hsp 83 antibodies in all samples as well as anti-Leishmania total antigen antibodies. When cross-reactivity was tested, chronic Chagas' disease patients (N = 10) did not show any reactivity. Therefore, we consider this L. infantum Hsp 83 to be a good antigen for routine use for serodiagnosis of tegumentary leishmaniasis

The role of oxidized low density lipoprotein (oxLDL) derived antigens and IgG autoantibodies in coronary artery disease (CAD)

Karolinska Univ Hosp, Ctr Mol Med, Dept Med, Stockholm, SwedenUniv São Paulo, Inst Biomed Sci, São Paulo, BrazilUnifesp, São Paulo, BrazilSante Pazzanese, Inst Cardiol, São Paulo, BrazilUnifesp, São Paulo, BrazilWeb of Scienc

Autoantibody response to chromatographic fractions from oxidized LDL in unstable angina patients and healthy controls

Levels of autoantibodies to oxidized low-density lipoprotein (oxLDL) have been correlated to atherosclerosis; however, contradictory results have been shown. To better understand the role of autoantibodies to oxLDL in atherogenesis, and their potential to predict risk of developing coronary artery disease we investigated the antibody response of unstable angina (UA) patients and healthy controls against chromatographic separated fractions of oxLDL. Five major peaks were detected after chromatographic separation of oxLDL and 10 fractions were collected. Surprisingly, when the response to high molecular weight fractions was analysed, we observed a significant increase in the levels of autoantibodies in controls compared to UA. in contrast, when the autoantibody response to intermediate and low molecular weight fractions was analysed, we observed that the UA group showed consistently higher levels compared with controls. Our data demonstrates that within oxLDL there are major fractions that can be recognized by autoantibodies from either UA patients or healthy individuals, and that the use of total oxLDL as an antigen pool may mask the presence of some antigenic molecules and their corresponding antibodies. Further studies are needed, but the analysis of antibody profiles may indeed open up a novel approach for evaluation and prevention against atherosclerosis.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Laboratorio de Investigacao Medica 38 (LIM 38)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Instituto do Milenio de Fluidos ComplexosInstitute of Cardiology Dante PazzaneseUniv São Paulo, Dept Immunol, Inst Biomed Sci, São Paulo, BrazilKarolinska Univ Hosp, Expt Cardiovasc Res Unit, Ctr Mol Med, S-17176 Stockholm, SwedenUniversidade Federal de São Paulo, Dept Biophys, São Paulo, BrazilInst Cardiol Dante Pazzanese, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Biophys, São Paulo, BrazilWeb of Scienc