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  • A relevant in vitro ELISA test in alternative to the in vivo NIH test for human rabies vaccine batch release. 24161572

    To assess the quality of vaccine batches before release, international regulation requires the control of potency of each lot of human rabies vaccines by the in vivo NIH challenge test. Meanwhile, the 3Rs strategy for animal testing encourages the replacement of the in vivo potency test by an in vitro assay. Consequently, since more than 10 years, an ELISA method has been implemented by ANSM in parallel to the NIH test for rabies vaccines lots. It consists in the evaluation of the glycoprotein content using a monoclonal antibody recognizing the trimeric native form of the glycoprotein. This ELISA method is able 1) to monitor the consistency of production with a similar profile than the NIH; 2) to detect a low quantity of glycoprotein in vaccines and 3) to agree with the manufacturer's NIH results by declaring a non compliant batch. This ELISA which characterizes the immunogenic form of the glycoprotein formulated in vaccines seems to be relevant to replace the NIH test and is a promising candidate to be standardized by a collaborative study.
    Document Type:
    Reference
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  • A sensitive and specific ELISA detects methionine sulfoxide-containing apolipoprotein A-I in HDL. 18832772

    Oxidized HDL has been proposed to play a key role in atherogenesis. A wide range of reactive intermediates oxidizes methionine residues to methionine sulfoxide (MetO) in apolipoprotein A-I (apoA-I), the major HDL protein. These reactive species include those produced by myeloperoxidase, an enzyme implicated in atherogenesis. The aim of the present study was to develop a sensitive and specific ELISA for detecting MetO residues in HDL. We therefore immunized mice with HPLC-purified human apoA-I containing MetO(86) and MetO(112) (termed apoA-I(+32)) to generate a monoclonal antibody termed MOA-I. An ELISA using MOA-I detected lipid-free apoA-I(+32), apoA-I modified by 2e-oxidants (hydrogen peroxide, hypochlorous acid, peroxynitrite), and HDL oxidized by 1e- or 2e-oxidants and present in buffer or human plasma. Detection was concentration dependent, reproducible, and exhibited a linear response over a physiologically plausible range of concentrations of oxidized HDL. In contrast, MOA-I failed to recognize native apoA-I, native apoA-II, apoA-I modified by hydroxyl radical or metal ions, or LDL and methionine-containing proteins other than apoA-I modified by 2e-oxidants. Because the ELISA we have developed specifically detects apoA-I containing MetO in HDL and plasma, it should provide a useful tool for investigating the relationship between oxidized HDL and coronary artery disease.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
  • A simple immuno-capture ELISA to estimate rabies viral glycoprotein antigen in vaccine manufacture. 16188454

    Rabies is an endemic, fatal zoonotic disease in the developing countries. Prevention and post-exposure therapy require safe and efficacious vaccines. The vaccine potency depends on the amount of immunogenic rabies viral glycoprotein antigen in the vaccine preparation. In order to estimate the rabies viral glycoprotein antigen, a specific monoclonal antibody was developed and used in an immuno-capture ELISA (IC-ELISA). The monoclonal antibody binds a conformational epitope on the natively folded rabies viral glycoprotein as indicated by specific, membrane fluorescence on unfixed, rabies virus infected murine neuroblastoma (MNA) cells and glycoprotein gene encoding plasmid transfected COS cells. In addition, the monoclonal antibody competes with and blocks a glycoprotein antigen site III binding monoclonal antibody (mAb-D1, Institut Pasteur, Paris, France). The monoclonal antibody was used in an IC-ELISA using an in-house standard to quantify the rabies viral glycoprotein antigen in 12 vaccine preparations with potency values ranging from 4 to 18 IU. The results indicated a good correlation with the NIH mouse potency assay (r=0.83). The immuno-capture ELISA described in this study can be used to quantify the immunogenic rabies viral glycoprotein antigen in the inactivated rabies viral antigen preparation in a simple and rapid format, which enables better vaccine formulation.
    Document Type:
    Reference
    Product Catalog Number:
    Multiple
    Product Catalog Name:
    Multiple