TY - JOUR A1 - Schneider, Rudolf A1 - Schmidt, S. A1 - Hanelt, S. A1 - Canitz, C. A1 - Hoffmann, H. A1 - Garbe, L.-A. T1 - Synthetic Strategies for the Modification of Diclofenac N2 - For many heterogeneous sensor applications as well as the synthesis of hapten antigens to produce antibodies, protein conjugates of the target substance are essential. A requirement is that the target substance already offers or is modified to contain a functionality that allows for coupling to a protein, that is, an amino acid residue. Ideally, to avoid shielding of the compound by the carrier protein, a sufficient distance to the protein surface should be provided. With its carboxyl function diclofenac (DCF) allows for direct binding to lysine residues after in situ synthesis of the NHS ester. One problem is that diclofenac as free acid tends to autocondensation, which results in low yields. Here we describe the 'insertion' of a C6 spacer via synthesis of the amide with 6-aminohexanoic acid. To carry out the reaction in solution, first the methyl ester of the amino acid had to be produced. Due to otherwise low yields and large cleaning efforts, solid-phase synthesis on Fmoc Ahx Wang resin is recommended. The crude product is mainly contaminated by cleavage products from the resin which were removed by chromatography. The structure of the highly pure hapten was completely determined by nuclear magnetic resonance (NMR) spectroscopy. KW - Diclophenac KW - Hapten synthesis KW - Solid-phase synthesis PY - 2017 U6 - https://doi.org/10.1055/s-0036-1588858 SN - 0936-5214 VL - 28 IS - 15 SP - 1984 EP - 1989 PB - Georg Thieme Verlag AN - OPUS4-43316 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schmidt, Stephan A1 - Hanelt, S. A1 - Canitz, C. A1 - Hoffmann, Holger A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Entwicklung monoklonaler Antikörper gegen Diclofenac N2 - Diclofenac (DCF) wird als nicht-steroidales, antientzündliches Arzneimittel gegen Fieber, Entzündungen, rheumatische Erkrankungen und Schmerzen eingesetzt. 70 % werden nach Einnahme wieder ausgeschieden. Aufgrund unvollständiger Elimination in den Kläranlagen lassen sich Rückstände im Trinkwasser und im Oberflächengewässer nachweisen. Diclofenac wird in der Umweltanalytik v.a. mittels GC-MS oder LCMS nachgewiesen. Bioanalytische Methoden machen sich die spezifische Antigen-Antikörper-Wechselwirkung zunutze, so z.B. Immunoassays. Die hierfür erforderlichen Antikörper werden durch Immunisierung mit einem Konjugat aus dem Analyten und einem Trägerprotein gewonnen. Die Kopplung kann direkt über die Carboxylfunktion von Diclofenac erfolgen. Um Abschirmungseffekte zu vermeiden, kann zwischen Protein und Analy1 ein Spacer sinnvoll sein. PY - 2015 SN - 1618-3258 SN - 1617-5301 IS - 4 SP - 107 EP - 109 PB - Wiley-VCH Verl. CY - Weinheim AN - OPUS4-35241 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Baldofski, Stefanie A1 - Canitz, C. A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Studies on the development of antibodies for the highly hydrophobic plasticizers DINCH and DEHT N2 - Diisononylcyclohexane-1,2-dicarboxylate (DINCH) and di-2-ethylhexyl terephthalate (DEHT), two of the most important substitutes for phthalate plasticizers, are used for a wide range of applications. Consequently, an increasing occurrence in urine and environmental samples is reported. Reliable and fast analytical methods for the quantification of these plasticizers are needed. So far, mainly GC-MS or LC-MS methods are used. We aimed to develop the first antibodies and immunoassays allowing for high-throughput analysis of samples. We designed two DINCH hapten structures and one DEHT hapten structure and employed hapten-protein conjugates for the immunization of rabbits. Sensitive competitive enzyme-linked immunosorbent assays (ELISAs) against each hapten using the produced polyclonal antibodies were established. Yet, binding of DINCH to the respective antibodies was not observed in neither direct nor indirect assay formats, even when using protein conjugates with the heterologous haptens and different carrier proteins in the indirect format. The use of surfactants and solvents in the sample buffer did not result in recognition of the plasticizers. Also, no binding of DEHT in ELISA employing the respective antibodies was detected. We speculate that the production of antibodies against these highly hydrophobic molecules is not possible via our route, however a different hapten design could overcome this obstacle. KW - Weichmacher KW - Phthalate KW - Antikörper KW - Selektivität KW - Ersatzstoffe KW - Polyclonal antibodies KW - Phthalate substitutes KW - Antibody production for hydrophobic molecules KW - DEHT KW - DINCH KW - Hapten design PY - 2018 U6 - https://doi.org/10.1016/j.ab.2017.12.006 SN - 0003-2697 VL - 543 SP - 90 EP - 96 PB - Elsevier Inc. CY - Amsterdam, NL AN - OPUS4-44015 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -