1 Quantification of Thiram in Honeybees: Development of a Chemiluminescent ELISA Stefano Girotti, Elisabetta Maiolini, Severino Ghini, Elida Ferri, Fabiana Fini Dipartimento di Scienza dei Metalli, Elettrochimica e Tecniche Chimiche, Università di Bologna, via San Donato 15, 40127 Bologna, Italy Patrice Nodet Laboratoire de Biodiversité et Ecologie Microbienne, ESMISAB, Technopôle Brest-Iroise, 29280 Plouzané, France Sergei Eremin Department of Chemical Enzymology, Faculty of Chemistry, M.V. Lomonosov Moscow State University, Moscow 119992, Russia Abstract: A Chemiluminescence Enzyme-Linked Immuno-Sorbent Assay (CL-ELISA) for determination and quantification of the fungicide thiram in honeybees was developed in an indirect competitive format. The assay was optimized by determining: the optimal coating conjugate concentration and anti-thiram antiserum dilution, the effect of the incubation time on the competitive step, the tolerance to organic solvents. The IC 50 and the limit of detection (LOD) values were 60 ng mL -1 and 9 ng mL -1 , respectively, similar to those of colorimetric ELISA with a calibration range of 9 – 15,000 ng mL -1 . Cross reactivity of some related compounds such as some dithiocarbamates, a thiocarbamate, the ethylenethiourea and the tetramethylthiourea were tested. The assay was then applied to honeybees sample extracts obtained by using the liquid-liquid extraction or the graphitized carbon-based solid phase extraction. hal-00519713, version 1 - 21 Sep 2010 Author manuscript, published in "Analytical Letters 41, 1 (2008) 46-55"