Biological monitoring of 2,4,5-trichlorophenol (I): preparation of antibodies and development of an immunoassay using theoretical models.
Antibodies against 2,4,5-trichlorophenol have been prepared after theoretical and molecular modeling chemical studies of three potential immunizing haptens with the aim to find out the one mimicking best the target analyte. Competitive direct and indirect ELISAs have been developed after screening a battery of haptenized enzyme tracers and coating antigens, respectively. The relation between the degree of heterology of the competitor and the resulting immunoassay detectability has been investigated according to the electronic similarities of the competitor haptens with the target analyte taking in consideration their pK(a) values. These studies have been performed using theoretical and molecular modeling tools to find out their electronic distribution at their minimum energetic levels. The results suggest that the competitors should have a high homology to produced assays with good detectability values. On the other hand detectability improves when lowering the hapten density of the competitors. An indirect competitive ELISA has been finally selected for further investigation. The immunoassay has an IC(50) value of 0.6 microg L(-)(1) and a limit of detection of 0.084 microg L(-)(1). The selectivity of the assay is high in relation to other chlorophenols frequently present in real samples. In contrast, the brominated analogues may also be recognized with this assay.
Nichkova
Mikaela
M
Department of Biological Organic Chemistry, IIQAB-CSIC, Jorge Girona, 18-26, 08034, Barcelona, Spain.
Galve
Roger
R
Marco
M-Pilar
MP
D000906:Antibodies / Q000737:chemistry* / Q000378:metabolism; D001667:Binding, Competitive; D002733:Chlorophenols / Q000032:analysis / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005069:Evaluation Studies as Topic; D006241:Haptens / Q000737:chemistry / Q000276:immunology; D008958:Models, Molecular; D012680:Sensitivity and Specificity
D000906:Antibodies; D002733:Chlorophenols; D006241:Haptens; C009534:2,4,5-trichlorophenol