Design and synthesis of immunoconjugates and development of competition inhibition enzyme-linked immunosorbent assay (CIEIA) for the detection of O-isopropyl methylphosphonofluoridate (sarin): An organophosphorous toxicant

Abstract

Three haptens of the organophosphorus (OP) toxicant ‘sarin’ having different spacer arm were designed and synthesized. Haptens were conjugated with BSA (bovine serum albumin) and ovalbumin (OVA) for raising antibody and coating antigen. High antibody titer with higher specificity was obtained from 4-(4-(isopropoxy(methyl)phosphoryloxy)phenylamino)-4-oxobutanoic acid (hapten B) having reasonable long spacer arm. For the standard curve, an IC 50 (inhibitory concentration) of free antigen was found to be 0.415 μg mL −1 on the basis of indirect competitive ELISA. The study revealed that heterology in competition inhibition enzyme immunoassay (CIEIA) produced remarkable improvement in the sensitivity and specificity of the assay. Under the optimized conditions, the quantitative working range was found to be 0.19–1.56 μg mL −1 with a limit of detection (LOD) of 0.05 μg mL −1. The antibodies showed negligible cross reactivity (CR) with other OP toxicants and pesticides, which makes the assay suitable for the selective detection of sarin.