Hapten Design and Antibody Production for a Highly Sensitive Broad-Spectrum Immunoassay for Detection of Seven Organochlorine Pollutants in Soil and Oilfield Chemicals

Organochlorine pollutants (OCPs), as a class of environmentally persistent and bioaccumulative toxic compounds, pose severe threats to ecosystems and human health. This study developed three novel haptens targeting seven chlorinated aromatic OCPs (CA-OCPs), including hexachlorobenzene, chlorothalonil (CLT), tetrachlorophthalide (TCP), pentachloronitrobenzene (PCNB), pentachloroaniline, pentachloroanisole, and pentachlorobenzene, using a shared aromatic scaffold strategy. Quantum chemical analysis of five haptens (three novel and two literature-based) and seven CA-OCPs assessed structural/electrostatic compatibility, enabling optimal hapten selection via structure-activity matching. Animal immunization produced a sensitive broad-spectrum antibody 6F11 (mAb-6F11) with half-maximal inhibitory concentrations of 0.48-16.15 ng/mL for CA-OCPs after heterologous coating optimization. Molecular docking revealed that the broad-spectrum recognition capability of mAb-6F11 is mediated by key residues (TYR-109, TYR-59, TYR-103, TRP-93, and ILE-52) within its binding pocket. Electronegative groups in CLT/TCP/PCNB enhanced the cross-reactivity through hydrogen bonds (TRP-93/GLY-106/TYR-109) and carbon-hydrogen bond interactions (TYR-105). A colloidal gold immunochromatographic assay (GICA) developed using mAb-6F11 enabled simultaneous on-site detection of all seven CA-OCPs in soil and oilfield chemicals (OFCs), with visual detection limits of 0.005-0.10 mg/kg (soil) and 0.5-10 mg/kg (OFCs). Field validation demonstrated strong consistency between GICA and gas chromatography-tandem mass spectrometry results, confirming the method's accuracy, stability, and reliability for preliminary screening of real samples.