Development of a sandwich vertical flow immunogold assay for rapid detection of oxytetracycline residue in fish tissues.

Highlights • Development, comparison and validation of silver enhanced, multiplex sandwich vertical flow immunogold assay (SVIA) for sensitive and rapid detection of OTC residues in edible fish tissues. • Combining silver enhancement with immunogold labelled MAb improved the sensitivity. • Employing anti-OTC capture rabbit antiserum and detector monoclonal antibody provided high specificity with a visual detection limit of 2 ng mL−1. • It is a semi-quantitative, simple, rapid, low cost assay compatible with field level analysis. Abstract A rapid multiplex silver enhanced ‘sandwich vertical flow immunogold assay (SVIA)’ based on disposable porous filter-membrane was developed for on-site detection of oxytetracycline (OTC) residues in fish tissues. Artificial antigens were synthesized to raise selective anti-OTC rabbit antiserum and highly specific monoclonal antibody (mAb). The assay consists of three layers. The first layer was composed of immobilized anti-OTC rabbit antiserum (capture antibody) onto the membrane, the second layer was the molecule of interest (OTC) and the third layer was refined with detector immunogold labeled mAb. The sensitivity of the silver enhanced SVIA was as low as 2 ng mL−1 (within 4 min) representing a 125-fold increase in sensitivity over the HRP labeled sandwich vertical flow assay. The reliability of developed technique was co-evaluated with HPLC and validated using 115 field fish samples from different regions of southern India. The results demonstrated that SVIA has the high potentiality to be established as a semi-quantitative routine on-site screening tool. [ABSTRACT FROM AUTHOR]