Quantitative determination of major capsaicinoids in serum by ELISA and time-resolved fluorescent immunoassay based on monoclonal antibodies

To monitor capsaicinoids in serum on-site, three new monoclonal antibodies (mAbs) were firstly proposed using a conjugate of 4-[(4-hydroxy-3-methoxybenzyl) amino]-4-oxobutanoic acid as the immunogen. Among them, the YQQD8 mAb showed the highest sensitivity and cross-reactivity to major capsaicinoids, such as capsaicin, dihydrocapsaicin and N-vanillylnonanamide. A competitive indirect enzyme-linked immunosorbent assay (icELISA) and a time-resolved fluorescent immunochromatographic assay (TRFICA) were established based on this mAb. The linear range was 1.1-27.0 ng mL−1 for icELISA and 1.9-62.5 ng mL−1 for TRFICA and the limit of detection (LOD) of TRFICA was 1.5 ng mL−1. To decrease the interference of sample components and increase accuracy, serum samples were diluted four times before assays. As a result, the linear range of serum samples was 4.6-107.9 ng mL−1 for icELISA and 7.6-250.0 ng mL−1 for TRFICA. Both icELISA and TRFICA showed good recoveries (91.0-112.8% for icELISA and 87.6-111.5% for TRFICA) and concordant results in spiked experiments. Overall, this is the first report of immunoassay based on the mAbs for quantitative determination of major capsaicinoids, and the results demonstrate that both methods can meet the demands of rapid on-site assay for capsaicinoids in serum samples.