Quantification of hemorphin-7 peptides by enzyme linked immunosorbent assay with secondary antibody

Abstract

This paper describes an enzyme linked immunosorbent assay (ELISA) for the quantification of bioactive peptides: hemorphin-7 peptides (LVV-hemorphin-7, VV-hemorphin-7 and hemorphin-7) using an anti-rabbit secondary antibody conjugated to peroxidase for sandwich antibody assay. Bovine serum albumin (BSA) conjugated to VV-hemorphin-7 was readily coated on a polystyrene microplate. Data acquisition on microtiter wells is performed by an ELISA reader. The standard curve was produced for 1×10 −13 to 2×10 −5 M VV-hemorphin-7. The minimum detectable amount of VV-hemorphin-7 is 5×10 −13 mol and the relative standard deviation was 8.8% for a 1×10 −6 M sample ( n=8). The ELISA procedure was selective with respect to structurally similar compounds. This method was applied to quantify hemorphin-7 peptides in bovine plasma.