Quantitative Determination of a Peanut Anionic Peroxidase by Specific Monoclonal Antibodies

A monoclonal antibody (46-12-C12) for use in a solid-phase enzyme-linkedimmunosorbent assay (ELISA) specific for an anionic peroxidase(APRX) from peanut (Arachis hypogaea L.) suspension cell mediumwas developed. The McAb (IgG₁) had a high affinity (2.77 ? 10¹¹)and specificity for APRX, and showed only weak interaction witha-amylase and virtually no reactivity with other enzymes, suchas MCPRX (peanut), minor CPRX (peanut), peroxidase (horseradish),RuBP case (spinach), -glucosidase (rice), ß-glucosidase(almonds), acid phosphatase (potato), catalase (bovine liver)and glucose-6-phosphatase (yeast). Sample dilution curves werefound to parallel the standard curve. The detection limit was0.002 ? 10^–12 mol APRX. The absorbance was linear at concentrationsbetween 0.004–24 ? 10^–12 mol APRX. Three hundredsamples could be analysed per day by one person, with a semi-automaticperformance. Using this assay, levels of APRX have been determinedin a number of biological extracts of different origin. Key words: Peanut, anionic peroxidase, monoclonal antibody, enzyme-linked immunosorbent assay (ELISA)