59D-18 |
Enzyme linked immunosorbent assay (ELISA) for Inca peanut (Plukenetia volubilis L.) albumin |
T. WESTMORELAND1, M. Venkatachalam1, K. H. Roux2, and S. K. Sathe1. (1) Department of Nutrition, Food and Exercise Sciences, Florida State University, 402 Sandels Bldg., College of Human Sciences, Tallahassee, FL 32306-1493, (2) Department of Biological Science, Florida State University Inca peanut (IP), a plant native to the high altitude rain forests of the Andean region of South America grows as a vine and produces tetralobular fruit containing one seed, with a nut like coat in each fruit. The seeds are valued for their high oil (49%) and protein (33%) content. Inca peanut seeds contain a nutritionally complete and a highly digestible water soluble (albumin) storage protein (IPA) that accounts for 31% total seed proteins. To our knowledge IPA is the first plant protein to contain all the essential amino acids required by an adult human. It is not known whether this protein is expressed only in Inca peanut seeds or it is also expressed in other commonly used seeds as human foods. It is therefore necessary to develop a sensitive assay that can detect trace amounts of IPA. Our objective was to use rabbit polyclonal antibodies (pAbs) raised against IPA in inhibition ELISA for the purpose of IPA detection. A rabbit was immunized and boosted with 1mg IPA in Freund's complete and incomplete adjuvants respectively. The assay was optimized for incubation temperature (4, 25, 370C) and time (1, 4, 24 h) for antigen coating and primary and secondary antibody incubation. Cross reactivity of pAbs against several seed protein extracts was reported as IC50 for IPA/IC50 for the test seed protein extract. Typically, primary and secondary antibody dilutions of 104 and 5 x 103, respectively, permitted reproducible results. Coating conditions did not influence the degree of antigen coating. The assay was not dependent on incubation time and temperature of the incubation steps.
Session 59D, Food Chemistry: Proteins and Physicochemical Properties
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