Quantitative Sandwich ELISA for the Determination of Lupine (Lupinusspp.) in Foods

Abstract

The use of lupine in foods has increased considerably during the past decade, reflected by a corresponding increase in reported lupine-induced allergic incidents. Lupine allergy may arise either by primary sensitization or by clinical cross-reactivity in peanut-allergic persons. Detection of lupine proteins in food has previously been based on the use of patient serum. A novel sandwich enzyme-linked immunosorbent assay (ELISA) for the detection and quantification of lupine in processed foods was developed, using a polyclonal rabbit antilupine capture antibody and a biotinylated conjugate of the same antibody for detection. The antibody was highly specific for lupine, apart from minor cross-reactivities to other legumes. The assay had a detection limit of 1 mug/g and was successfully used to quantify lupine protein in various food matrixes. Recoveries ranged from 60 to 116%, while the intra-and interassay coefficients of variation were <6% and <21%, respectively.

Characterization of IgE Binding to Lupin, Peanut and Almond with Sera from Lupin-Allergic Patients

“…All protein extracts, if not otherwise stated, were prepared as described previously [17]. In brief, homogenized samples (2 g) were extracted with 10 ml of 0.1 M Tris-0.5 M glycine, pH 8.7, overnight at 45°C, and centrifuged at 39,200 g for 25 min at 4°C.…”

“…In earlier studies performed in this laboratory, anti-lupin polyclonal and monoclonal antibodies have been generated, characterized and used in the development of enzyme-linked immunosorbent assay (ELISA) methods for the detection of lupin in foods [16,17,18]. In the comprehensive specificity testing of the antibodies, the monoclonal antibody was found to cross-react with almond, which was a somewhat unexpected finding because these plants are phylogenetically distant.…”

Narrow‐leafed lupin (Lupinus angustifolius L.) β‐conglutin proteins modulate the insulin signaling pathway as potential type 2 diabetes treatment and inflammatory‐related disease amelioration

“…In this regard, allergenic proteins of 71, 59 and 34 kDa [14] have been identified and isolated using sera from lupin allergen patients, with protein sizes matching alpha, beta and gamma families, respectively. Lupin allergy may arise either by primary sensitisation, or by clinical cross-reactivity in peanut-allergic persons [15,16]. Various reports suggest the common clinical pattern of lupin allergy is the triggering of an allergic reaction via ingestion of lupin in peanut-allergic individuals.…”

Opinion of the Scientific Panel on Dietetic products, nutrition and allergies [NDA] related to the evaluation of lupin for labelling purposes

“…The use of serum from allergic patients in such methods hampers standardisation, and the limited availability of such serum prevents methods depending on this from becoming routine analyses. A quantitative sandwich ELISA that utilizes polyclonal rabbit anti-lupin antibodies can however be used as a routine method to detect lupin in food products with a detection limit of 1 mg lupin protein per 1 kg of food (Holden et al, 2005). Generally, the extensive immunological cross-reactivities between legumes will represent a problem in the development of lupin-specific immunological assays Ibáñez et al, 2003;.…”