Enzyme immunoassay for the quantitative determination of Lysozyme in food.
Background: Hen’s egg (Gallus gallus) is very rich of proteins and represents an important food source for humans. While proteins of egg yolk only have minor allergenicity, many proteins of egg white are known to be allergenic. In addition to ovalbumin, ovotransferrin, ovomucoid and livetin, lysozyme represents an important allergen. Primarily lysozyme is used as a preservative in wine and cheese industry. For allergic persons the consumption of lysozyme represents a critical problem. Already very low amounts of the allergen can cause allergic reactions, which may lead to anaphylactic shock in severe cases. Because of this, lysozyme allergic persons must strictly avoid the consumption of lysozyme containing food. Nondeclared addition of lysozyme in food is hazardous for allergic people. Cross-contamination, mostly in consequence of the production process, is also problematic. Since July 1, 2012 the European Union requests allergen labeling for wine if milk or egg proteins are used during the production and are still present at a detection level of 0.25 mg/L or greater. Thus for the detection of lysozyme residues, sensitive assay systems are required.
Lysozyme ELISA represents a highly sensitive detection system for lysozyme and is particularly capable of the quantification of lysozyme residues in wine and cheese.
Description: The Lysozyme quantitative test is based on the principle of the enzyme linked immunosorbent assay. An antibody directed against lysozyme is bound on the surface of a microtiter plate. Lysozyme containing samples or standards are given into the wells of the microtiter plate. After 20 minutes incubation at room temperature, the wells are washed with diluted washing solution to remove unbound material. A peroxidase conjugated second antibody directed against lysozyme is given into the wells, and after 20 minutes of incubation the plate is washed again. A substrate solution is added and incubated for 20 minutes, resulting in the development of a blue colour. The colour development is inhibited by the addition of a stop solution, and the colour turns yellow. The yellow colour is measured photometrically at 450 nm. The concentration of lysozyme is directly proportional to the colour intensity of the test sample.
- The kit contains reagents for 96 determinations;
- Microtiter plate consisting of 12 strips with 8 breakable wells;
- Lysozyme standards: 0, 25, 50, 100, 250 ppb;
- ELISA reader at 450 nm;
- Standard range: 25 - 250 ppb;
- Sensitivity analytical: wine: 2 ppb; others: 10 ppb.