Novel elisa testing system

Details Novel elisa testing system Novel elisa testing system

G - Physics

01

N

G01N 33/543 (2006.01) G01N 37/00 (2006.01)

Patent

CA 2300687

Enzyme-linked immunosorbent assay (ELISA) is perhaps one of the most widespread assay methods used extensively in the field of immunology. This assay works on the basis of known amounts of antigens or antibodies adhered onto a solid surface made of polystyrene. The polystyrene-made wells are opened at only one side and each well is eight mm in diameter, ten mm in depth and holds up to a maximum of 300 µL, in volume. Every 8 or 12 wells joins together to form a 96-well or a 48-well combination unit. The inner wall of each well is smooth with a transparent flat base. Generally, approximately 100 µL of known antigens or antibodies is added to each well occupying one- third of the well. Known amounts of antigens or antibodies are allowed to adhere onto the polystyrene surface at a specific temperature for over 10 hours. This process is often known as "coating". Following this, protein solutions at high concentrations or solutions with concentrated proteins and glucose are allowed to further adhere onto the polystyrene wells to ensure maximum occupancy of the well surfaces. In addition, this second coating offers better protection on the activity of the antigens or antibodies. This second process is termed "blocking". The commonly used enzyme (horseradish peroxidase) can be linked either to the antigens or the antibodies, but in either case the solid-phase reactant is unlabeled. ELISA can be made quantitative by setting it up as a competitive inhibition assay in which free or unknown amounts of antigens (or antibodies) in samples compete with enzyme- labeled antigens (or antibodies) for binding sites on the polystyrene solid phase coated with known amounts of antigens or antibodies. Following appropriate washings, the concentration of enzyme bound to the immune complex is estimated by the addition of its substrate that changes colour when the substrate is acted on by the enzyme. This highly- specific immunoassay has thusfar become one of the most popular immunoassays in immunology and immunology-related studies.

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