Faster ELISA with Better Results
White Paper Apr 22, 2016
The search for biomarkers for the early detection of human diseases such as cancer, or the identification of novel drug targets in discovery requires an assay that delivers specific, accurate, and sensitive quantification of target proteins. The ideal assay uses raw sample types, such as plasma and serum, which eliminate any bias that results from the sample-preparation process, avoids lengthy and messy sample purification, and reduces the time to results. Protein identification methods like mass spectrometry or antibody-based approaches like western blotting or immunofluorescence can be used, but none of these approaches provides linear quantification of target protein in raw sample types. Only ELISA meets the dual requirements of working with crude biofluids and achieving accurate quantification of the target protein.
The most sensitive and specific format for ELISA is the “sandwich” format, which requires two different antibodies that bind to at least two distinct sites on the target protein. The capture antibody is pre-coated onto a surface, commonly the wells of a 96-well microplate, and selectively binds to the target protein. After a wash step, the detector antibody, which is modified with a detection tag, is added and binds to a second site on the target protein, forming a sandwich complex. However, as a conventional sandwich ELISA may take three hours or more to complete, less time consuming approaches are in demand.
One novel way to speed up an ELISA is to use a homogeneous format where in the antibody/analyte sandwich complex is formed in solution in a single step. SimpleStep ELISA® technology uses a sandwich ELISA format with a novel, streamlined protocol. In a single step, the complete sandwich complex forms in the well and is anchored to the plate with an immunoaffinity tag. Only one incubation and one wash step are required, compared to multiple incubation and wash steps for a conventional ELISA.