A solid phase sandwich enzyme-linked-immunosorbent assay (ELISA).
The assay is a solid phase sandwich enzyme linked-immunosorbent assay (ELISA) for the in vitro quantitative determination of mouse Tau in mouse brain homogenates, cell extracts, buffered solutions, or cell culture media. The assay will recognize both natural and recombinant mTau.
The assay uses a monoclonal capture antibody specific for mouse tau that has been coated onto multiwell strips. Standard dilutions and samples are incubated 2 hours at RT. Tau antigen binds to the capture antibody. After a wash, a detection antibody specific for the non-phosphorylated or phosphorylated protein is incubated 1 hr at RT, which results in binding to the immobilized Tau protein. An anti-rabbit IgG-HRP binds to the immobilized protein completing the four-member sandwich. The reaction is visualized by tetramethylbenzidine (TMB) substrate, followed by the stop solution. The intensity of the yellow color measured in a multiwell plate reader at 450 nm is directly proportional to the concentration of Tau in original sample. The unknown concentrations are calculated from the standard curve run with each assay
Features and Benefits
Tau ELISA is a simple, sensitive, and specific alternative to immunoblotting and RIA.
The specificity was confirmed by peptide competition. The data shows that only the peptide corresponding to the region surrounding serine 199 containing the phosphoserine blocks the ELISA signal.