Detection Method:Sandwich
Test principle:This assay employs a two-site sandwich ELISA to quantitate DKK4 in samples. An antibody specific for DKK4 has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and anyDKK4 present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for DKK4 is added to the wells. After washing, Streptavidin conjugated Horseradish Peroxidase (HRP) is added to the wells. Following a wash to remove any unbound avidin-enzyme reagent, a substrate solution is added to the wells and color develops in proportion to the amount of DKK4 bound in the initial step. The color development is stopped and the intensity of the color is measured.
Product Overview:All DKKs have several potential sites for cleavage by furin-type proteases. Northern blot analysis detected no expression of DKK4, but RT-PCR analysis detected DKK4 expression in cerebellum, T-cell, esophagus, and lung cDNA libraries. Western blot analysis showed that DKK4 is secreted as 40-, 30- to 32-, and 15- to 17-kD proteins; these proteins were not significantly affected by N-glycanase treatment. N-terminal sequence analysis of the different forms suggested that DKK4 is proteolytically processed, possibly by intracellular furin-type proteases. Functional analysis determined that DKK4 blocks Xenopus Wnt8, Wnt3a, and Wnt2b, but not Dsh or Fz8, induction of a secondary axis in frog embryos, indicating that DKKs antagonize WNT function upstream of WNT receptors.
