Detection Method:Sandwich
Test principle:This assay employs a two-site sandwich ELISA to quantitate DOT1L in samples. An antibody specific for DOT1L has been pre-coated onto a microplate. Standards and samples are pipetted into the wells and anyDOT1L present is bound by the immobilized antibody. After removing any unbound substances, a biotin-conjugated antibody specific for DOT1L 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 DOT1L bound in the initial step. The color development is stopped and the intensity of the color is measured.
Product Overview:DOT1L is inactive against free core histones, but shows significant histone methyltransferase activity against nucleosomes. DOT1L is a nucleosomal histone-3 (H3)-specific methyltransferase that requires the SAM and sequences between amino acids 351 and 472 for activity. Overexpression of DOT1L in human embryonic kidney cells significantly increased methylation of lys79 of H3, but had no effect on lys4 or lys9. Methylation of lys79, attributed to DOT1L, was regulated during the cell cycle in synchronized HeLa cells. The deduced 1,537-amino acid protein has a calculated molecular mass of 165 kD and contains a SAM (sterile alpha motif) domain for protein-protein interaction. DOT1L does not, however, have the SET domain found in most histone lysine methyltransferases.
