Development of monoclonal antibodies against bovine mucin core 2 beta 6 N-acetylglucosaminyltransferase

Molecular cloning techniques have been used to produce abundant amounts of recombinant glycosyltransferases for biochemical studies. We recently clone d a cDNA which encoded bovine mucin core 2 beta 6N-acetylglucosaminyl trans ferase (C2TF). Poly-histidine-C2TF fusion protein was generated from the cl oned cDNA in the E. coli Xpress system and used to produce monoclonal antib odies (MAbs). We obtained seven hybridomas which secreted MAbs against bovi ne C2TF in mouse ascites with titers ranging from 1:1280 to 1:40960 as asse ssed by immunofluorescence assay (IF). Isotyping revealed that all seven MA bs were IgG (4 IgG1, 2 IgG2b and 1 IgG2a). The affinity constants (M(-)1) f or these MAbs range from 5.4 x 10(7) to 1.2 x 10(9). These MAbs recognized bovine C2TF in tissue sections and on Western blottings. Six of these MAbs reacted with human core 2-M enzyme and one with both core 2-L and core 2-M enzymes on Western blottings. Therefore, These antibodies should be useful for further study of bovine and human core 2 enzymes.