Cloning and characterization of a single chain antibody to glucose oxidase from a murine hybridoma.
Glucose oxidase (GOD) is an oxidoreductase catalyzing the reaction of glucose and oxygen to peroxide and gluconolacton (EC 1.1.3.4.). GOD is a widely used enzyme in biotechnology. Therefore the production of monoclonal antibodies and antibody fragments to GOD are of interest in bioanalytics and even tumor therapy. We describe here the generation of a panel of monoclonal antibodies to native and heat inactivated GOD. One of the hybridomas, E13BC8, was used for cloning of a single chain antibody (scFv). This scFv was expressed in Escherichia coli XL1-blue with the help of the vector system pOPE101. The scFv was isolated from the periplasmic fraction and detected by western blotting. It reacts specifically with soluble active GOD but does not recognize denatured GOD adsorbed to the solid phase. The same binding properties were also found for the monoclonal antibody E13BC8.
Department of Biotechnology, Institute of Biochemistry and Biology, University of Potsdam, Karl-Liebknecht-Strasse 24/25, D-14476 Golm, Germany.
Frank
F
Sellrie
Jörg A
JA
Schenk
Olaf
O
Behrsing
Oliver
O
Drechsel
Burkhard
B
Micheel
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics*; D001234:Aspergillus niger / Q000201:enzymology; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D005949:Glucose Oxidase / Q000276:immunology*; D006825:Hybridomas / Q000276:immunology; D007128:Immunoglobulin Fragments / Q000096:biosynthesis / Q000235:genetics*; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis / Q000235:genetics; D007147:Immunoglobulin Light Chains / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data
D000911:Antibodies, Monoclonal; D017931:DNA Primers; D007128:Immunoglobulin Fragments; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; C055966:immunoglobulin Fv; D005949:Glucose Oxidase