Development of a bispecific monoclonal antibody as a universal immunoprobe for detecting biotinylated macromolecules

Bispecific monoclonal antibody (BsMab) combining two different antigen binding sites, anti-biotin and anti-HRPO paratopes, could be used as a universal immunoprobe for detecting all biotinylated macromolecules. First, a mouse hybridoma cell line secreting monospecific anti-biotin Mab was generated and characterized. Second, a quadroma cell line which could continuously secrete bsMab (anti-biotin x anti-HRPO) was developed by a nonselective microelectrofusion method. The supernatant containing bsMab was collected from tissue culture medium and purified with two affinity columns. This bsMab has comparable avidity to commercial streptavidin-HRPO when tested against biotinylated macromolecules. Compared to streptavidin, this bsMab can bind the enzyme and thus eliminate the need for chemical conjugation. This bsMab can be used as a promising immunoprobe for detecting many macromolecules bearing biotin markers, such as protein, phage, liposome and DNA in different bioassay systems.