OBTAINING A FUNCTIONAL RECOMBINANT ANTIRHESUS-(D) ANTIBODY USING THE BACULOVIRUS-INSECT CELL EXPRESSION SYSTEM

The cloning and production of a human anti-rhesus (Rh) D monoclonal an tibody (mAb) using the baculovirus-insect cell expression system is de scribed. This monoclonal recombinant antibody R.D7C2 derived from a hu man parental IgM lambda immunoglobulin was obtained after immortalizat ion of lymphocytes by Epstein-Barr virus (EBV). The human heavy (VH) a nd light (VL) variable regions were cloned from the parental cell line and genetically fused to the human constant IgG1 heavy (H) and light (L) chain genes (gamma l and lambda, respectively). ii recombinant bac ulovirus was constructed that directs the co-expression of genes encod ing both genetically fused heavy and light chains under the control of two late and strong baculovirus promoters. After infecting the Spodop tera frugiperda (Sf9) insect cell line with this baculovector, a compl ete IgG1 mAb was secreted in the culture medium indicating that each i mmunoglobulin chain was correctly processed and assembled with a funct ional glycosylation into a tetrameric form. In vitro analysis showed t hat the functional properties of R.D7C2; using agglutination tests wer e efficient for the specific recognition of Rh-D-positive red blood ce lls (RBC). In addition, R.D7C2 showed effector functions of the gamma l heavy chain resulting in the lysis of Rh+ papain RBC by an antibody- directed cellular cytotoxicity mechanism. These results demonstrate th at R.D7C2 can be produced in the baculovirus-insect cell expression sy stem as a source for potential therapeutic application in the treatmen t of the haemolytic disease of the newborn.