Review articleProduction of monoclonal antibodies: Strategy and tactics
Abstract
A myeloma line has been developed which produces no globulin chains of its own, has a duplication time of 8.7 h, fuses effectively with B-lymphoblasts and produces stable hybrids.
An enhancing effect of macrophages on hybridoma yields has been observed. Among the fusing agents tested, PEG of mol.wt. 4000 gave the best results, 20°C being the optimum working temperature. The maintenance medium of choice has been found to be Iscove's with 10% FCS. Direct exposure of fusion cultures to a selective medium with hypoxanthine, aminopterine and thymidine reduced the labor involved and increased the yield. A mechanical device for changing the medium has been designed. The replacement of standard trays by microtrays resulted in a higher frequency of surviving hybrids.
By using a feeder layer, the spleen cell input can be reduced 50-fold. At such low multiplicities the positive cultures arise predominantly from single hybrids, eliminating the need for subsequent cloning.
The hybrids can be labelled and will yield in serum-free medium. Since at least a third of them inherit the fast growth rate of their myeloma parent and keep producing over 2000 antibody molecules per second, readaptation to ascitic growth is also superfluous.
A simplified technique of producing monoclonal antibodies is given in detail, together with the experimental evidence prompting modifications of the classical method of Köhler and Milstein (1975, 1976).
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