Cell fusion, exemplified by the creation of hybridomas secreting MAbs, is widely used in bioscience. Fusion techniques, however, are time consuming, unpredictable, and inefficient, largely because current methods lack control of cell paring, a necessary step in producing hybrids capable of growth in culture. Acoustic theory, supported by recent experiments, predicts that cells of like size selectively pair in an ultrasound field. We will develop and optimize a new method, electro-acoustic fusion (EAF), which uses ultrasound to induce cell pairing prior to or after electroporation in an electric field. The project seeks to determine the optimum conditions for acoustic pairing, and the sequence and parameters for electrofusion, leading to production of mouse hybridomas. Anticipated benefits include: smaller number of cells required than for conventional chemical fusion methods; elimination of toxic chemicals, and use of physiological media for fusion, leading to greater viability; earlier appearance of hybridoma clones; selective formation of myeloma-B lymphocyte hybrids over nonviable hybrids or syncytia; and more reproducible results. Although the experiments emphasize hybridoma technology, EAF should have widespread applications in somatic cell hybridization (e. g., plant cells, in vitro fertilization, prokaryotic and eukaryotic cell hybridization, and production of hybridomas of human origin) as a better alternative to the increasingly prevalent use of electrofusion instruments, which use alternating current in unphysiological hypotonic media to form cell aggregates prior to DC electrofusion. An EAF instrument could contribute to U. S. competitiveness in the electrofusion marketplace, which is increasing challenged by instrument manufacturers in Europe and Japan.Awardee's statement of the potential commercial applications of the research:In Phase II, we will develop a flexible, full-featured electro-acoustic fusion instrument prototype. The instrument would allow users to efficiently create somatic cell hybrids in physiological media directly in culture plates, thus avoiding trauma attendant to chemical and electrofusion methods.National Institute of Allergy and Infectious Diseases (NIAID)
