Establishment of a Tetraploid, Immunoglobulin-Producing Cell Line from the Hybridization of Two Human Lymphocyte Lines

Abstract

We here report the establishment of a seemingly permanent hybrid cell line formed by fusion of the cells of two biochemically mutant human lymphocyte lines. One parental line (UM-1-6TGr) was deficient in hypoxanthine-guanine phosphoribosyl transferase (IMP: pyrophosphate phosphoribosyltransferase, EC 2.4.2.8), and had two marker chromosomes. The second parental line (UM-21-5) was a clonal derivative of a citrullinemic lymphocyte line, and was, like the line of origin, dificient in argininosuccinic acid synthetase [_L-Citrulline: _L-aspartate ligase (AMP-forming), EC 6.3.4.5]. This line also had a marker chromosome, which was a B5 with a very prominent secondary constriction. After trypsinization of both parental lines, followed by addition to the fusion mixture of β-propiolactone-inactivated Sendai virus, the cells were placed in a doubly selective medium (hypoxanthine-aminopterin-thymidine-containing medium in which the arginine was replaced with citrulline) to prevent the proliferation of the mutant parents. Under selective conditions, 97-99% of cells were found to be tetraploid, containing the three marker chromosomes; and the specific activities of the hybrid line transferase and synthetase were intermediate between normal and mutant line values. Furthermore, the UM-1-6TGr and UM-21-5 lines were producers of gamma and mu heavy chains of immunoglobulin, and of kappa light chains, as determined by immunodiffusion and immunofluorescence, and the hybrid line continued to synthesize and to secrete detectable levels of these same immunoglobulins. These studies demonstrate the genic and cytogenetic stability of this hybridized lymphocyte cell line, and prove that hybridization per se does not extinguish the activity of either the regulatory of structural genes involved in immunoglobulin synthesis.