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A new apparatus was constructed which enables the use of the electrofusion method to obtain polynuclear cells of various mammalian cell lines, erythrocytes and plant protoplasts. This technique was applied to both suspensions and monolayers. Electrical and other physical parameters were monitored to find optimal conditions for mutual contact of cells (dielectrophoresis) and subsequent fusion. In the suspension technique, dielectrophoresis of mouse erythrocytes occurred at a field frequency of 20 kHz and a strength of 500 V.cm-1, whereas cultured mammalian cells and plant protoplasts required a frequency of 1-1.4 MHz and a strength of 250-800 V.cm-1. Fusion of cells was induced after the application of 1 to 10 high-voltage pulses of 1-5 kV.cm-1, 10-36 microseconds duration. After these high-voltage pulses were to the monolayer of mouse L cells, about 12% viable homokaryons were obtained.

Hybrid cells (HY SS2 and HY SS6) arising from the fusion of diploid cells of the mouse lymphosarcoma LS/BL and L cells resistant to 8-azaguanine (HGPRT-) showed slower growth and a longer generation time than the parent lines. The inter- and intrachromosomal timing and patterns of early chromosome DNA replication of parent cells was preserved in the hybrid genome and was not influenced by loss of telocentric chromosomes from LS/BL or L (HGPRT-) cells. Thus DNA chromosome replication sequences are not dependent on the presence of a complete set of chromosomes of the parent cells and do not therefore seem to be a result of interaction between chromosomes not segregated in the hybrid genome.

Incorporation of isologous 3H-DNA of exogenous origin into chromosomes of the L cells was investigated by means of autoradiographical technique. It was shown that during short time intervals of incubation of 3H-DNA with the host cells (1--3 hr), nearly double quantity of the mitoses was labelled, when compared with 3H-TdR used as a control. During longer time intervals, no difference between number of the mitoses labelled with 3H-TdR and exogenous isologous 3H-DNA was observed.

Ninety-six clonal populations were derived from a wild mouse neuroblastoma cell population C 1300 in a serum-free medium containing commercially available serum growth-promoting proteins (GPP). From among these 96 lines the clonal population E 7 was chosen for further work because it displayed maximum spontaneous morphological differentiation. The neuroblastoma clonal population differs morphologically from the original population; it was defined both cytogenetically and by means of growth parameters. The cells of the neuroblastoma clone E 7 are hypertetraploid with two chromosome number modals - 88 and approximately 180-200. The majority of telocentric chromosomes in metaphases with a modal number of 88 chromosomes are identical with the chromosomes of mouse diploid cells. The cell generation time is 22 hours. The cells of the clonal population E 7 are highly sensitive to the action of ethanolamine, which induces morphological differentiation, so that the processes of 30% of the cells in the population are over 40 micron long. Electrophysiological studies showed that the cells of the neuroblastoma clonal population E 7 retain the character of excitable cells and they are thus suitable for studying some of the properties of nervous tissue cells.

The initial and terminal patterns of DNA replication in the chromosomes of the hybrids arising from a fusion of mouse lymphosarcoma LS/BL cells and L cells (HGPRT-) were studied. The replication of chromosomes in the hybrids compared to the parent cells was not modified by the alterations in generation time of the hybrids not by segregation of telocentric chromosomes. It can be assumed that there is a gross control system in LS/BL X L hybrids that is responsible for the patterns and timing of chromosome DNA replication and does not depend on interactions between genomes of the parent cells.

The karyotype of HY 5/3--4 hybrid cells formed by fusion of diploid and heteroploid murine cells (LS/BLxR-AG/20) consisted of 60--75 chromosomes (modal number 72). The G-banding technique made it possible to identify the origin of the chromosomes; it is concluded that due to chromosomal segregation the hybrid genome lost preferentially the chromosomes derived from the diploid parental line LS/BL. Even after long-term culture in vitro, the chromosomal complement of hybrid cells retained all bi-armed chromosomes and most of the telocentrics, derived from the heteroploid parental cells.

The autoradiographic investigation of L cells and chinese hamster cells for the presence of mycoplasmas (A. laidlawii and M. hyorhinis) using uridine/uracil (UdR/U) testing is a rapid and reliable method suitable for the serial checking of even a small number of cells. It depends on a reduced incorporation of [3H]uridine and an increased uptake of [3H]uracil into the RNA of mycoplasma-infected cells, shown in autoradiograms by the density of the grains and their distribution. Results obtained by the autoradiographic technique correspond approximately to specific activity values of RNA-infected cells after the incorporation of [3H]uridine and [3H]uracil.

Chinese hamster cell line (line K 431, HGPRT+) and its mutant (line K II, HGPRT-) are similar in the numberical composition of the karyotype (modal number 22 chromosomes), but differ in its structureal constitution (occurrence of trisomy, breaks in the centromeric region, chromosome losses and others). Deletion and rearrangement of some heterochromatic segments of the X chromosome appeared with a higher frequency in HGPRT- line and can constitute one of the conceivable causes of the HGPRT activity loss in the mutant line.