References

Notes: A simple method for production of clones from almost all the cells of a HeLa population is descrived. The technique can be used to titrate the number of cells capable of reproduction in a suspension and should find application in genetic and metabolic studies involving the action on animal cells of viruses, drugs, nutritional stresses, and mutagenic agents.
Reference ID: 2423

Notes: Cloning of HeLa S3 from HeLa was reported.
Reference ID: 367

Notes: A tichnique developed in our laboratory for plating single mammalian cells in petri dishes containing nutrient medium permits each cell to grow in isolation to form a macroscopic colony (1-3). The method is simplicity and accuracy, to the standard agar plating procedure of quantitative bacteriology. It has been used for demonstrationof the existence of mutant human cell strains and for the isolation of clonal cell lines with stable differentiating characteristics (4). It has provided accurate and reproducible analysis of growth curves of single mammalian cells under a variety of conditions (1-3) and has made possible quantitative studies of the action of them of agents such as ionizing radiation (5), antibodies, drugs, and hormones (6).
Reference ID: 1889

Notes: Clonal strains of HeLa cells have been isolated which vary with respect to plaque-forming ability with poliovirus over a tenfold range. This phenomenon appearea to be mediated through a heritable cellular difference and is not due to a host range virus variation. The differences in susceptibility are unexplained but are not due to the quantity of virus adsorbed, even though differences in adsorption rate were detected.
Reference ID: 1890

Notes: The major advances of the science of genetics have come about through the use of two fundamental types of operations. The first and oldest consists in mating of selected, multicellular plants or animals to produce large numbers of offspring, among which the distribution of various inferited characters is examined. This procedure has produced that body of knowledge known as classical genetics and which, beginning systematically with the studies of Mendel, has delineadted the processes which sustain and modulate biological heredity in all living forms. This experimental approach, which essentially examines genetic mechanisms in the germ cells, has been extremely successful in elucidating hereditary phenomena in organisms as diverse as Drosophila and Zea mays, but is difficult to apply to the slowly and less extensively reproducing organisms like the mammals.
Reference ID: 1888

Notes: The replication site of DNA in HeLa S3 cells has been studied with "M-band" technique after Tremblay and others. When the cells are mixed with sodium lauroyl sarcosinate and Magnesium ion directly on the sucrose gradient and spun, virtually all the DNA is caught in the M-band, suggesting that mammalian cell DNA is attached to the supernatant.Pulse-labeled DNA is more trsistant to shearing as compared with the buld of DNA and chased away from the M-band during a subsequent growth. These results indicate that, as in the case of bacteria, the replication points of mammalian cell DNA are on the mambrane.
Reference ID: 1613

Notes: In this paper we describe a new technique of cloning by use of agar plates and its application to replica plating. It was found that most cell lines form colonies on the surface of solid agar, although the plating efficiency and size of colony is dependent on specimens and concentrations of agar and agarose used. When 0.5 % noble-agar was used as substrate, plating efficiencies were obtained ...................
Reference ID: 926

Note:HeLa S3 (Epitheloid carcinoma, cervix, human) Current medium for propagation: Ham's F-12 medium, 90%; FBS, 10%. HeLa S3 is a clonal derivative of the parent HeLa line (see ATCC CCL 2). S3 was cloned in 1955 by T.T. Puck, P.I. Marcus, and S.J. Cieciura (J. Exp. Med. 103: 273-284, 1956) in classical studies of great ingenuity that introduced microbiological plating techniques to the study of mammalian cell culture systems (T.T. Puck and P.I. Marcus, Proc. Natl. Acad. Sci. USA 41: 432-437, 1955). Of several HeLa clones derived at that time, S3 was found to be especially hardy and had a plating efficiency of 100% (T.T. Puck and H.W. Fisher, J. Exp. Med. 104: 427-434, 1956). After development as a clone the cells were adapted to growth in Medium N16HHF constituted as follows: Solution N16, 40%; Saline F, 30%; human serum, 20%; horse serum, 10% plus antibiotics (R.G. Ham and T.T. Puck, Methods Enzymol. 5: 90-119, 1962). Later the cells were adapted to growth in Ham's F12 medium plus 10% FBS. *The HeLa S3 clone has been very useful in the clonal analysis of mammalian cell populations relating to chromosomal variation (T.T. Puck, Rev. Mod. Phys. 31: 433-448, 1959), cell nutrition (G. Sato, H.W. Fisher, and T.T. Puck, Science (Washington, DC) 126: 961-964, 1957) and plaque-forming ability (J.E. Darnell, Jr. and T.K. Sawyer, Virology 8: 223-229, 1959). Clone S3 has also been found to be readily adaptable to growth in suspension culture for biochemical studies of viruses and cells (J.E. Darnell, Jr. et al., J. Exp. Med. 110: 445-450, 1959; E.P. Cohen and H. Eagle, J. Exp. Med. 113: 467-474, 1961). A culture in approximately the 400th passage was submitted to the American Type Culture Collection in February, 1972. *[DESCRIPTION OF REPOSITORY REFERENCE SEED STOCK] *Number of Serial Subcultures from Tissue of Origin: Unknown. Freeze Medium: Culture medium, 95%; dimethyl sulfoxide (DMSO), 5%; antibiotic-free. Viability: Approximately 90% (dye exclusion). Culture Medium: Ham's F12 medium, 90%; FBS, 10%; antibiotic-free. Growth Characteristics of Thawed Cells: An inoculum of 10(5) viable cells/ml in the above culture medium at 37C multiplies approximately 15-fold within 7 days provided the medium is renewed 3 times weekly. Plating Efficiency: Approximately 14% in the above culture medium. Morphology: Epithelial-like. Karyology: Chromosome Frequency Distribution 50 Cells: 2n = 46 *Cells: 1 1 1 1 1 5 3 6 7 9 7 3 4 1 *Chromosomes: 51-58-60-62 63 64 65 66 67 68 69-71 72-74 *A medium-sized metacentric marker is present in 100% of the cells. HeLa Markers: One copy of M1, one copy of M2, two copies of M3, and one copy of M4. Sterility: Tests for mycoplasma, bacteria, and fungi were negative. Species: Confirmed as human by immunofluorescence test. Virus Susceptibility: Susceptible to poliovirus type 1, adenovirus type 5 and vesicular stomatitis (Indiana Strain) virus. Reverse Transcriptase: Not detected. Isoenzymes: G6PD type A. Submitted by: T.T. Puck, University of Colorado Medical Center, Denver, CO. Prepared and characterized by: ATCC, Rockville, MD.
RefID: 4093

9. Macville,M., Schrock,E., Padilla-Nash,H., Keck,C., Ghadimi,B.M., Zimonjic,D., Popescu,N., and Ried,T. Comprehensive and definitive molecular cytogenetic characterization of HeLa cells by spectral karyotyping, Cancer Res, 59: 141-150, 1999.

Notes: Genome Technology Branch, National Human Genome Research Institute, NIH, Bethesda, Maryland 20892, USA mmacville@patholaznnl
Abstract: We revisited the cytogenetic alterations of the cervical adenocarcinoma cell line HeLa through the use of spectral karyotyping (SKY), comparative genomic hybridization (CGH), and fluorescence in situ hybridization (FISH). SKY analysis unequivocally characterized all abnormal chromosomes. Chromosomal breakpoints were primarily assigned by simultaneous assessment of SKY painted chromosomes and inverted 4,6-diamidino2-phenylindole banding from the same cell. Twenty clonally abnormal chromosomes were found. Comparison with previously reported HeLa G-banding karyotypes revealed a remarkably stable cytogenetic constitution because 18 of 20 markers that were found were present before. The classification of 12 markers was refined in this study. Our assignment of the remaining six markers was consistent with those described in the literature. The CGH map of chromosomal copy number gains and losses strikingly matched the SKY results and was, in a few instances, decisive for assigning breakpoints. The combined use of molecular cytogenetic methods SKY, CGH, and FISH with site-specific probes, in addition to inverted 4,6-diamidino-2-phenylindole or conventional G-banding analysis, provides the means to fully assess the genomic abnormalities in cancer cells. Human papillomaviruses (HPVs) are frequently integrated into the cellular DNA in cervical cancers. We mapped by FISH five HPV18 integration sites: three on normal chromosomes 8 at 8q24 and two on derivative chromosomes, der(5)t(5;22;8)(qll;q11q13;q24) and der(22)t(8; 22)(q24;q13), which have chromosome 8q24 material. An 8q24 copy number increase was detected by CGH. Dual-color FISH with a c-MYC probe mapping to 8q24 revealed colocalization with HPV18 at all integration sites, indicating that dispersion and amplification of the c-MYC gene sequences occurred after and was most likely triggered by the viral insertion at a single integration site. Numerical and structural chromosomal aberrations identified by SKY, genomic imbalances detected by CGH, as well as FISH localization of HPV18 integration at the c-MYC locus in HeLa cells are common and representative for advanced stage cervical cell carcinomas. The HeLa genome has been remarkably stable after years of continuous cultivation; therefore, the genetic alterations detected may have been present in the primary tumor and reflect events that are relevant to the development of cervical cancer
Reference ID: 5247