Chromosome Research

, Volume 8, Issue 1, pp 37–44 | Cite as

Early Phase Karyotype Analysis of Chromosome Segregation After Formation of Mouse–Mouse Hybridomas with Chromosome Painting Probes

  • Leo Wollweber
  • Hiltrud Münster
  • Sabine Hoffmann
  • Käthe Siller
  • Karl Otto Greulich

Abstract

FISH analysis with chromosome painting probes allows, better than karyotyping after Giemsa banding, the study of chromosome segregation after hybridoma formation. FISH is particularly useful for intraspecies hybrids and allows visualization of small chromosome fragments. Cell hybrids were constructed between P3 × 63Ag8.653 mouse myeloma cells and lymphocytes from BALB/c mice by PEG fusion and by selection in hypoxanthine–azaserine medium. Three hybridomas (A4, D8, F10) were selected and, after cloning, the cells were cultivated in vitro over a period of 28 days. During this time in culture, air-dried metaphase spreads were prepared by standard methods. For FISH chromosome painting, digoxigenin- and biotin-labeled mouse chromosome painting probes and rhodamine–antidigoxigenin antibodies and fluorescein–avidin were used for dual color detection. Total chromosome numbers and the numbers of mouse chromosomes 1, X, 6 and 12 were estimated as function of days in culture. Mean chromosome numbers of 78 (D8), 82 (F10) and 150 (A4) were observed. The major rearrangements of chromosome numbers occured in the first 28 days in culture and did not change significantly between day 28 and day 56. Mouse chromosome #12, which had the largest chromosome fragments in the parent myeloma, remained stable while the number of X chromosomes, which were significantly fragmented already in the parent myeloma, decreased by approximately 50%.

chromosome number chromosome painting FISH hybridomas mouse 

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References

  1. Abbott C, Povey S (1995) Somatic Cell Hybrids. Oxford: IRL Press, p 25.Google Scholar
  2. Carter NP (1994) Cytogenetic analysis by chromosome painting. Cytometry 18: 2–10.PubMedGoogle Scholar
  3. Castillo FJ, Mullen LJ, Grant BC et al. (1994) Hybridoma stability. Dev Biol Stand 83: 55–64.PubMedGoogle Scholar
  4. Chen TR (1979) Cytogenetics of somatic cell hybrids. I. Progression of stemlines in continuous uncloned cultures of man-mouse cell hybrids. Cytogenet Cell Genet 23: 221–230.PubMedGoogle Scholar
  5. Cieplinski W, Reardon P, Testa MA (1983) Non-random human chromosome distribution in human-mouse myeloma somatic cell hybrids. Cytogenet Cell Genet 35: 93–99.PubMedGoogle Scholar
  6. Cowell JK (1984) A photographic representation of the variability in the G-banded structure of the chromosomes in the mouse karyotype. A guide to the identification of the individual chromosomes. Chromosoma 89: 294–320.PubMedGoogle Scholar
  7. Cremer T, Lichter P, Borden J, Ward DC, Manuelidis L (1988) Detection of chromosome aberrations in metaphase and interphase tumor cells by in situ hybridization using chromosome-specific library probes. Hum Genet 80: 235–246.PubMedGoogle Scholar
  8. Croce CM, Shander M, Martinis J, Cicurel L, D'Ancona GG, Koprowski H (1980) Preferential retention of human chromosome 14 in mouse 3 human B cell hybrids. Eur J Immunol 10: 486–488.Google Scholar
  9. Ekong R, Wolfe J (1998) Advances in fluorescent in situ hybridisation. Curr Opin Biotech 9: 19–24.PubMedGoogle Scholar
  10. Gardner JS, Chiu AL, Maki NE, Harris JF (1985) A quantitative stability analysis of human monoclonal antibody production by heteromyeloma hybridomas, using an immunofluorescent technique. J Immunol Meth 85: 335–346.Google Scholar
  11. Kearney JF, Radbruch A, Liesegang B, Rajewsky K (1978) A new mouse myeloma cell line that has lost immunoglobulin expression but permits the construction of antibody-secreting hybrid cell lines. J Immunol 123: 1548–1550.Google Scholar
  12. Köhler G (1980) Immunoglobulin chain loss in hybridoma lines. Proc Natl Acad Sci USA 77: 2197–2199.PubMedGoogle Scholar
  13. Kontsek P, Novak M, Kontsekova E (1988) Karyotype analysis of hybridomas producing monoclonal antibodies against different antigens. Folia Bio (Praha) 34: 99–104.Google Scholar
  14. Lichter P (1997) Multicolor FISHing: what's the catch? Trends Genet 13: 475–479.CrossRefPubMedGoogle Scholar
  15. Lichter P, Cremer T, Borden J Manuelidis L, Ward DC (1988) Delineation of individual human chromosomes in metaphase cells by in situ suppression hybridization using recombinant DNA libraries. Hum Genet 80: 224–234.PubMedGoogle Scholar
  16. Liyanage M, Coleman A, du Manoir S et al. (1996) Multicolour spectral karyotyping of mouse chromosomes. Natl Genet 14: 312–315.Google Scholar
  17. Matsuta Y, Chapman VM (1995) Application of fluorescence in situ hybridization in genome analysis of the mouse. Electrophoresis 16: 261–272.PubMedGoogle Scholar
  18. Miller OJ, Miller DA, Dev VG, Tantravahi R, Croce CM (1976) Expression of human and suppression of mouse nucleolus organizer activity in mouse-human somatic cell hybrids. Proc Natl Acad Sci USA 73: 4531–4535.PubMedGoogle Scholar
  19. Norum RA, Migeon BR (1974) Non-random loss of human markers from man-mouse somatic cell hybrids. Nature 251: 72–74.PubMedGoogle Scholar
  20. Nowak JS (1985) Loss of antibody production accompanied by chromosome loss in a cloned hybrid line secreting antibodies to sheep red blood cells. Experientia 41: 88–89.PubMedGoogle Scholar
  21. Pinkel D Straume T, Gray JW (1986) Cytogenetic analysis using quantitative, high-sensitivity, fluorescence hybridization. Proc Natl Acad Sci USA 83: 2934–2938.PubMedGoogle Scholar
  22. Potter M (1983) Immunoglobulin and immunoglobulin genes. In Foster HL, Small JD, Fox JG, eds. The Mouse in Biomedical Research, Vol. III. New York: Academic Press, pp 347–380.Google Scholar
  23. Ried T, Schröck E, Ning Y, Wienberg J (1998) Chromosome painting: a useful art. Hum Mol Genet 7: 1619–1626.PubMedGoogle Scholar
  24. Rodova MA, Tsoi LA, Kushch AA, Novokhatskii AS (1985) Karyological analysis of hybridoma lines producing monoclonal antibodies to viral antigens. Tsitol Genet 19: 425–428.PubMedGoogle Scholar
  25. Rushton AR (1976) Quantitative analysis of human chromosome segregation in man-mouse somatic cell hybrids. Cytogenet Cell Genet 17: 243–253.PubMedGoogle Scholar
  26. Schröder J, Autio K, Jarvis JM, Milstein C (1980) Chromosome segregation and expression of rat immunoglobulins in rat/mouse hybrid myelomas. Immunogenetics 10: 125–131.PubMedGoogle Scholar
  27. Schröder J, Sutinen ML, Suomalainen HA (1981) Chromosome segregation in lymphocyte hybrids. Hereditas 94: 77–82.PubMedGoogle Scholar
  28. Shi YP, Mohapatra G, Miller J et al. (1997) FISH probes for mouse chromosome identification. Genomics 45: 42–47.PubMedGoogle Scholar
  29. Volgareva GM (1985) Karyologic research on murine B-cell hybridomas. Tsitologiia 27: 1394–1403.PubMedGoogle Scholar
  30. Wollweber L, Fritzke H, Ozegowski J-H, Gerlach D, Köhler W (1994) Production and partial characterization of monoclonal antibodies against erythrogenic toxins type A and C from Streptococcus pyogenes. Hybridoma 13: 403–408.PubMedGoogle Scholar
  31. Zhil'tsova MA, Trofimova MN, Novikov VV (1989) Karyological analysis of hybridoma cells after prolonged cultivation. Zh Mikrobiol Epidemiol Immunobiol 6: 99–102.PubMedGoogle Scholar

Copyright information

© Kluwer Academic Publishers 2000

Authors and Affiliations

  • Leo Wollweber
    • 1
  • Hiltrud Münster
    • 2
  • Sabine Hoffmann
    • 2
  • Käthe Siller
    • 2
  • Karl Otto Greulich
    • 2
  1. 1.Institut für Molekulare BiotechnologieJenaGermany
  2. 2.Institut für Molekulare BiotechnologieJenaGermany

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