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. 1982 Aug 11;10(15):4703–4713. doi: 10.1093/nar/10.15.4703

Production and characterization of monoclonal antibody against 10S DNA polymerase alpha from calf thymus.

S Masaki, H Shiku, T Kaneda, O Koiwai, S Yoshida
PMCID: PMC321123  PMID: 7133996

Abstract

One hybridoma cell line that produces an antibody directed against 10S DNA polymerase alpha purified from calf thymus was obtained. The monoclonality of the antibody was tested by sodium dodecyl sulfate polyacrylamide gel electrophoresis, isoelectrofocusing and antibody subclass determination. The antibody specifically recognized the 10S DNA polymerase alpha and 6.5S DNA polymerase alpha-2 from calf thymus, but not 6.5S DNA polymerase alpha-1. The antibody precipitated both polypeptides of 140-150,000 and 46-50,000 dalton of 10S DNA polymerase alpha. The antibody also recognized the DNA polymerase alpha purified from human cells, but did pig DNA polymerase alpha only partially. The antibody did not crossreact with rat DNA polymerase alpha, calf DNA polymerase beta, virus DNA polymerase and E. coli DNA polymerase I. This antibody will be a useful tool for studying the mechanism of DNA replication in eukaryotic cells.

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Selected References

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  1. Dippold W. G., Lloyd K. O., Li L. T., Ikeda H., Oettgen H. F., Old L. J. Cell surface antigens of human malignant melanoma: definition of six antigenic systems with mouse monoclonal antibodies. Proc Natl Acad Sci U S A. 1980 Oct;77(10):6114–6118. doi: 10.1073/pnas.77.10.6114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Galfre G., Howe S. C., Milstein C., Butcher G. W., Howard J. C. Antibodies to major histocompatibility antigens produced by hybrid cell lines. Nature. 1977 Apr 7;266(5602):550–552. doi: 10.1038/266550a0. [DOI] [PubMed] [Google Scholar]
  3. Kaneda T., Kuroda S., Koiwai O., Yoshida S. Purification of terminal deoxynucleotidyl transferase from pig thymus: identification of 42,000 and 57,000 dalton species. J Biochem. 1981 Nov;90(5):1421–1427. doi: 10.1093/oxfordjournals.jbchem.a133608. [DOI] [PubMed] [Google Scholar]
  4. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  5. Koiwai O., Masaki S., Yoshida S. Stimulating factor for calf thymus DNA polymerase beta. Biochem Biophys Res Commun. 1981 Dec 15;103(3):990–996. doi: 10.1016/0006-291x(81)90907-4. [DOI] [PubMed] [Google Scholar]
  6. Köhler G., Milstein C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature. 1975 Aug 7;256(5517):495–497. doi: 10.1038/256495a0. [DOI] [PubMed] [Google Scholar]
  7. LITTLEFIELD J. W. SELECTION OF HYBRIDS FROM MATINGS OF FIBROBLASTS IN VITRO AND THEIR PRESUMED RECOMBINANTS. Science. 1964 Aug 14;145(3633):709–710. doi: 10.1126/science.145.3633.709. [DOI] [PubMed] [Google Scholar]
  8. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  9. Masaki S., Yoshida S. 10-S DNA polymerase from calf thymus which copies both poly(rA) . oligo(dT) and activated DNA. Biochim Biophys Acta. 1978 Nov 21;521(1):74–88. doi: 10.1016/0005-2787(78)90250-2. [DOI] [PubMed] [Google Scholar]
  10. Nakamura H., Morita T., Yoshida S. Differential inhibition of mammalian DNA polymerases by X-irradiated DNA. Int J Radiat Biol Relat Stud Phys Chem Med. 1980 Oct;38(4):449–458. doi: 10.1080/09553008014551811. [DOI] [PubMed] [Google Scholar]
  11. TAnaka M., Yoshida S. Altered sensitivity to 1-beta-D-arabinofuranosylcytosine 5'-triphosphate of DNA polymerase alpha from leukemic blasts of acute lymphoblastic leukemia. Cancer Res. 1982 Feb;42(2):649–653. [PubMed] [Google Scholar]
  12. Yoshida S., Masaki S., Koiwai O. Further characterization of a poly(rA) . oligo(dT)-dependent activity of multiple DNA polymerase alpha from calf thymus. Biochim Biophys Acta. 1981 Jul 27;654(2):194–200. doi: 10.1016/0005-2787(81)90172-6. [DOI] [PubMed] [Google Scholar]

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