Identification, developmental regulation, and response to heat shock of two antigenically related forms of a major nuclear envelope protein in Drosophila embryos: application of an improved method for affinity purification of antibodies using polypeptides immobilized on nitrocellulose blots - PubMed (original) (raw)

Identification, developmental regulation, and response to heat shock of two antigenically related forms of a major nuclear envelope protein in Drosophila embryos: application of an improved method for affinity purification of antibodies using polypeptides immobilized on nitrocellulose blots

D E Smith et al. J Cell Biol. 1984 Jul.

Abstract

An affinity-purification method has been developed for the rapid, efficient, and precise elution of antibodies specifically bound to antigens immobilized on nitrocellulose after blot transfer from SDS polyacrylamide gels. The applicability of this technology has been demonstrated using antisera raised against the nuclear matrix-pore complex-lamina fraction prepared from Drosophila melanogaster embryos. In so doing, we have established the existence in whole embryo lysates, of two nearly identical forms of the predominant 74-kilodalton polypeptide previously identified in lower resolution studies of the nuclear matrix-pore complex-lamina fraction. These species, distinguishable on the basis of a slight difference in SDS PAGE mobilities on low concentration polyacrylamide gels, are immunochemically cross-reactive and have been localized exclusively to the nuclear periphery (nuclear envelope) by indirect immunofluorescence analyses of cryosections. The steady-state levels of these two polypeptides have been examined in total embryo lysates both as a function of embryogenesis and in response to heat shock. The larger species is not detectable in early embryos but approaches levels approximately equal to that of the smaller form by about the temporal midpoint of embryonic development. In response to heat shock, this larger form appears to be converted nearly quantitatively into the lower molecular weight polypeptide. These results, as well as the general reliability of the nitrocellulose blot immunoaffinity-purification methodology, have been substantiated through the use of monoclonal antibodies.

PubMed Disclaimer

Similar articles

• Identification of a major polypeptide of the nuclear pore complex.
  Gerace L, Ottaviano Y, Kondor-Koch C. Gerace L, et al. J Cell Biol. 1982 Dec;95(3):826-37. doi: 10.1083/jcb.95.3.826. J Cell Biol. 1982. PMID: 7153248 Free PMC article.
• A myosin heavy-chain-like polypeptide is associated with the nuclear envelope in higher eukaryotic cells.
  Berrios M, Fisher PA. Berrios M, et al. J Cell Biol. 1986 Sep;103(3):711-24. doi: 10.1083/jcb.103.3.711. J Cell Biol. 1986. PMID: 2943745 Free PMC article.

Cited by

• The 46/50 kDa phosphoprotein VASP purified from human platelets is a novel protein associated with actin filaments and focal contacts.
  Reinhard M, Halbrügge M, Scheer U, Wiegand C, Jockusch BM, Walter U. Reinhard M, et al. EMBO J. 1992 Jun;11(6):2063-70. doi: 10.1002/j.1460-2075.1992.tb05264.x. EMBO J. 1992. PMID: 1318192 Free PMC article.
• cpc-1, the general regulatory gene for genes of amino acid biosynthesis in Neurospora crassa, is differentially expressed during the asexual life cycle.
  Ebbole DJ, Paluh JL, Plamann M, Sachs MS, Yanofsky C. Ebbole DJ, et al. Mol Cell Biol. 1991 Feb;11(2):928-34. doi: 10.1128/mcb.11.2.928-934.1991. Mol Cell Biol. 1991. PMID: 1824959 Free PMC article.
• cDNA and amino acid sequences of the cell adhesion protein receptor recognizing vitronectin reveal a transmembrane domain and homologies with other adhesion protein receptors.
  Suzuki S, Argraves WS, Pytela R, Arai H, Krusius T, Pierschbacher MD, Ruoslahti E. Suzuki S, et al. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8614-8. doi: 10.1073/pnas.83.22.8614. Proc Natl Acad Sci U S A. 1986. PMID: 2430295 Free PMC article.
• Production of the CYS3 regulator, a bZIP DNA-binding protein, is sufficient to induce sulfur gene expression in Neurospora crassa.
  Paietta JV. Paietta JV. Mol Cell Biol. 1992 Apr;12(4):1568-77. doi: 10.1128/mcb.12.4.1568-1577.1992. Mol Cell Biol. 1992. PMID: 1532230 Free PMC article.

References

1. 1. EMBO J. 1983;2(3):469-75 - PubMed
2. 1. Immunochemistry. 1969 Jan;6(1):43-52 - PubMed
3. 1. Nature. 1970 Aug 15;227(5259):680-5 - PubMed
4. 1. Histochemie. 1970;23(2):180-4 - PubMed
5. 1. Anal Biochem. 1973 Dec;56(2):502-14 - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources

• Full Text Sources

  • Europe PubMed Central
  • Ovid Technologies, Inc.
  • PubMed Central
  • Silverchair Information Systems

• Other Literature Sources

  • The Lens - Patent Citations Database

• Molecular Biology Databases

  • FlyBase