Abstract
Polyacrylamide gel electrophoresis has proved to be an extremely powerful tool for the analysis of complex mixtures of proteins (see Chapter 2). However, while the resolving power of this method has progressively increased (the two-dimensional gel system of O’Farrell1 for example being capable of resolving more than 1600 different proteins) any further analysis of the separated proteins has been limited. The majority of the protein molecules lie buried within the gel matrix and are therefore not readily accessible to further investigation. Though methods have been developed to overcome this problem (for example by elution of the proteins2 or by direct in-situ analysis using antisera3,4 or other protein probes5) these methods are in general time-consuming, insensitive and also frequently lead to a loss of resolution.
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Further Reading
Schleicher and Schuell Inc., USA. Methods for the transfer of DNA, RNA and protein to nitrocellulose and diazotized paper solid supports. By: Barinaga. M. et al., 1981.
Bio-Rad Laboratories.’ southern, Western and Electroblotting’, Bulletin 1080 EG, (June 1981).
References
P.H. O’Farrell and P.Z. O’Farrell, ‘Two dimensional polyacrylamide gel electrophoretic fractionation’ in G. Stein. J. Stein and L.J. Kleinsmith, Chromatin and chromosomal protein research, Methods in Cell Biology 16. (Academic Press, New York, London, Toronto, Sydney. San Francisco, 1977), Part 1, 407–420.
M. Bustin, R.B. Hopkins and I. Isenberg, ‘Immunological relatedness of high mobility group chromosomal proteins from calf thymus’, J. Biol. Chem., 253 (1978), 1694–1699.
W.S. Adari, D. Jurivich and U.W. Goodenough, ‘Localization of cellular antigens in sodium dodecylsulphate-polyacrylamide gels’, J. Cell Biol., 79 (1978), 281–285.
K. Burridge, ‘Changes in cellular glycoproteins after transformation: Identification of specific glycoproteins and antigens in sodium dodecyl sulphate gels’, Proc. Natl. Acad. Sci. USA, 73 (1976), 4457–4461.
M.C. Snabes, A.E. Boyd III and J. Bryan, ‘Detection of actin-binding proteins in human platelets by 125I actin overlay of polyacrylamide gels’, J. Cell Biol., 90 (1981), 809–812.
E.M. Southern, ‘Detection of specific sequences among DNA fragments separated by gel electrophoresis’, J. Mol. Biol., 98 (1975), 503–517.
Ibid.
B. Bowen. J. Steinberg. U.K. Laemmli and H. Weintraub, ‘The detection of DNA-binding proteins by protein blotting’, Nucleic Acids Res., 8 (1980), 1–20.
H. Towbin, T. Stahelin and J. Gordon, ‘Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets. Procedure and some applications’, Proc. Natl. Acad. Sci. USA. 76 (1979), 4350–4354.
Ibid.
W.N. Burnett, ‘“Western blotting”. Electrophoretic transfer of proteins from SDS-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A’, Anal. Biochem., 112 (1981), 195–203.
W. Gibson, ‘Protease-facilitated transfer of high-molecular-weight proteins during electrotransfer to nitrocellulose’, Anal. Biochem., 118 (1981), 1–3.
D.W. Cleveland, S.G. Fischer, M.W. Kirschner and U.K. Laemmli, ‘Peptide mapping by limited proteolysis in sodium dodecyl sulphate and analysis by gel electrophoresis’, J. Biol. Chem., 252 (1977), 1102–1106.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
M. Bittner, P. Kupferer and C.F. Morris, ‘Electrophoretic transfer of proteins and nucleic acids from slab gels to diazobenzyloxymethyl cellulose or nitrocellulose sheets’, Anal. Biochem., 102 (1980), 459–471.
T.P. St John and R.W. Davis, ‘Isolation of galactose-inducible DNA sequences from Saccharomyces cerevisiae by differential plaque filter hybridisation’, Cell, 16 (1979), 443–452.
Burnette, “Western blotting”.
J.C. Alwine, D.J. Kemp and G.R. Stark, ‘Method for detection of specific RNAs in agarose gels by transfer to diazobenzyloxymethyl paper and hybridization with DNA probes’, Proc. Natl Acad. Sci. USA, 74 (1977), 5350–5354.
J. Renart, J. Reiser and G.R. Stark, ‘Transfer of proteins from gels to diazobenzyloxymethyl paper and detection with antisera: A method for studying antibody specificity and antigen structure’, Proc. Natl. Acad. Sci. USA, 76 (1979), 3116–3120.
J. Symington, M. Green and K. Brackman, ‘Immunoautoradiographic detection of proteins after electrophoretic transfer from gels to diazo-paper: Analysis of adenovirus encoded proteins’, Proc. Natl. Acad. Sci. USA, 78 (1981), 177–181.
L. Clarke, R. Hitzeman and J. Carbon, ‘Selection of specific clones from colony banks by screening with radioactive antibody’, Methods in Enzymology, (Academic Press, New York, London), 68 (1979), 436–442.
B.S. Bhullar, J. Hewitt and E.P.M. Candido, ‘The large high mobility group proteins of rainbow trout are localized predominantly in the nucleus and nucleoli of a cultured trout cell line’, J. Biol. Chem., 256 (1981). 8801–8806.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
Towbin, Stahelin and Gordon, ‘Electrophoretic transfer of proteins’.
Ibid.
Burnette, “Western blotting”.
Bittner, Kupferer and Morris, ‘Electrophoretic transfer of proteins’.
D. Denharft, ‘A membrane-filter technique for the detection of complementary DNA’, Biochem. Biophys. Res. Commun., 23 (1966), 641–646.
Towbin, Stahelin and Gordon, ‘Electrophoretic transfer of proteins’.
Burnette, “Western blotting”.
V. Shen, T.C. King, V. Kumar and B. Daugherty, ‘Monoclonal antibodies to Escherichia coli 50S ribosomes’, Nucleic Acids Res., 8 (1980), 4639–4649.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
R.S. Gilmour, A. Lang, J.R. Gu, C. Johnston and J. Paul, DNA binding proteins in chromatin structure and function’, Cell Biol. Int. Repts., 5 (1981), 45–53.
Bowen, Steinberg. Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
Gilmour, Lang, Gu, Johnston and Paul, ‘DNA-binding proteins’.
Burnette, “Western blotting”.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
H.A. Erlich, J.R. Levinson, S.N. Cohen and H.O. McDevitt, ‘Filter affinity transfer. A new technique for the in-situ identification of proteins in gels’, J. Biol. Chem., 254 (1979), 12240–12247.
Burnette, “Western blotting”.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
Erlich, Levinson, Cohen and McDevitt, ‘Filter affinity transfer’.
Towbin, Stahelin and Gordon, ‘Electrophoretic transfer of proteins’.
Burnette, ‘“Western blotting”’.
Symington, Green and Brackman, ‘Immunoautoradiographic detection of proteins’.
Bhullar, Hewitt and Candido, ‘The large high mobility group proteins’.
Erlich, Levinson, Cohen and McDevitt, ‘Filter affinity transfer’.
Burnette, ‘“Western blotting”’.
Renart, Reiser and Stark, ‘Transfer of proteins’.
Bhullar, Hewitt and Candido, ‘The large high mobility group of proteins’.
Towbin, Stahelin and Gordon, ‘Electrophoretic transfer of proteins’.
Erlich, Levinson, Cohen and McDevitt, ‘Filter affinity transfer’.
Towbin, Stahelin and Gordon, ‘Electrophoretic transfer of proteins’.
Ibid.
R.T.M.J. Vaessen, J. Kreike and G.S.P. Groot, ‘Protein transfer to nitrocellulose filters’, FEBS Letts., 124 (1981), 193–196.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
Gilmour, Lang, Gu, Johnston and Paul, ‘DNA-binding proteins’.
Bowen, Steinberg, Laemmli and Weintraub, ‘The detection of DNA-binding proteins’.
Ibid.
Erlich, Levinson, Cohen and McDevitt, ‘Filter affinity transfer’.
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© 1983 John M. Walker and Wim Gaastra
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Gooderham, K. (1983). Protein Blotting. In: Walker, J.M., Gaastra, W. (eds) Techniques in Molecular Biology. Springer, Dordrecht. https://doi.org/10.1007/978-94-011-6563-1_3
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DOI: https://doi.org/10.1007/978-94-011-6563-1_3
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