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Immunofixation Electrophoresis for Identification of Proteins and Specific Antibodies

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Electrophoretic Separation of Proteins

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1855))

Abstract

Immunofixation electrophoresis (IFE) is a technique for the identification of proteins within complex mixtures after separation by either conventional zone electrophoresis or isoelectric focusing. Most commonly antigens (which are often immunoglobulins) are separated by electrophoresis followed by precipitation with specific antibodies in situ. However, immunoglobulins with specific reactivity can be also precipitated with the proper antigens after electrophoresis in reverse or reversed IFE. Because of its great versatility, potentially high sensitivity, ease to perform and customize, and relatively low cost with no requirement for expensive instrumentation, manual IFE remains a valuable tool for both clinical diagnostic testing and research. Any low-viscosity body fluid specimen or, possibly, culture fluid could be tested with IFE if proper antibodies (or antigens in reverse[d] IFE) are available. After pretreatment with chaotropic and/or reducing agents, even high-viscosity specimens might be amenable to testing with IFE.

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References

  1. Afonso E (1964) Quantitative immunoelectrophoresis of serum proteins. Clin Chim Acta 10:114–122

    Article  CAS  Google Scholar 

  2. Wilson AT (1964) Direct immunoelectrophoresis. J Immunol 92:31–34

    Google Scholar 

  3. Alper CA, Johnson AM (1969 and 1993) Immunofixation electrophoresis: a technique for the study of protein polymorphism. Vox Sang 17:445–452, Vox Sang 65:76

    Article  CAS  Google Scholar 

  4. Johnson AM, Cleve H, Alper C (1975) Variants of the group-specific component system as demonstrated by immunofixation electrophoresis. Report of a new variant, Gc Boston (Ge B). Am J Hum Genet 27:728–736

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Johnson AM (1976) Genetic typing of alpha1-antitrypsin by immunofixation electrophoresis, identification of subtypes of Pi M. J Lab Clin Med 87:152–163

    CAS  PubMed  Google Scholar 

  6. Lieberman J, Gaidulis L (1976) Simplified alpha1-antitrypsin phenotyping by immunofixation of acid-starch gels. J Lab Clin Med 87:710–716

    CAS  PubMed  Google Scholar 

  7. Grunbaum BW, Zajac PL (1977) Rapid phenotyping of the group specific component by immunofixation on cellulose acetate. J Forensic Sci 22:586–589

    Article  CAS  Google Scholar 

  8. Cleve H, Constans J, Berg S et al (1981) Gc revisited: six further Gc-phenotypes delineated by isoelectric focusing and by polyacrylamide gel electrophoresis. Hum Genet 57:312–316

    Article  CAS  Google Scholar 

  9. Mauff G, Hummel K, Pulverer G (1975) Properdin factor B (glycine-rich beta-glycoprotein or C3 proactivator)-polymorphism: genetic and biochemical aspects. First application to paternity cases. Z Immunitatsforsch Exp Klin Immunol 150:327–338

    CAS  PubMed  Google Scholar 

  10. Arnaud P, Wilson GB, Koistinen J et al (1977) Immunofixation after electrofocusing: improved method for specific detection of serum proteins with determination of isoelectric points. I. Immunofixation print technique for detection of alpha-1-protease inhibitor. J Immunol Methods 16:221–231

    Article  CAS  Google Scholar 

  11. Baumstark JS (1978) Quantitative immunofixation of proteins following zone electrophoresis in agarose gel: application to the determination of the stoichiometry of the alpha1-antitrypsin-elastase interaction. J Immunol Methods 23:79–89

    Article  CAS  Google Scholar 

  12. Morrison R, Noppinger K, Brown MG Jr (1985) Immunofixation of complement component C3 phenotypes in bloodstains after cellulose acetate electrophoresis. J Forensic Sci 30:1221–1225

    Article  CAS  Google Scholar 

  13. Cawley LP, Minard BJ, Tourtellotte WW et al (1976) Immunofixation electrophoretic techniques applied to identification of proteins in serum and cerebrospinal fluid. Clin Chem 22:1262–1268

    CAS  PubMed  Google Scholar 

  14. Chang CH, Inglis NR (1979) Convenient immunofixation electrophoresis on cellulose acetate membrane. Clin Chim Acta 65:91–97

    Google Scholar 

  15. Ritchie RF, Smith R (1976) Immunofixation. I. General principles and application to agarose gel electrophoresis. Clin Chem 22:497–499

    CAS  PubMed  Google Scholar 

  16. Ritchie RF, Smith R (1976) Immunofixation. II. Application to typing of alpha1-antitrypsin at acid pH. Clin Chem 22:1735–1737

    CAS  PubMed  Google Scholar 

  17. Ritchie RF, Smith R (1976) Immunofixation. III. Application to the study of monoclonal proteins. Clin Chem 22:1982–1985

    CAS  PubMed  Google Scholar 

  18. Kyle RA, Greipp PR (1978) The laboratory investigation of monoclonal gammopathies. Mayo Clin Proc 53:719–739

    CAS  PubMed  Google Scholar 

  19. Johnson AM (1982) Immunofixation electrophoresis and electrofocusing. Clin Chem 28:1797–1800

    CAS  PubMed  Google Scholar 

  20. Keren DF (1999) Procedures for the evaluation of monoclonal immunoglobulins. Arch Pathol Lab Med 123:126–132

    CAS  PubMed  Google Scholar 

  21. Freedman MS, Thompson EJ, Deisenhammer F et al (2005) Recommended standard of cerebrospinal fluid analysis in the diagnosis of multiple sclerosis: a consensus statement. Arch Neurol 62:865–870

    Article  Google Scholar 

  22. Laurenzi MA, Link H (1978) Localization of the immunoglobulins G, A and M, beta-trace protein and gamma-trace protein on isoelectric focusing of serum and cerebrospinal fluid by immunofixation. Acta Neurol Scand 58:141–147

    Article  CAS  Google Scholar 

  23. Bateman N, Jones NS (2000) Rhinorrhoea feigning cerebrospinal fluid leak: nine illustrative cases. J Laryngol Otol 114:462–464

    CAS  PubMed  Google Scholar 

  24. Bleier BS, Debnath I, O'Connell BP et al (2011) Preliminary study on the stability of beta-2 transferrin in extracorporeal cerebrospinal fluid. Otolaryngol Head Neck Surg 144:101–103

    Article  Google Scholar 

  25. Ottó S (1982) Reversed immunofixation agar gel electrophoresis. Immunol Lett 4:85–86

    Article  Google Scholar 

  26. Mehta PD, Patrick BA, Thormar H et al (1982) Oligoclonal IgG bands with and without measles antibody activity in sera of patients with subacute sclerosing panencephalitis (SSPE). J Immunol 129:1983–1985

    CAS  PubMed  Google Scholar 

  27. Storstein A, Monstad SE, Honnorat J et al (2004) Paraneoplastic antibodies detected by isoelectric focusing of cerebrospinal fluid and serum. J Neuroimmunol 155:150–154

    Article  CAS  Google Scholar 

  28. Stich O, Rauer S (2007) Antigen-specific oligoclonal bands in cerebrospinal fluid and serum from patients with anti-amphiphysin- and anti-CV2/CRMP5 associated paraneoplastic neurological syndromes. Eur J Neurol 14:650–653

    Article  CAS  Google Scholar 

  29. Bossuyt X, Bogaerts A, Schiettekatte G et al (1998) Detection and classification of paraproteins by capillary immunofixation/subtraction. Clin Chem 44:760–764

    CAS  PubMed  Google Scholar 

  30. Litwin CM, Anderson SK, Philipps G et al (1999) Comparison of capillary zone and immunosubtraction with agarose gel and immunofixation electrophoresis for detecting and identifying monoclonal gammopathies. Am J Clin Pathol 112:411–417

    Article  CAS  Google Scholar 

  31. Yang Z, Harrison K, Park YA et al (2007) Performance of the Sebia CAPILLARYS 2 for detection and immunotyping of serum monoclonal paraproteins. Am J Clin Pathol 128:293–299

    Article  CAS  Google Scholar 

  32. Paquette DM, Banks PR (2001) Detection of specific antibodies using immunosubtraction and capillary electrophoresis instrumentation. Electrophoresis 22:2391–2397

    Article  CAS  Google Scholar 

  33. Pascali E, Pezzoli A, Chiarandini A (1982) Immunofixation: application to the identification of “difficult” monoclonal components. Clin Chem 28:1404–1405

    CAS  PubMed  Google Scholar 

  34. Leglise D, Menez JF, Person B et al (1982) A detailed lipidograph: enzymatic determination of cholesterol, phospholipids and glycerides in plasma lipoprotein after a cellulose acetate electrophoretic procedure (author’s transl). [Article in French]. Clin Chim Acta 118:265–277

    Article  CAS  Google Scholar 

  35. Nauck M, Winkler K, März W et al (1995) Quantitative determination of high-, low-, and very-low-density lipoproteins and lipoprotein(a) by agarose gel electrophoresis and enzymatic cholesterol staining. Clin Chem 41:1761–1767

    CAS  PubMed  Google Scholar 

  36. Winkler K, Nauck M, Siekmeier R et al (1995) Determination of triglycerides in lipoproteins separated by agarose gel electrophoresis. J Lipid Res 36:839–847

    Google Scholar 

  37. García-Sánchez C, Torres-Tamayo M, Juárez-Meavepeña M et al (2011) Lipid plasma concentrations of HDL subclasses determined by enzymatic staining on polyacrylamide electrophoresis gels in children with metabolic syndrome. Clin Chim Acta 412:292–298

    Article  Google Scholar 

  38. Cejka J, Kithier K (1979) IgD myeloma protein with “unreactive” light chain determinants. Clin Chem 25:1495–1498

    CAS  PubMed  Google Scholar 

  39. Netto D, Vladutiu AO (1981) A simple technique for identification of “unreactive” light chains of immunoglobulins. Clin Chim Acta 116:253–260

    Article  CAS  Google Scholar 

  40. Rabhi H, Ghaffor M, Abbadi MC (1989) Spontaneous enzymatic cleavage of IgD myeloma protein giving a pattern of delta heavy chain disease. Arch Inst Pasteur Alger 57:135–140

    CAS  PubMed  Google Scholar 

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Author information

Authors and Affiliations

  1. Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, MD, USA

    Gyorgy Csako

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  1. Gyorgy Csako
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Editors and Affiliations

  1. Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA

    Biji T. Kurien

  2. Department of Medicine, University of Oklahoma Health Sciences Center, Oklahoma City, OK, USA

    R. Hal Scofield

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Csako, G. (2019). Immunofixation Electrophoresis for Identification of Proteins and Specific Antibodies. In: Kurien, B., Scofield, R. (eds) Electrophoretic Separation of Proteins. Methods in Molecular Biology, vol 1855. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8793-1_17

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  • DOI: https://doi.org/10.1007/978-1-4939-8793-1_17

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8792-4

  • Online ISBN: 978-1-4939-8793-1

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