Methods in Immunolocalization of Autoantigens

  • Martin Blüthner
Part of the Springer Lab Manual book series (SLM)

Abstract

The immune system recognizes and eliminates foreign antigens by a complex network of systems. To effectively fulfill this task the immune system has to discriminate between foreign and self antigens. The mechanisms by which this discrimination is accomplished shall not be discussed here (there are numerous excellent textbooks on immunology available). However, in autoimmune disorders this discrimination fails to a certain extent due to mechanisms currently only vaguely understood. Linked with these diseases is the occurrence of circulating autoantibodies. These are antibodies directed against self antigens of a defined nature. Autoantibodies may have a direct effect on the etiology of the disease, such as in Grave’s disease. Here, the target antigen of the autoimmune response is the acetylcholine receptor of the motoric endplate; this leads to a severe disturbance in the nervous system. In other cases, the so-called systemic autoimmune disorders, the connection between etiology and target antigen is not obvious. To what extent these auto antibodies are involved in the pathogenesis of the corresponding disease is not exactly known. In the systemic diseases the target antigen commonly resides within the cell, especially within the cell nucleus.
Table 7.1.

Features of some of the most important nuclear autoantigensa

Antigen

Molecular identity

Disease

Indirect cytoimmunofluorescence

Western blot

SM

RNP

Scl-70

CENP

Fibrillarin

PM/Scl

Ro (SSA)

La (SSB)

AMA

Core proteins of snRNP’s

U1-snRNP

Topoisomerase I

Centromere-associated proteins CENP-A, B, C

Protein component of U3-snoRNP

Nucleolar particle of unknown function

Protein component of hY-RNP

RNA-polymerase III associated factor

Subunits of mitochondrial 2-oxo-acid dehydrogenase complex

SLE

SLE overlap

Scleroderma

Scleroderma

Scleroderma

Polymyositis scleroderma overlap

Sjögren’s syndrome

Sjögren’s syndrome

PBC

Speckled nuclear staining no staining of nucleoli

Similar to Sm, occasionally more granular

Fine speckled nuclear staining, frequent staining of the nucleolus

Centromere staining

Clumpy, nucleolar staining

Homogeneous, nucleolar staining

Fine, granular nuclear staining

Fine, granular nuclear staining

Large, granular cytoplasmic staining frequently accompanied by multiple nuclear speckles (sp100 antigen)

B-protein (26–20 kDa)

70 kDa 30 kDa proteins

100 kDa protein (degradation product of 70 kDa)

CENP-A (19.5 kDa) CENP-B (80 kDa)

36 kDa protein

100 kDa, 75 kDa proteins some smaller proteins

60 kDa, 52 kDa proteins

48 kDa protein

70–74 kDa protein frequently 100 kDa (sp100 antigen)

Keywords

Cellulose Vortex Sucrose Acetone Foam 

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References

  1. Blüthner M, Bautz FA(1992) Cloning and characterization of the cDNA coding for a polymyositis-scleroderma overlap syndrome-related nucleolar 100 kDa protein. J Exp Med 176: 973–980Google Scholar
  2. Gey GD, Coffman WD, Kubicek MD (1952) Tissue culture studies of the proliferative capacity of cervical carcinoma and normal epithelium. Cancer Res 12: 264–265Google Scholar
  3. Guldner HH, Lakomek HJ, Bautz FA (1983) Identification of human Sm and ( U1) RNP antigens by immunoblotting. J Immunol Meth 64: 45–59CrossRefGoogle Scholar
  4. Kalckar HM (1947) Differential spectrophotometrie of purine compounds by means of specific enzymes. III. Studies of the enzymes of purine metabolism. J Biol Chem 167: 461–475PubMedGoogle Scholar
  5. Krohne GR, Stick R, Kleinschmidt J, Moll R, Franke WW, Hausen P (1982) Immunological localization of a major karyoskeletal protein in nucleoli of oocytes and somatic cells of Xenopus laevis. J Cell Biol 94: 749–754PubMedCrossRefGoogle Scholar
  6. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685PubMedCrossRefGoogle Scholar
  7. Olmsted JB (1981) Affinity purification of antibodies from diazotized paper blots of heterogeneous protein samples. J.Biol.Chem. 256: 11955–11957PubMedGoogle Scholar
  8. Smith DE, Fisher PA (1984) 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. J Cell Biol 99: 20–28PubMedCrossRefGoogle Scholar
  9. Tan EM (1989) Antinuclear antibodies: Diagnostic markers for autoimmune diseases and probes for cell biology. Adv.Immunol 44: 93–151PubMedCrossRefGoogle Scholar
  10. Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci USA 76: 4350–4354PubMedCrossRefGoogle Scholar
  11. van Venrooij WJ, Maini RN (1994) Manual of biological markers of disease. Kluwer Academic, DordrechtGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1998

Authors and Affiliations

  • Martin Blüthner
    • 1
  1. 1.Institute of Molecular GeneticsUniversity of HeidelbergHeidelbergGermany

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