Advertisement

Autoantibodies pp 255-262 | Cite as

Determination of Autoantibodies by Line Immunoblotting

  • Louise Sternbæk
  • Tina Friis
  • Gunnar Houen
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1901)

Abstract

Autoantibody detection is a useful and obligatory tool for clinicians and researchers in diagnosing autoimmune diseases. Line blotting is a simple, sensitive, and flexible technique for fast semiquantitative detection of multiple antibodies. Line blotting enables the detection of antibodies on membrane strips coated with thin parallel lines of several purified, biochemically characterized antigens, which are fixed onto a synthetic support. Each strip can contain more than ten antigens, thus allowing simultaneous screening for multiple antibodies. Here, we describe the principle of line blotting and how the buffer composition can interfere with the results of autoantibody detection.

Key words

Line blotting Immunoblotting Autoantibodies Sensitivity Scanning densitometry 

References

  1. 1.
    Baslund B, Wieslander J (1994) A rapid and semi-quantitative method for the detection of autoantibodies by multiple spot immunoassay. J Immunol Methods 169(2):183–193CrossRefGoogle Scholar
  2. 2.
    Noya O, Losada S, Toledo M, Gauna A, Lorenzo MA, Bermúdez H, Alarcón de Noya B (2015) Improvements and variants of the multiple antigen blot assay-MABA: an immunoenzymatic technique for simultaneous antigen and antibody screening. Methods Mol Biol. 1312:301–319CrossRefGoogle Scholar
  3. 3.
    Stott DI (2000) Immunoblotting, dot-blotting, and ELISPOT assays: methods and applications. J Immunoassay 21(2–3):273–296CrossRefGoogle Scholar
  4. 4.
    Mishra M, Tiwari S, Gomes AV (2017) Protein purification and analysis: next generation Western blotting techniques. Expert Rev Proteomics 14(11):1037–1053CrossRefGoogle Scholar
  5. 5.
    Kurien BT, Scofield RH (2015) Western blotting: an introduction. Methods Mol Biol. 1312:17–30CrossRefGoogle Scholar
  6. 6.
    Stott DI (1989) Immunoblotting and dot blotting. J Immunol Methods 119(2):153–187CrossRefGoogle Scholar
  7. 7.
    Noya O, Losada S, Toledo M, Alarcón de Noya B (2009) The multiple antigen blot assay: a simple, versatile and multipurpose immunoenzymatic technique. Methods Mol Biol. 536:237–251CrossRefGoogle Scholar
  8. 8.
    Dorri Y (2015) Immunoblotting of antigens: whole, strip, and new-line nitrocellulose membrane immunoblotting using the chemiluminescence technique. Methods Mol Biol. 1314:79–94CrossRefGoogle Scholar
  9. 9.
    Kurien BT (2009) Strip immunoblotting of multiple antigenic peptides to nitrocellulose membrane. Methods Mol Biol 536:191–199CrossRefGoogle Scholar
  10. 10.
    Gallo-Oller G, Ordoñez R, Dotor J (2018) A new background subtraction method for Western blot densitometry band quantification through image analysis software. J Immunol Methods 467:1–5CrossRefGoogle Scholar
  11. 11.
    Gassmann M, Grenacher B, Rohde B, Vogel J (2009) Quantify Western blots: pitfalls of densitometry. Electrophoresis 30:1845–1855CrossRefGoogle Scholar
  12. 12.
    Murphy K, Janeway CA, Travers P, Walport M, Mowart A, Weaver CT (2012) Janeway’s immuno biology, 8th edn. Garland Science, New YorkGoogle Scholar
  13. 13.
    Borg EJ, Risselada AP, Kelder JC (2011) Relation of systemic autoantibodies to the number of extraglandular manifestations in primary Sjögren’s syndrome: a retrospective analysis of 65 patients in the Netherlands. Semin Arthritis Rheum 40(6):547–551CrossRefGoogle Scholar
  14. 14.
    Ciechomska M, Laar JV, O’Reilly S (2015) Current frontiers in systemic sclerosis pathogenesis. Exp Dermatol 24(6):401–406CrossRefGoogle Scholar
  15. 15.
    Saito H, Takahashi A, Abe K, Okai K, Katsushima F, Monoe K, Kanno Y, Ohira H (2012) Autoantibodies by line immunoassay in patients with primary biliary cirrhosis. Fukushima J Med Sci 58(2):107–116CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Louise Sternbæk
    • 3
  • Tina Friis
    • 1
  • Gunnar Houen
    • 1
    • 2
  1. 1.Department of AutoimmunologyStatens Serum InstitutCopenhagenDenmark
  2. 2.Department of Biochemistry and Molecular BiologyUniversity of Southern DenmarkOdenseDenmark
  3. 3.Phase Holographic ImagingLundSweden

Personalised recommendations