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Autoantibodies pp 113-131 | Cite as

Measurement of Autoantibodies to Gastric H+,K+-ATPase (ATP4A/B) Using a Luciferase Immunoprecipitation System (LIPS)

  • Edith Lahner
  • Ilaria Marzinotto
  • Cristina Brigatti
  • Howard Davidson
  • Janet Wenzlau
  • Lorenzo Piemonti
  • Bruno Annibale
  • Vito LampasonaEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1901)

Abstract

The Luciferase Immuno Precipitation System (LIPS) enables the detection of specific serum antibodies by immunoprecipitation of recombinant antigens tagged with a luciferase reporter. Here we describe LIPS assays for the quantification of autoantibodies to the H+, K+-ATPase A (ATP4A) and B (ATP4B) subunits, two serological markers of autoimmune atrophic gastritis and pernicious anemia. In particular, we will describe the expression of luciferase-tagged recombinant ATP4A and ATP4B, their immunoprecipitation with test sera, the recovery and washing of immune-complexes with a protein-A coated resin, and the quantification of autoantibodies by addition of a luciferase substrate and the measurement of the light output from captured luciferase-tagged antigens.

Key words

Atrophic gastritis Autoimmune gastritis Autoantibodies ATP4A ATP4B Gastric proton pump H+,K+-ATPase LIPS Luciferase immunoprecipitation system EIA Enzyme immunoassay 

Notes

Acknowledgments

E.L. and B.A. acknowledge support from University Sapienza 2013–2014. H.W.D. and J.M.W. acknowledge support from NIH grant R01 DK052068 (to H.W.D.). I.M., C.B., L.P., and V.L. acknowledge that their work was carried out within the framework of the “Ivascomar project, Cluster Tecnologico Nazionale Scienze della Vita ALISEI, Italian Ministry of Research” and with the support of the Associazione Italiana per la Ricerca sul Cancro (AIRC, bando 5 × 1000 N_12182). The authors wish to thank Carlo Lombardoni for his help in drawing Fig. 1.

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Copyright information

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

Authors and Affiliations

  • Edith Lahner
    • 1
  • Ilaria Marzinotto
    • 2
  • Cristina Brigatti
    • 3
  • Howard Davidson
    • 4
  • Janet Wenzlau
    • 4
  • Lorenzo Piemonti
    • 3
  • Bruno Annibale
    • 1
  • Vito Lampasona
    • 2
    Email author
  1. 1.Medical-Surgical Department of Clinical Sciences and Translational Medicine, Sant’Andrea HospitalSapienza University of RomeRomeItaly
  2. 2.Division of Genetics and Cell BiologyIRCCS San Raffaele Scientific InstituteMilanItaly
  3. 3.Beta Cell Biology Unit, Diabetes Research InstituteIRCCS San Raffaele Scientific InstituteMilanItaly
  4. 4.Barbara Davis Center for DiabetesUniversity of Colorado Anschutz Medical CampusAuroraUSA

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