Double One-Dimensional Electrophoresis (D1-DE) Adapted for Immunoproteomics

  • Youcef Shahali
  • Hélène Sénéchal
  • Pascal Poncet
Part of the Methods in Molecular Biology book series (MIMB, volume 1871)


The classical proteomics approach for the identification of allergen candidates consists on the separation of proteins by high-resolution two-dimensional electrophoresis (2-DE) with subsequent IgE immunoblotting and further analysis of IgE-reactive protein spots with mass spectrometry. In this approach at least two gels most be run. One gel is used for staining and the other is for immunoblotting by antibodies labeled with specific immunostains. Additional functional characterizations require either protein purification or 2-DE replicates and appear to be time- and reagent-consuming. Here we described a modified double one-dimensional electrophoresis (D1-DE) allowing the conversion of a protein spot previously visualized by 2-DE into an extended protein band. In D1-DE, the purity of the protein of interest is similar to 2-DE spots, but its abundance is many times higher than what can be found in a 2-DE single spot allowing many other functional analyses from a single D1-DE separation.

Key words

Double one-dimensional electrophoresis D1-DE 2-DE Allergens Proteomics 


  1. 1.
    Nony E, Le Mignon M, Brier S, Martelet A, Moingeon P (2016) Proteomics for allergy: from proteins to the patients. Curr Allergy Asthma Rep 16:64CrossRefGoogle Scholar
  2. 2.
    Mousavi F, Majd A, Shahali Y, Ghahremaninejad F, Shoormasti RS, Pourpak Z (2017) Immunoproteomics of tree of heaven (Ailanthus atltissima) pollen allergens. J Proteome 154:94–101CrossRefGoogle Scholar
  3. 3.
    Hoffmann-Sommergruber K (2016) Proteomics and its impact on food allergy diagnosis. EuPA Open Proteom 12:10–12CrossRefGoogle Scholar
  4. 4.
    Tiotiu A, Brazdova A, Longé C, Gallet P, Morisset M, Leduc V et al (2016) Urtica dioica pollen allergy: clinical, biological, and allergomics analysis. Ann Allergy Asthma Immunol 117:527–534CrossRefGoogle Scholar
  5. 5.
    Poncet P, Sénéchal H, Clement G, Purohit A, Sutra JP, Desvaux FX et al (2010) Evaluation of ash pollen sensitization pattern using proteomic approach with individual sera from allergic patients. Allergy 65:571–580CrossRefGoogle Scholar
  6. 6.
    D’Amato A, Bachi A, Fasoli E, Boschetti E, Peltre G, Sénéchal H et al (2010) In-depth exploration of Hevea brasiliensis latex proteome and “hidden allergens” via combinatorial peptide ligand libraries. J Proteome 73:1368–1380CrossRefGoogle Scholar
  7. 7.
    Shahali Y, Sutra JP, Fasoli E, D’Amato A, Righetti PG, Futamura N et al (2012) Allergomic study of cypress pollen via combinatorial peptide ligand libraries. J Proteome 77:101–110CrossRefGoogle Scholar
  8. 8.
    Shahali Y, Sutra JP, Peltre G, Charpin D, Sénéchal H, Poncet P (2010) IgE reactivity to common cypress (C. Sempervirens) pollen extracts: evidence for novel allergens. W Allergy Organ J 3:229–234CrossRefGoogle Scholar
  9. 9.
    Shahali Y, Nicaise P, Brazdova A, Charpin D, Scala E, Mari A et al (2014) Complementarity between microarray and immunoblot for the comparative evaluation of IgE repertoire of French and Italian cypress pollen allergic patients. Folia Biol 60:192Google Scholar
  10. 10.
    Shahali Y, Sutra JP, Haddad I, Vinh J, Guilloux L, Peltre G et al (2012) Proteomics of cypress pollen allergens using double and triple one-dimensional electrophoresis. Electrophoresis 33:462–469CrossRefGoogle Scholar
  11. 11.
    Altland K, Silke R, Hackler R (1981) Demonstation of human prealbumin by double one-dimensional slab gel electrophoresis. Electrophoresis 2:148–155CrossRefGoogle Scholar
  12. 12.
    Blum H, Beier H, Gross HJ (1987) Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis 8:93–99CrossRefGoogle Scholar
  13. 13.
    Demeulemester C, Peltre G, Laurent M, Panheleux D, David B (1987) Cyanogen bromide-activated nitrocellulose membranes: a new tool for immunoprint techniques. Electrophoresis 8:71–73CrossRefGoogle Scholar
  14. 14.
    Shahali Y, Sutra JP, Hilger C, Swiontek K, Haddad I, Vinh J et al (2017) Identification of a polygalacturonase (Cup s 2) as the major CCD-bearing allergen in Cupressus sempervirens pollen. Allergy 72:1806–1810CrossRefGoogle Scholar
  15. 15.
    Sénéchal H, Šantrůček J, Melčová M, Svoboda P, Zídková J, Charpin D et al (2018) A new allergen family involved in pollen food-associated syndrome: Snakin/gibberellin-regulated proteins. J Allergy Clin Immunol 141:411–414CrossRefGoogle Scholar

Copyright information

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

Authors and Affiliations

  • Youcef Shahali
    • 1
  • Hélène Sénéchal
    • 2
  • Pascal Poncet
    • 2
    • 3
  1. 1.Razi Vaccine and Serum Research InstituteAgricultural Research, Education and Extension Organization (AREEO)KarajIran
  2. 2.Allergy and Environment Team, Biochemistry DepartmentArmand Trousseau Children Hospital (AP-HP)ParisFrance
  3. 3.Institute PasteurCenter for Innovation and Technological ResearchParisFrance

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