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

Youcef Shahali (1) , Hélène Sénéchal (2) , Pascal Poncet (2, 3)

1 AREEO - Agricultural Research, Education and Extension Organisation
2 CHU Trousseau [APHP]
3 Centre d'Innovation et Recherche Technologique / Center for Innovation and Technological Research

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.

Keywords: Double one-dimensional electrophoresis; D1-DE; 2-DE; Allergens; Proteomics

Domains: Life Sciences [q-bio]; Biochemistry [q-bio.BM]; Health; Allergology

Youcef Shahali, Hélène Sénéchal, Pascal Poncet. Double One-Dimensional Electrophoresis (D1-DE) Adapted for Immunoproteomics. Functional Proteomics, 1871, Springer New York; Humana Press, pp.133-141, 2018, Methods in Molecular Biology, 978-1-4939-8814-3. ⟨10.1007/978-1-4939-8814-3_9⟩. ⟨pasteur-03718453⟩