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Article Dans Une Revue Biochimica et Biophysica Acta Proteins and Proteomics Année : 2015

Atomic force microscopy analysis of IgG films at hydrophobic surfaces: A promising method to probe IgG orientations and optimize ELISA tests performance

Résumé

IgG films are widely used in the field of immunoassays, especially in (double) antibody-sandwich ELISA tests where capture antibodies are coated on surfaces like polystyrene or hydrophobic self-assembled monolayers (h-SAMs). It is critical to analyze-at a molecular scale and under liquid conditions the structure of the deposited IgG film in order to quantitatively address the efficiency of the ELISA test in terms of antigen detection. In this communication, we report an atomic force microscopy (AFM) analysis evidencing a strong relationship between immunological activities of mouse monoclonal anti-human interleukin-2 (IL-2) and 6 (IL-6) antibodies, thickness and roughness of the IgG monolayer adsorbed onto h-SAMs, and surface concentration of IgG molecules. Indirect information may be further obtained on antibody orientation. Collating the results obtained by AFM and those from ELISA tests leads us to conclude that antibodies like anti-IL-6 forming flat monolayers should be more efficient under ELISA detection conditions. In addition, the concentration of IgG in the coating suspension should be optimized to obtain a monolayer heavily populated by "end-on" adsorbed molecules, an orientation that is desirable for enhancing ELISA tests performance.
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Dates et versions

hal-01159533 , version 1 (03-06-2015)

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Pierre de Thier, Jalal Bacharouche, Jérôme F.L. Duval, Salaheddine Skali-Lami, G. Francius. Atomic force microscopy analysis of IgG films at hydrophobic surfaces: A promising method to probe IgG orientations and optimize ELISA tests performance. Biochimica et Biophysica Acta Proteins and Proteomics, 2015, 1854 (2), pp.138-145. ⟨10.1016/j.bbapap.2014.12.001⟩. ⟨hal-01159533⟩
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