End-on Covalent Antibody Immobilization on Dual Polarization Interferometry Sensor Chip for Enhanced Immuno-sensing

Hong Yan Song, Jonathan Hobley, Xiaodi Su, Xiaodong Zhou

研究成果: Article同行評審

11 引文 斯高帕斯(Scopus)


End-on immobilization of antibody is an important technology to greatly improve the sensitivity of immuno-sensing. The end-on, or constant fragment (Fc) site-specifically oriented antibodies expose more active antigen binding sites (Fab) and tend to capture more analytes from the solution. In this study, dual polarization interferometry (DPI), which is the only available equipment that can concurrently obtain the mass, thickness, density, and refractive index in real time, was applied to investigate two end-on antibody immobilization methods, as exemplified by prostate-specific antigen (PSA) antibody. In the first method, antibody was immobilized via the saccharide chain linked to Fc portion of the antibody, by chelation to the surface-bound boronic acid. In the second method, antibody was partially reduced by tris(2-carboxyethyl) phosphine (TCEP) under mild conditions, followed by covalent conjugation to the surface via thiol-maleimide reaction. Time-resolved measurement from DPI verifies the end-on conformation of the antibody on the sensor surface. The second method shows better detection performance, with enhanced sensitivity and reproducibility than the first method, due to the optimal alignment of the antibody. Finally, these two methods were compared with the protein G-based antibody orientation control and random antibody immobilization in terms of the mass, thickness, density, refractive index, reproducibility, and stability. The detailed antibody immobilization parameters obtained in this paper are of great importance in the developing of solid-liquid phase immunosensors with enhanced detection sensitivity.

頁(從 - 到)851-858
出版狀態Published - 2014 八月

All Science Journal Classification (ASJC) codes

  • 生物技術
  • 生物物理學
  • 生物化學


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