Abstract
In solid-phase analytical techniques, a reduction in nonspecific binding at the solid-phase surface is key to obtaining excellent assay performance. We have developed a new preparation method to minimize nonspecific protein binding in the solid phase. Microwave irradiation (2.45 GHz, 500 W) was used to produce a highly protein-resistant, dense coating layer of bovine serum albumin (BSA) that was acid-denatured at pH 4.0. The protein resistance in the BSA-coated magnetic bead was assessed by measuring the enzyme activity of alkaline phosphatase-labeled mouse immunoglobulin G (ALP-IgG) that nonspecifically bound to the surface. In addition, we observed the BSA-coated beads by atomic force microscopy (AFM) and also quantitatively measured the bound BSA, enabling us to propose a model for the denaturation processes caused by microwave irradiation. Compared to conventional BSA-coated beads prepared at pH 8.0 without microwave treatment, the activity of nonspecifically bound ALP-IgG was markedly reduced to 3%. Our results strongly suggest that a meshwork film of extensively associated BSA molecules is formed on the bead surface. This new technique should be useful for the development of polymer supports for use in solid-phase separation analyses, such as heterogeneous immunosorbent assay and column chromatography.
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E. Ishikawa, S. Hashida, T. Kohno, and K. Hirota, Clin. Chim. Acta, 194, 51 (1990).
J. E. Butler, Methods Enzymol., 73, 482 (1981).
S. Yoshitake, M. Imagawa, E. Ishikawa, Y. Niitsu, I. Urushizaki, M. Nishiura, R. Kanazawa, H. Kurosaki, S. Tachibana, N. Nakazawa, and H. Ogawa, J. Biochem., 92, 1413 (1982).
N. Miura, N. Asaka, N. Shinyashiki, and S. Mashimo, Biopolymers, 34, 357 (1994).
H. Bohr and J. Bohr, Phys. Rev. E, 61, 4310 (2000).
C. Y. Wu, S. T. Chen, S. H. Chiou, and K. T. Wang, J. Protein Chem., 11, 45 (1992).
A. Morales-Rubio, C. J. Salvador, and M. de la Guardia, Microchem. J., 47, 270 (1993).
A. Higo and M. Shimazaki, J. Home Econ. Jpn., 35 (11), 778 (1984).
L. D. Gapochka, M. G. Gapochka, A. F. Korolyov, A. V. Roshchin, A. P. Sukhorukov, N. N. Sysoev, and I. V. Timoshkin, Crit. Rev. Biomed. Eng., 28, No. 5–6, 40 (2000).
P. K. Smith, R. I. Krohn, G. T. Hermanson, A. K. Mallia, F. H. Gartner, M. D. Provenzano, E. K. Fujimoto, N. M. Goeke, B. J. Olson, and D. C. Klenk, Anal. Biochem., 150, 76 (1985).
I. Bronstein, B. Edwards, and J. C. Voyta, J. Biolumin. Chemilumin., 4, 99 (1989).
T. Peters, Jr., “All about Albumin,” Academic Press, San Diego, CA, 1996.
P. Bagchi and S. M. Birnbaum, J. Colloid Interface Sci., 83, 460 (1981).
H. Shirahama and T. Suzawa, J. Colloid Interface Sci., 104, 416 (1985).
N. B. Holland and R. E. Marchant, J. Biomed. Mater. Res., 51, 307 (2000).
T. Matsumoto and H. Inoue, J. Colloid Interface Sci., 160, 105 (1993).
V. J. C. Lin and J. L. Koenig, Biopolymers, 15, 203 (1976).
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Kawaguchi, S., Higo, Y. Effects of Microwave Irradiation on Nonspecific Protein Binding in the Solid Phase Coated with Bovine Serum Albumin. Polym J 37, 109–117 (2005). https://doi.org/10.1295/polymj.37.109
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DOI: https://doi.org/10.1295/polymj.37.109