Quantitative measurement of p53-p53 antibody interactions by quartz crystal microbalance: A modelsystem for immunochemical calibration


We are developing methods to quantify antibody interactions that include a quartz crystal microbalance (QCM) system to measure, on a molecular basis, the interaction of p53 and anti-p53 antibodies. Previously, as a model system, we developed a measurement device consisting of p53 protein (human wild type), characterized by mass spectroscopy and immobilized at various concentrations on a glass slide. The device is designed as a control to be used with immunohistochemical (IHC) assays that incorporate commercially available anti-p53 antibodies and probes. In the current study, p53 protein is covalently immobilized on a silicon dioxide-coated quartz crystal and the resonance frequency shift is followed in-situ. The functionalized sensor is then incubated with the anti-p53 antibody, which provides a direct gravimetric measure of the antibody-antigen binding. This previously described method allows the comparison of the surface immobilized protein concentrations with estimates obtained by fluorescence measurement. The p53 functionalized QCM system offers an independent measure of surface immobilized protein concentration and is essential in development of our IHC calibration prototypes. These results provide a benchmark for comparing antibody systems that may be used in developing other molecular diagnostic assays such as immunocytochemical analysis and the detection of biomarker proteins in blood and urine.

Share and Cite:

H. Atha, D. and Reipa, V. (2012) Quantitative measurement of p53-p53 antibody interactions by quartz crystal microbalance: A modelsystem for immunochemical calibration. Journal of Biophysical Chemistry, 3, 211-220. doi: 10.4236/jbpc.2012.33024.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Jakupciak, J.P., Gallant, N.D., Smith, A.H., Becker, M.L., Tona, A. and Atha, D.H. (2009) Improved methods and standards for telomerase detection. Biotechnic & Histochemistry, 84, 1-12. HUdoi:10.1080/10520290903039060U
[2] Reipa, V., Purdum, G. and Choi, J. (2010) Measurement of nanoparticle concentration using quartz crystal microgravimetry. The Journal of Physical Chemistry B, 114, 16112-16117. HUdoi:10.1021/jp103861mU
[3] Marx, K.A. (2003) Quartz crystal microbalance: A useful tool for studying thin polymer films and complex biomolecular systems at the solution-surface interface. Biomacromolecules, 4, 1099-1120. HUdoi:10.1021/bm020116jU
[4] Liu, Y.-C., Wang, C.-M. and Hsiung, K.-P. (2001) Comparison of different protein immobilization methods on quartz crystal microbalance surface in flow injection immunoassay. Analytical Biochemistry, 299, 130-135. HUdoi:10.1006/abio.2001.5409UH
[5] Marques, J.T., Rebouillat, D., Ramana, C.V., Murakani, J., Hill, J.E. and Gudkov, A. (2005) Down-regulation of p53 by double-stranded RNA modulates the antiviral response. Journal of Virology, 79, 11105-11114. HUdoi:10.1128/JVI.79.17.11114.2005U
[6] Atha, D.H., Manne, U., Grizzle, W.E., Wagner, P.D., Srivastava, S. and Reipa, V. (2010) Standards for immunohistochemical imaging: A protein reference device for biomarker quantitation. Journal of Histochemistry & Cytochemistry, 58, 1005-1014. HUdoi:10.1369/jhc.2010.956342U
[7] Hook, F., Kasemo, B., Nylander, T., Fant, C., Sott, K. and Elwing, H. (2001) Variations in coupled water, viscoelastic properties, and film thickness of a Mefp-1 protein film during adsorption and cross-linking: A quartz crystal microbalance with dissipation monitoring, ellipsometry, and surface plasmon resonance study. Analytical Chemistry, 73, 5796-5804. HUdoi:10.1021/ac0106501U
[8] Sauerbrey, G. (1959) Verwendung von schwing quarzen zur wagung dunner schichten und zur mikrowagung. Zeitschrift für Physik a Hadrons and Nuclei, 155, 206- 222. HUdoi:10.1007/BF01337937U
[9] Hook, F., Rodahl, M., Brzezinski, P. and Kasemo, B. (1998) Energy dissipation kinetics for protein and antibody-antigen adsorption under shear oscillation on a quartz crystal microbalance. Langmuir, 14, 729-734. HUdoi:10.1021/la970815uU
[10] Hengerer, A., Kosslinger, C., Decker, J., Hauck, S., Queitsch, I. and Wolf, H., (1999) Determination of phage antibody affinities to antigen by a microbalance sensor system. BioTechniques, 26, 956-964.
[11] Hovgaard, M.B., Dong, M., Otzen, D.E. and Besenbacher, F. (2007) Quartz crystal microbalance studies of multilayer glucagon fibrillation at the solid-liquid interface. Biophysical Journal, 93, 2162-2169. HUdoi:10.1529/biophysj.107.109686U
[12] Feldoto, Z., Lundin, M., Braesch-Andersen, S. and Blomberg, E. (2011) Adsorption of IgG on/in a PAH/PSS multilayer film: Layer structure and cell response. Journal of Colloid and Interface Science, 354, 31-37. HUdoi:10.1016/j.jcis.2010.10.006U
[13] Hook, F., Voros, J., Rodahl, M., Kurrat, R., Boni, P. and Ramsden, J.J. (2002) A comparative study of protein adsorption on titanium oxide surfaces using in sity ellipsometry, optical waveguide lightmode spectroscopy, and quartz crystal microbalance/dissipation. Colloids and Surfaces B: Biointerfaces, 24, 155-170. HUdoi:10.1016/S097-7765(01)00236-3U
[14] Brewer, S.H., Glomm, R., Johnson, M.C., Knag, M.K. and Franzen, S. (2005) Probing BSA binding to citrate-coated gold nanoparticles and surfaces. Langmuir, 21, 9303-9307. HUdoi:10.1021/la50588tU
[15] Shen, D., Huang, M., Chow, L.M. and Yang, M. (2001) Kinetic profile of the adsorption and conformational change of lysozyme on self-assembled monolayers as revealed by quartz crystal resonator. Sensors and Actuators, B77, 664-670.
[16] Yang, M., Chung, F.L. and Thompson, M. (1993) Acoustic network analysis as a novel technique for studying protein adsorption and denaturation at surfaces. Analytical Chemistry, 65, 3713-3716. HUdoi:10.21/ac00072a027U
[17] Giamblanco, N., Yaseen, M., Zhavnerko, G., Lu, J.R. and Marletta, G. (2011) Fibronectin conformation switch induced by coadsorption with human serum albumin. Langmuir, 27, 312-319. HUdoi:10.1021/la104127qU
[18] Richart, L., Variola, F., Rosei, F., Wuest, J.D. and Nanci, A. (2010) Adsorption of proteins on nanoporous Ti surfaces. Surface Science, 604, 1445-1451. HUdoi:10.1016/jsusc.2010.05.007U
[19] Hammersam, A.G., Foss, M., Chevallier, J. and Besenbacher, F. (2005) Adsorption of fibrinogen on tantalum oxide, titanium oxide and gold studied by the QCM-D technique. Colloids and Surfaces B: Biointerfaces, 43, 208-215. HUdoi:10.1016/j.colsurfb.2005.04.007U
[20] Yao, C., Zhu, T., Qi, Y., Zhao, Y., Xia, H. and Fu, W. (2010) Development of a quartz crystal microbalance biosensor with aptamers as bio-recognition element. Sensors, 10, 5859-5871. HUdoi:10.3390/s100605859U
[21] Tagaya, M., Ikoma, T., Takemura, T., Hanagata, N., Okuda, M., Yoshioka, T. and Tanaka, J. (2011) Detection of interfacial phenomena with osteoblast-like cell adhesion on a hydroxyapatite and oxidized polystyrene by the quartz crystal microbalance with dissipation. Langmuir, 27, 7635-7644. HUdoi:10.1021/1a200008zU

Copyright © 2021 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.