Quantification of specific bindings of biomolecules by magnetorelaxometry

被引：39

作者：

Eberbeck D. ^{[1
]}

Bergemann C. ^{[2
]}

Wiekhorst F. ^{[1
]}

Steinhoff U. ^{[1
]}

Trahms L. ^{[1
]}

机构：

[1] Physikalisch-Technische Bundesanstalt, D-10587 Berlin

[2] Chemicell GmbH, D-12103 Berlin

来源：

Journal of Nanobiotechnology | 2008年 / 6卷 / 1期

关键词：

Association reactions - Binding sites - Bioassay - Magnetic materials - Nanoparticles - Proteins - Rate constants - SQUIDs;

D O I：

10.1186/1477-3155-6-4

中图分类号：

学科分类号：

摘要：

The binding reaction of the biomolecules streptavidin and anti-biotin antibody, both labelled by magnetic nanoparticles (MNP), to biotin coated on agarose beads, was quantified by magnetorelaxometry (MRX). Highly sensitive SQUID-based MRX revealed the immobilization of the MNP caused by the biotin-streptavidin coupling. We found that about 85% of streptavidin-functionalised MNP bound specifically to biotin-agarose beads. On the other hand only 20% of antibiotin-antibody functionalised MNP were specifically bound. Variation of the suspension medium revealed in comparison to phosphate buffer with 0.1% bovine serum albumin a slight change of the binding behaviour in human serum, probably due to the presence of functioning (non heated) serum proteins. Furthermore, in human serum an additional non-specific binding occurs, being independent from the serum protein functionality. The presented homogeneous bead based assay is applicable in simple, uncoated vials and it enables the assessment of the binding kinetics in a volume without liquid flow. The estimated association rate constant for the MNP-labelled streptavidin is by about two orders of magnitude smaller than the value reported for free streptavidin. This is probably due to the relatively large size of the magnetic markers which reduces the diffusion of streptavidin. Furthermore, long time non-exponential kinetics were observed and interpreted as agglutination of the agarose beads. © 2008 Eberbeck et al; licensee BioMed Central Ltd.

引用

共 25 条

[1]

Mutschler E., Geisslinger G., Kroemer H.K., Schafer-Korting M., Arzneimittelwirkungen, (2005)

[2]

Karlsson R., Roos H., Reaction kinetics, Principles and Practice of Immunoassay, pp. 99-122, (1997)

[3]

Burke T.J., Loniello K.R., Beebe J.A., Ervin K., Development and application of fluorescence polarization assays in drug discovery, Comb Chem High Throughput Screen, 6, 3, pp. 183-194, (2003)

[4]

Boozer C., Kim G., Cong S., Guan H., Londergan T., Looking towards label-free biomolecular interaction analysis in a high-throughput format: A review of new surface plasmon resonance technologies, Curr Opin Biotechnol, 17, 4, pp. 400-405, (2006)

[5]

Hoa X.D., Kirk A.M., Tabrizian M., Towards integrated and sensitive surface plasmon resonance biosensors: A review of recent progress, Biosens Bioelectron, 23, 2, pp. 151-160, (2007)

[6]

Kotitz R., Fannin P.C., Trahms L., Time domain study of Brownian and Néel relaxation in ferrofluids, J Magn Magn Mat, 149, 1-2, pp. 42-46, (1995)

[7]

Weitschies W., Kotitz R., Bunte T., Trahms L., Determination of relaxing or remanent nanoparticle magnetization provides a novel binding-specific technique for the evaluation of immunoassays, Pharm Pharmacol Lett, 7, pp. 1-4, (1997)

[8]

Kotitz R., Trahms L., Koch H., Weitschies W., Ferrofluid relaxation for biomagnetic imaging, Biomagnetism: Fundamental Research and Clinical Applications, pp. 785-788, (1995)

[9]

Lange J., Kotitz R., Haller A., Trahms L., Semmler W., Weitschies W., Magnetrelaxometry-a new binding specific detection method based on magnetic nanoparticles, J Magn Magn Mat, 252, pp. 381-383, (2002)

[10]

Grossman H.L., Myers W.R., Vreeland V.J., Bruehl R., Alper M.D., Bertozzi C.R., Clarke J., Detection of bacteria in suspension by using a superconducting quantum interference device, Proc Natl Acad Sci, 101, pp. 129-134, (2004)

← 1 2 3 →