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Issue 12, 2011
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Kinetics of immunoassays with particles as labels: effect of antibody coupling using dendrimers as linkers

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Abstract

In this article, we report on poly(amidoamine) dendrimers (PAMAM) as coupling agents for recombinant single-chain (ScFv) antibodies to nanoparticle (NP) labels, for use in immunoassay. We present a simple theory for the kinetics of particle capture onto a surface by means of an antibodyantigen reaction, in which the important parameter is the fraction of the particle surface that is active for reaction. We describe how increasing the generation number of the linking dendrimers significantly increased the fraction of the NP surface that is active for antigen binding and consequently also increased the assay kinetic rates. Use of dendrimers for conjugation of the NP to the antibody resulted in a significantly higher surface coverage of active antibody, in comparison with mono-valent linker chemistry. As a direct consequence, the increase in effective avidity significantly out-weighed any effect of a decreased diffusion coefficient due to the NP, when compared to that of a molecular dye-labelled antibody. The signal to noise ratio of the G4.5 dendrimer-sensitised nanoparticles out-performed the dye-labelled antibody by approximately four-fold. Particle aggregation experiments with the multi-valent antigen CRP demonstrated reaction-limited aggregation whose rate increased significantly with increasing generation number of the dendrimer linker.

Graphical abstract: Kinetics of immunoassays with particles as labels: effect of antibody coupling using dendrimers as linkers

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Supplementary files

Article information


Submitted
07 Jan 2011
Accepted
01 Apr 2011
First published
04 May 2011

Analyst, 2011,136, 2533-2541
Article type
Paper

Kinetics of immunoassays with particles as labels: effect of antibody coupling using dendrimers as linkers

V. Gubala, C. Crean (née Lynam), R. Nooney, S. Hearty, B. McDonnell, K. Heydon, R. O'Kennedy, B. D. MacCraith and D. E. Williams, Analyst, 2011, 136, 2533
DOI: 10.1039/C1AN15017K

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