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Posted: Jun 16, 2011
Simple double-bead assay method for detecting cancer markers in biological samples
(Nanowerk News) The integration of nanoparticles into lab-on-a-chip devices has the potential to improve point-of-care applications. Of particular interest are the so-called double-bead immunoassays where magnetic nanoparticles are used in combination with gold nanoparticles. The magnetic nanoparticles are employed to isolate and concentrate the target analyte. The gold nanoparticles can then selectively bind the analyte isolated by the magnetic nanoparticles. Moreover, gold nanoparticles have interesting optical properties, making them suitable for signal transduction.
Schematic representation of the performed double-bead sandwich assay (A: incubation; B: separation).
Imec and K.U.Leuven have now exploited the use of gold and magnetic nanoparticles for detecting cancer markers in biological samples (serum). An innovative and simple approach was used to quantify the analyte concentration. The method is simple, generally applicable and cheap. Moreover, it can easily be implemented within a microfluidic device for point-of-care applications and as such help to elaborate personal diagnostics.
The detection of PSA was chosen to demonstrate the effectiveness of the proposed double-bead assay in both buffer and serum environment. Prior to the measurements, commercially available magnetic nanoparticles and gold nanoparticles were biofunctionalized with monoclonal antibodies (301 and 310 respectively). A key step for the covalent biofunctionalization of magnetic nanoparticles is the pre-concentration, i.e., the maximization of the electrostatic force between the antibody and the nanoparticle. The PSA binding event was quantified by measuring the unbound gold nanoparticle fraction using UV-vis spectroscopy, which provides a fast and cheap optical measurement. In addition, the sandwich complexes were visualized by scanning electron microscopy (SEM).
SEM image of the sandwich structures obtained by capturing PSA (0, 2, 20, 200, 2000 ng/mL) using MNP 310 and GNP-301 (20, 40, 50, 60, 80, 100 nm).
The double-bead assay was performed using gold nanoparticles of different sizes (20-100nm). Not all sizes were equally suited: an optimal detection signal was found for a gold nanoparticle size of 60nm. The occurrence of this optimum can be explained by two opposite trends: on the one hand, larger gold nanoparticles are functionalized less efficiently, on the other hand, larger gold nanoparticles give rise to larger optical signals. By using the optimal 60nm gold nanoparticles, clinically relevant concentrations for the detection of PSA were obtained. A detection limit of 5-8ng/mL, a sensitivity of 0.6-0.8(pg/mL)-1 and a dynamic range of 3 orders of magnitude were achieved without any further amplification. By increasing the concentration of the magnetic nanoparticles, an even further increase in assay sensitivity was obtained.
Since the biofunctionalization of the nanoparticles is very simple and generally applicable, the method described in this research can be used for the detection of other relevant cancer markers.