Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2015 Aug;11(31):3782-8.
doi: 10.1002/smll.201500112. Epub 2015 Apr 11.

Measuring Binding Kinetics of Antibody-Conjugated Gold Nanoparticles With Intact Cells

Affiliations
Free PMC article

Measuring Binding Kinetics of Antibody-Conjugated Gold Nanoparticles With Intact Cells

Linliang Yin et al. Small. .
Free PMC article

Abstract

Antibody-conjugated nanomaterials have attracted much attention because of their applications in nanomedicine and nanotheranostics, and amplification of detection signals. For many of these applications, the nanoconjugates must bind with a cell membrane receptor (antigen) specifically before entering the cells and reaching the final target, which is thus important but not well understood. Here, a plasmonic imaging study of the binding kinetics of antibody-conjugated gold nanoparticles with antigen-expressing cells is presented, and the results are compared with that of the nanoparticle-free antibody. It is found that the nanoconjugates can significantly affect the binding kinetics compared with free antibody molecules, depending on the density of the antibody conjugated on the nanoparticles, and expressing level of the antigen on the cell membrane. The results are analyzed in terms of a transition from monovalent binding model to a bivalent binding model when the conjugation density and expressing level increase. These findings help optimize the design of functional nanomaterials for drug delivery and correct interpretation of data obtained with nanoparticle signal amplification.

Keywords: binding kinetics; membrane proteins; nanomedicine; plasmonics.

Figures

Figure 1
Figure 1. Monitoring the binding kinetics of nano-conjugates with single intact cells
(a) Schematic illustration of the plasmonic imaging setup. A p-polarized beam is directed onto the gold-coated glass coverslip through the prism underneath to excite the surface plasmon. The reflected light is captured by a CCD camera to produce a plasmonic image of individual adherent cells. (b) The structure of a Herceptin@AuNP nano-conjugate.
Figure 2
Figure 2. In situ binding kinetics of nano-conjugates with SKBR3 cells
(a) A typical plasmonic image of SKBR3 cells, where the bright spots represent individual cells. (b) Typical sensorgrams showing the association and dissociation of nano-conjugates with surface bound Herceptin (black dots) and free Herceptin (blue dots) with SKBR3 cells. The black and red curves are fitted curves with the bi-valent (for pink dots) and mono-valent binding models (for blue dots), respectively. (c) Mapping of nano-conjugate distribution on SKBR3 cells, showing negligible non-specific interactions of nano-conjugates in the gold regions surrounding the cells. (d) Individual sensorgrams of the nano-conjugates with multiple SKBR3 cells (grey), average sensorgram over the individual cells (pink), and the fit of the average sensorgram the bi-valent binding model (black). The cyan curve represents the initial binding of Herceptin to Her2 on the cell membrane via one pair of Herception-Her2 interaction, and the magenta curve is the binding via a second pair of Herception-Her2 interaction. (e) Schematic illustration of the bi-valent binding model.
Figure 3
Figure 3. In situ binding kinetics of nano-conjugates with JIMT1 cells
(a) A typical plasmonic image of several JIMT1 cells adherent on a gold chip. (b) Mass distribution of nano-conjugates on JIMT1 cells, showing negligible non-specific interactions of nano-conjugates in the surrounding gold regions. (c) Sensorgrams showing the association and dissociation of nano-conjugates (pink dots) and free Herceptin (blue dots) with JIMT1 cells, where the black curve is a fit with the bi-valent binding model. (d) Plasmonic signal amplification due to the presence of gold nanoparticles. (e) Individual sensorgrams of the nano-conjugates with multiple JIMT1 cells (grey), average sensorgram over the different cells (pink), where the fit of the average sensorgram the bi-valent binding model (black). The cyan curve represents the initial binding of Herceptin to Her2 on the cell membrane via one pair of Herception-Her2 interaction, and the magenta curve is the binding via a second pair of Herception-Her2 interaction. (f) Schematic illustration of the loss of bi-valent binding due to increased inter-molecular distance of Her2 receptors.
Figure 4
Figure 4. Influence of Herceptin conjugation density on the binding mechanisms of nano-conjugates
(a) Sensorgrams with Herceptin to AuNP molar ratios at 150 (pink curve), 64 (blue curve) and 25 (magenta curve), respectively. (b) The sensorgram of nano-conjugates with the lowest Herceptin to AuNP molar ratio fits typical monovalent binding model (panel b inset). (c) The sensorgram of nano-conjugates with the highest Herceptin to AuNP molar ratio fits the bi-valent binding model (panel c inset). Cyan and magenta lines represent the initial mono-valent attachment and the formation of bi-valent binding, respectively.

Similar articles

See all similar articles

Cited by 3 articles

Publication types

Feedback