Feasibility and constraints of particle targeting using the antigen-antibody interaction

Abstract

This work is concerned with the surface modification of fluorescent silica nanoparticles by a monoclonal antibody (M75) and the specific bioadhesion of such particles to surfaces containing the PG domain of carbonic anhydrase IX (CA IX), which is a trans-membrane protein specifically expressed on the surfaces of several tumor cell lines. The adhesion strength of antibody-bearing silica nanoparticles to antigen-bearing surfaces was investigated under laminar flow conditions in a microfluidic cell and compared to the adhesion of unmodified silica nanoparticles and nanoparticles coupled with an unspecific antibody. Adhesion to cancer cells using flow cytometry was also investigated and in all cases the adhesion strength of M75-modified nanoparticles was significantly stronger than for the unmodified or unspecific nanoparticles, up to several orders of magnitude in some cases. The specific modification of nano- and microparticles by an antibody-like protein therefore appears to be a feasible approach for the targeting of tumor cells.

Fig. 1. (a) Schematic illustration of SiO₂ nanoparticles modified by a specific monoclonal antibody and their interactions with the trans-membrane antigen of a tumor cell. (b) Schematic of the principle of adhesion force measurement in a laminar flow field.

Fig. 2. SiO₂ nanoparticles: (a) TEM image (the scale bar represents 200 nm) and (b) size distribution obtained by DLS and TEM.

Fig. 3. (a) Divided adhesion cell (upper part with inflow and outflow hoses, bottom part with milled space for plastic slides and rubber spacer) and (b) assembled adhesion cell connected to a syringe pump.

Fig. 4. Fluorescence response of the ELISA-like test for 4 types of particles: (a) linear scale and (b) logarithmic scale.

Fig. 5. Effect of aging on particle adhesion in an ELISA-like test repeated after 1 and 7 months.

Fig. 6. Fluorescence microscopy images of SiO₂-M75 particles deposited on a PG-MBP modified slide after increasing the exposure time (the scale bar represents 100 μm).

Fig. 7. Time dependence of surface coverage by SiO₂-M75 particles evaluated from the fluorescence microscopy images shown in Fig. 6.

Fig. 8. Fluorescence microscopy image of an interface between an area containing the PG-MBP antigen domain with deposited SiO₂-M75 particles and the surrounding blocked area (the scale bar represents 100 μm).

Fig. 9. Fluorescence microscopy images of adhesion at three different fluid velocities for four particle types: (a) SiO₂-M75; (b) SiO₂-IgG-X; (c) SiO₂-BSA; and (d) SiO₂ (the scale bar represents 100 μm).

Fig. 10. (a) Influence of shear rate inside the flow cell on the surface coverage for all four types of particles. (b) Effect of increasing shear rate on the removal of the M75 modified particles.

Fig. 11. Comparison of flow cytometry results (fluorescence intensity distribution) for the adhesion of all four particle types to HT-29 cells. (a) Fluorescence intensity distribution at 80× dilution. (b) Comparison of the mean fluorescence intensity at several dilutions.

Fig. 12. Summary of the flow cytometry results on HT-29 and NIH 3T3 cells (mean fluorescence intensity) for all four particle types as a function of dilution.