Optical Response of Magnetic Fluorescent Microspheres Used for Force Spectroscopy in the Evanescent Field

Abstract

Force spectroscopy based on magnetic tweezers is a powerful technique to manipulate single biomolecules and study their interactions. The resolution in a magnetic probe displacement, however, needs to be commensurate with molecular sizes. To achieve the desirable sensitivity in tracking displacements of the magnetic probe, some recent approaches have combined magnetic tweezers with total internal reflection fluorescence microscopy. In this situation, a typical force probe is a polymer microsphere containing two types of optically active components -a pure absorber (magnetic nanoparticles for providing the pulling force) and a luminophore (semiconducting nanoparticles or organic dyes for fluorescent imaging). In order to fully assess the system's capability for tracking the position of the force probe with sub-nanometer accuracy, we developed a body-of-revolution formulation of the method of auxiliary sources (BOR-MAS) to simulate absorption, scattering, and fluorescence of microscopic spheres in an evanescent electromagnetic field. The theoretical formulation uses the axial symmetry of the system to reduce the dimensionality of the modeling problem and produces excellent agreement with the reported experimental data on forward scattering intensity. Using the BOR-MAS numerical model, we investigated the probe detection sensitivity for a high numerical aperture objective. The analysis of both backscattering and fluorescence observation modes shows that the total intensity of the bead image decays exponentially with the distance from the surface (or the length of a biomolecule). Our investigations demonstrate that the decay lengths of observable optical power are smaller than the penetration depth of the unperturbed excitation evanescent wave. In addition, our numerical modeling results illustrate that the expected sensitivity for the decay length changes with the incident angle, tracking the theoretical penetration depth for a two-media model, and is sensitive to the bead size. The BOR-MAS methodology developed in this work for near field modeling of bead tracking experiments fully describes the fundamental photonic response of microscopic BOR probes at the sub-wavelength level and can be used for future improvements in the design of these probes or in the setup of bead tracking experiments.

Dielectrophoretic tweezers as a platform for molecular force spectroscopy in a highly parallel format

“…47, 51, 52 Both methods are compatible with flat electrodes, however, for structured electrodes, TIRF microscopy is preferred because only total intensity of the probe fluorescence or scattering intensity is needed to map the vertical position of the probe. 53 We used through-objective laser TIRF in our experiments with DEP tweezers. To achieve TIR conditions at the bottom of the microwells, the incident laser beam must be prevented from entering the SU-8 resist layer, since otherwise propagating light conditions will be achieved effectively throughout the whole sample.…”

Mitigation of eddy current heating during magnetic nanoparticle hyperthermia therapy

“…MAS is a robust and accurate numerical technique for evaluating electromagnetic wave propagation and scattering problems [61, 62]. In this method, boundaries between materials of differing electrical parameters are defined and discretized into pairs of points along a surface.…”

Magnetic nanoparticle hyperthermia: predictive model for temperature distribution

“…The electromagnetic model is based on the Method of Auxiliary Sources (MAS), a robust, accurate numerical technique for solving electromagnetic wave propagation and scattering problems 15, 16 . The AMF source was modeled based on coil schematics and magnetic core material properties (Fluxtrol Inc., Aubrun Hills, MI, USA).…”