Insights into the Formation of DNA–Magnetic Nanoparticle Hybrid Structures: Correlations between Morphological Characterization and Output from Magnetic Biosensor Measurements

Understanding the binding mechanism between probe-functionalized magnetic nanoparticles (MNPs) and DNA targets or amplification products thereof is essential in the optimization of magnetic biosensors for the detection of DNA. Herein, the molecular interaction forming hybrid structures upon hybridization between DNA-functionalized magnetic nanoparticles, exhibiting Brownian relaxation, and rolling circle amplification products (DNA-coils) is investigated by the use of atomic force microscopy in a liquid environment and magnetic biosensors measuring the frequency-dependent magnetic response and the frequency-dependent modulation of light transmission. This approach reveals the qualitative and quantitative correlations between the morphological features of the hybrid structures with their magnetic response. The suppression of the high-frequency peak in the magnetic response and the appearance of a new peak at lower frequencies match the formation of larger sized assemblies upon increasing the concentration of DNA-coils. Furthermore, an increase of the DNA-coil concentration induces an increase in the number of MNPs per hybrid structure. This study provides new insights into the DNA–MNP binding mechanism, and its versatility is of considerable importance for the mechanistic characterization of other DNA-nanoparticle biosensor systems.

nm (see inset in Figure S4) can be associated to free MNPs and it correlates with peak 1 of the    Optomagnetic system description and optomagnetic measurement principle.
As illustrated in Figure   The optomagnetic measurement principle is based on the rotational dynamics of magnetic nanoparticles (MNPs). The MNPs employed in this study have a remnant magnetic moment, which implies that the dominating relaxation mechanism upon a reversal of the magnetic field direction is a physical rotation of the particle, known as Brownian relaxation. The characteristic frequency for Brownian relaxation dynamics is given by where is the dynamic viscosity of the carrier liquid, is the thermal energy, ℎ is the hydrodynamic volume, and ℎ is the hydrodynamic diameter of the relaxing entity (e.g. a single MNP). 2-4 The dynamic magnetic behavior can be described in term of the magnetic susceptibility with real (in-phase) and imaginary (out-of-phase) parts ′ and ′′ , respectively. In case of a sinusoidal magnetic field ℎ 0 ( ), the time dependent linear magnetic response can be expressed as where ′ = 0 ( ) and ′′ = 0 ( ). At low frequencies the MNPs are able to rotate and follow the magnetic field, and the response is in-phase with the applied field. Therefore ′ is maximal. The rotation of the MNPs starts to lag behind the applied field at higher frequencies, which leads to a decrease in the in-phase component ′ and a corresponding increase in the outof-phase component ′′. The out-of-phase component ′′ attains its maximum value at the Brownian relaxation frequency .
A simple approach to account for a distribution of MNP sizes was introduced by Cole and Cole 5 according to the following expression for the complex magnetic susceptibility where α is the Cole-Cole parameter (ranging from 0 to 1, a measure of the nanoparticle size distribution width), = (2 ) −1 is the Brownian relaxation time, = 2 is the angular frequency of the applied field and 0 and ∞ are the zero and high frequency limits of .
The dynamics is determined by the rotational behavior of the individual MNPs, which follows the Brownian relaxation dynamics. The modulation of the transmitted light is found in the complex second harmonic voltage output from the photodetector where 2 ′ and 2 ′′ are the in-phase and out-of-phase signals, respectively. The modulation is measured using a lock-in amplifier with the AC magnetic field excitation as reference. From the perspective of transmitted light, the MNP ensemble will scatter light equally for a positive and negative magnetic field of the same amplitude. We therefore assume that the photodetector signal can be described as where 0 represents the un-modulated part of the transmitted light (used here for normalization), where 2 (0) = 4 0 3√2 ⁄ is the zero-frequency limit of 2 ′′ .
The sign of depends on the optical scattering properties and the measurement geometry. For a geometry where the transmission is measured perpendicular to the axis of the applied magnetic field, as used in the present study, it is generally found that is negative for MNPs with sizes smaller than about 130 nm for blue laser light (λ = 405 nm). For even larger scattering entities, first becomes positive (e.g. for 250 nm MNPs) and then negative (e.g. for 500 nm MNPs). This originates from the oscillation of the scattering cross-section with particle size as can be accounted for by Mie scattering theory.