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Optical Transmittance for Analysis of the Dynamics of Self-Assembled Rotating Chains of Superparamagnetic Micro- and Nano Beads in Solution

Optically monitoring biosensing procedures based on the dynamics of an aqueous solution containing functionalized superparamagnetic beads in the application of the external rotating magnetic field has been developed for a rapid, highly sensitive, and inexpensive bioassay. Typically, the dynamics of micrometer diameter beads is observed by conventional optical microscopes. For greater affinity to biomolecules, there is a demand which necessitates the use of nanometer sized superparamagnetic beads, comparable size to actual biomolecules. However, a limited amount of work for monitoring the dynamics of nanometer sized beads has been performed thus far due to the maximum resolution of microscopes. Here, we propose a novel protocol enabling us monitor the dynamics of nanometer-diameter beads via change in the optical transmittance.


Introduction
Biosensing protocols based on functionalized superparamagnetic beads (SPBs) have widely been studied for developing inexpensive, rapid, and highly-sensitive biodiagnosis of disorder [1] [2], even testing the use of illegal drag [3]. Several sorts of biosensing protocols utilizing the functionalized SPBs have been introduced over past decades. The optical, magnetic, chemical, and mechanical properties depend mostly on the particle shape, size, composition, and more importantly the applied external magnetic field. Among many, an optical mean to measure the intensity of fluorescence to determine the biorecognition of interest has particularly been utilized [4]. However, the optical method requires expensive equipments, such as a fluorometer and a fluorescence microtiter plate reader, thus it is limited only in affluent research centers and hospitals [5]. For that reason, it finds difficult the protocol to become prevailed. In order to familiarize a biosensing protocol, it must be inexpensive, highly sensitive, and more importantly easy to use [6].
Many researchers have attempted to realize the requisites with using conventional optical microscopes. However, this mean has not been able to response to a recent trend due to its limitation, extremely challenging to observe magnetic beads smaller than intrinsic maximum resolution of a microscope [7]. The recent trend puts great focus on the use of the functionalized SPBs less than 300-nm-diameter comparable size to actual biomolecules for greater affinity to target biomolecules [8].
Recently, there has been considerable interest in magneto-optic devices which combine magnetic and optical phenomena. The optical transmittance of an aqueous solution containing SPBs under an applied external magnetic field is one of the essential areas of magneto-optical effects. Some reports show that the optical transmittance of the magnetic fluids can be modulated by varying the external magnetic field [9][10][11][12]. In addition to magnetic field effect, the wavelength effect on the transmission of the magnetic fluids was also reported [13].
Piecing together published reports, the optical transmittance is greatly dependent on the external magnetic field and its direction, the wavelength of the light, and the orientation of the magnetically induced superparamagnetic bead chains.
Here we show a potential application of a magneto optical method to optical biosensing protocols. Our biosensing new protocol have already been published elsewhere [14]. We need to more research of the effect of bead size, composition, and configuration on the transmittance of the light passing through a column filled with an aqueous solution containing a kind of superparamagnetic bead has been investigated as a function of the applied external magnetic field and frequency. The beads we used were 2.8-μm-diameter SPBs (consisting nanometer sized magnetic beads, such as Fe 3 O 4 , typically 5-10 nm in diameter, are embedded into polystyrene matrix) and 250-nm-diameter SPBs (composing of crosslinked nanometer sized magnetic beads with dextran). Our procedure enables to monitor the dynamics of sub-300-nm-diameter SPBs in application of the rotating external magnetic field, although the dynamics of such beads is not well observable under an optical microscope.

Experimental
The experiments were performed using two suspensions of super-paramagnetic particles     achieved higher transmittance by geometrical shadowing effects [17,18].  Hz. The chain tended to become shorter when higher frequency was used.
Although there was a distinction of transmittance behavior between two kinds of beads, ΔT as a function of frequency made no difference between sorts of beads as shown in figure 6. Once the frequency reached threshold, ΔT dramatically dropped and eventually reached to zero. At this moment, it was thought that none of magnetically induced chain could be remained as a linear chain. Based on the optical images in figure 7, the length of magnetically induced chains became shorter as frequency increased. Ideally, by knowing the frequency at ΔT = 0, we could estimate dipole-dipole interaction of the magnetic beads we used. A considerable application of this mean to bioassay would be to determine the biorecognition forces among biomolecuels.

Conclusion
We have investigated the effect of the composition of magnetic bead on optical transmission of light passing through an aqueous solution containing the SPBs as a function of the external magnetic field and the orientation of magnetically induced bead chains. For the solution containing 2.8-μm-diameter SPBs, the transmission increased as the orientation of the chains became parallel to the direction of the incident light. On the contrary, the transmission declined for the aqueous solution containing 250-nm-diameter SPBs under the same condition. Despite the fact that transmission behaviors made distinction, when focusing on the difference between maximum and minimum transmission, ΔT, as function of frequency, we noticed that ΔT reached to zero at higher frequency. Based on results, our procedure enables to monitor the dynamics of sub-300-nm-diameter SPBs without using any expensive, difficult, and unwieldy experimental equipments, although the dynamics of such beads is not well observable under an optical microscope. Thus, our protocol has a great potential to apply into the bioassay field.