Microfluidics as an emerging paradigm for assisted reproductive technology: A sperm separation perspective.

Millions of people are subject to infertility worldwide and one in every six people, regardless of gender, experiences infertility at some period in their life, according to the World Health Organization. Assisted reproductive technologies are defined as a set of procedures that can address the infertility issue among couples, culminating in the alleviation of the condition. However, the costly conventional procedures of assisted reproduction and the inherent vagaries of the processes involved represent a setback for its successful implementation. Microfluidics, an emerging tool for processing low-volume samples, have recently started to play a role in infertility diagnosis and treatment. Given its host of benefits, including manipulating cells at the microscale, repeatability, automation, and superior biocompatibility, microfluidics have been adopted for various procedures in assisted reproduction, ranging from sperm sorting and analysis to more advanced processes such as IVF-on-a-chip. In this review, we try to adopt a more holistic approach and cover different uses of microfluidics for a variety of applications, specifically aimed at sperm separation and analysis. We present various sperm separation microfluidic techniques, categorized as natural and non-natural methods. A few of the recent developments in on-chip fertilization are also discussed.

Electric Field-Assisted Molecularly Imprinted Polymer-Modified QCM Sensor for Enhanced Detection of Immunoglobulin.

In this study, an electric-field-assisted molecularly imprinted polymer (EFAMIP) as an enhanced form of MIP was developed to improve the MIP-modified quartz crystal microbalance (QCM) biosensors. While exerting a vertical electric field, polymerization of methacrylic acid in the presence of immunoglobulin G (IgG) as the template was initiated, and later, after the template removal process, the EFAMIPs were obtained. The polymer surface characterization was conducted by using a scanning electron microscope. The impact of electric field direction on IgG binding sites, forming either EFAMIP-Fab or EFAMIP-Fc, was assessed. Next, the static measurement results in liquid for EFAMIP-modified QCM and MIP-modified QCM were compared. While encompassing IgG, EFAMIP-modified QCMs exhibited up to a 113.5% higher frequency shift than typical MIP in time-limited detection. The final frequency shift of EFAMIP, which determines the detection limit of IgG, was improved up to 12.5% compared to typical MIP. Moreover, the EFAMIP-Fab performance was promising for the selective detection of IgG in a solution containing different types of immunoglobulins.

Bio-inspired progressive motile sperm separation using joint rheotaxis and boundary-following behavior.

Infertility, as a daunting ever-increasing challenge, poses a worldwide issue to both couples and the healthcare sector. According to the World Health Organization, half of infertility cases are attributed to male factor infertility, either partly or completely. Semen parameters of concern including sperm count, morphology, and motility are deemed to play a vital role in the insemination process. Density gradient centrifugation, being a clinically established procedure for improving on the mentioned parameters, has long been proven to inflict damage on the DNA content of the sperm cells, inducing DNA fragmentation. Herein, a bio-inspired microfluidic device is proposed that capitalizes on the geometry of the uterotubal junction (UTJ) of the female reproductive tract, which can act as a rheological barrier. The device leverages sperm rheotaxis and boundary-following behavior which have been considered as major migratory mechanisms used by sperm during the fertilization process in the female body. The device consists of a series of parallel channels that guide progressive motile sperms into the main sorting channel, where the hydrodynamic barriers created by two consecutive UTJ-like constrictions select sperms based on their propulsive velocity and linearity of motion. The sequential sorting employed here allows for the fractionation of the sperm population into two subpopulations with varying degrees of motility. Both sorted populations showed a significant increase in straight line velocity, reaching 63.4 ± 14.4 µm s-1 and 74 ± 13.8 µm s-1 in the first and second pools, respectively from 35.2 ± 27.2 µm s-1 in raw semen. Additionally, sorted populations demonstrated over 30% reduction in DNA fragmentation index, an indication that the proposed device selects for undamaged sperms with high quality. Apart from the biological superiority of the sorted sperms, this device presents itself as an easy and clinically-applicable method for the separation of progressive motile sperms, while at the same time, benefiting from a straightforward procedure for sperm retrieval.

High-Efficiency Inertial Separation of Microparticles Using Elevated Columned Reservoirs and Vortex Technique for Lab-on-a-Chip Applications.

The discovery of circulating tumor cells (CTCs) has envisioned an excellent outlook for cancer diagnosis and prognosis. Among numerous efforts proposed for CTCs isolation, vortex separation is a well-known method for capturing CTCs from blood due to its applicability, low sample volume requirement, and ability to retain cell viability. It is a label-free, passive, low-cost, and automated method, making it an ideal solution for lab-on-a-chip applications. The previous designs that employed vortex technology have shown reaching high throughput and 70% separation efficiency although it was after three processing cycles which are not desired. Inspired by our earlier design, in this work, we redesigned the chip geometry by elevating the columned reservoir height to capture more particles and consequently reduce particle-particle collision, eventually improving efficiency. So, a height-variable chip with fewer elevated columned reservoirs (ECRs) was employed to isolate 20 µm microparticles representing CTCs from 8 µm microparticles. Also, numerical simulations were conducted to investigate the third axis contribution to the separation mechanism. The new design with ECRs resulted in a 14% increase in average efficiency, reaching ∼80% ± 8.3% in microparticle separation and 61% purity. Moreover, the proposed chip geometry demonstrated more than three times higher capacity in retaining orbiting particles up to 1300 in peak performance without sacrificing efficiency compared to earlier single-layer designs. We came up with an upgraded injection system to facilitate this chip characterization. We also presented an effortless and straightforward approach for purging air bubbles trapped inside the reservoirs to preserve regular chip operation, especially where there is a mismatch between channel and reservoir heights.

Ultrasensitive miniaturized planar microwave sensor for characterization of water-alcohol mixtures.

Designing a low-cost, compact, yet sensitive planar microwave sensor for complex permittivity measurement is highly desired for numerous applications though quite challenging. Here, in this research, an ultrasensitive planar microwave sensor is proposed which is based on an electric LC structure. The core sensor was fabricated on an FR-4 substrate using a simple fabrication process, then integrated within a polymethylmethacrylate microfluidic channel for straightforward liquid delivery to the sensing region. The resonance frequency of the bare sensor was designed to occur at 4.14 GHz while empty and shifted to 0.88 GHz when deionized water flows into the channel. The sensor response has been characterized for different mixture ratios of methanol and ethanol with deionized water. Next, the complex permittivity of the resulted binary mixtures has been extracted by the Debye model through a least square fitting method. The calculated average sensitivity is 1.45% which stands above most of sensors reported in the literature. Besides, the sensor has a small footprint with dimensions of 3.6 × 3.8 mm[Formula: see text] making it a suitable candidate for integration with point-of-care testing devices.

Nonlinear dielectric spectroscopy biosensor for SARS-CoV-2 detection.

The coronavirus disease caused by the SARS-CoV-2 virus has affected people worldwide for more than two years. Here we present a new diagnostic method based on nonlinear dielectric spectroscopy to detect the presence of the SARS-CoV-2 virus in swab samples. A known current is injected into the virus sample suspension, and the biomarker is the third harmonic detected in the power spectrum of the recorded signal. Computational modeling of harmonic production supports the hypothesis of ion channels (the E-protein) with nonlinear current-voltage characteristics being present on the virus envelope as a possible origin of harmonics. The developed system is able to distinguish between positive and negative samples with 5-10 dBc (decibels relative to the carrier) higher third harmonic ratios in positive samples, in agreement with the computational estimation. Our early results demonstrate that this method can detect the virus in solution. This is the first time harmonic signatures are used to detect SARS-CoV-2 in swab samples.

Simulation and fabrication of an integrating well-aligned silicon nanowires substrate for trapping circulating tumor cells labeled with Fe3O4 nanoparticles in a microfluidic device.

Introduction: Circulating tumor cells (CTCs) are the transformed tumor cells that can penetrate into the bloodstream and are available at concentrations as low as 1-100 cells per milliliter. To trap CTCs in the blood, one valid and mature technique that has been developed is the magnetophoresis-based separation in a microfluidic channel. Recently, nanostructured platforms have also been developed to trap specific targeted and marker cells in the blood. We aimed to integrate both in one platform to improve trapping. Methods: Here, we developed a numerical scheme and an integrated device that considered the interaction between drag and magnetic forces on paramagnetic labeled cells in the fluid as well as interaction of these two forces with the adhesive force and the surface friction of the nanowires substrate. We aimed on developing a more advanced technique that integrated the magnetophoretic property of some Fe3O4 paramagnetic nanoparticles (PMNPs) with a silicon nanowires (SiNWs) substrate in a microfluidic device to trap MDA-MB231 cell lines as CTCs in the blood. Results: Simulation indicated assuming that the nanoparticles adhere perfectly to the white blood cells (WBCs) and the CTCs, the magnetic moment of the CTCs was almost one order of magnitude larger than that of the WBCs, so its attraction by the magnetic field was much higher. In general with significant statistics, the integrated device can trap almost all of the CTCs on the SiNWs substrate. In the experimental section, we took advantage of the integrated trapping techniques, including micropost barriers, magnetophoresis, and nanowires-based substrate to more effectively isolate the CTCs. Conclusion: The simulation indicated that the proposed device could almost trap all of the CTCs onto the SiNWs substrate, whereas trapping in flat substrates with magnetophoretic force was very low. As a result of the magnetic field gradient, magnetophoretic force was applied to the cells through the nanoparticles, which would efficiently drive down the nanoparticle-tagged cells. For the experimental validation, anti-EpCAM antibodies for specific binding to tumor cells were used. Using this specific targeting method and by statistically counting, it was shown that the proposed technique has excellent performance and results in the trapping efficiency of above 90%.

Corkscrew point spread function for far-field three-dimensional nanoscale localization of pointlike objects.

We describe the corkscrew point spread function (PSF), which can localize objects in three dimensions throughout a 3.2 µm depth of field with nanometer precision. The corkscrew PSF rotates as a function of the axial (z) position of an emitter. Fisher information calculations show that the corkscrew PSF can achieve nanometer localization precision with limited numbers of photons. We demonstrate three-dimensional super-resolution microscopy with the corkscrew PSF by imaging beads on the surface of a triangular polydimethylsiloxane (PDMS) grating. With 99,000 photons detected, the corkscrew PSF achieves a localization precision of 2.7 nm in x, 2.1 nm in y, and 5.7 nm in z.

Three-dimensional localization precision of the double-helix point spread function versus astigmatism and biplane.

Wide-field microscopy with a double-helix point spread function (DH-PSF) provides three-dimensional (3D) position information beyond the optical diffraction limit. We compare the theoretical localization precision for an unbiased estimator of the DH-PSF to that for 3D localization by astigmatic and biplane imaging using Fisher information analysis including pixelation and varying levels of background. The DH-PSF results in almost constant localization precision in all three dimensions for a 2 µm thick depth of field while astigmatism and biplane improve the axial localization precision over smaller axial ranges. For high signal-to-background ratio, the DH-PSF on average achieves better localization precision.

Three-dimensional tracking of single mRNA particles in Saccharomyces cerevisiae using a double-helix point spread function.

Optical imaging of single biomolecules and complexes in living cells provides a useful window into cellular processes. However, the three-dimensional dynamics of most important biomolecules in living cells remains essentially uncharacterized. The precise subcellular localization of mRNA-protein complexes plays a critical role in the spatial and temporal control of gene expression, and a full understanding of the control of gene expression requires precise characterization of mRNA transport dynamics beyond the optical diffraction limit. In this paper, we describe three-dimensional tracking of single mRNA particles with 25-nm precision in the x and y dimensions and 50-nm precision in the z dimension in live budding yeast cells using a microscope with a double-helix point spread function. Two statistical methods to detect intermittently confined and directed transport were used to quantify the three-dimensional trajectories of mRNA for the first time, using ARG3 mRNA as a model. Measurements and analysis show that the dynamics of ARG3 mRNA molecules are mostly diffusive, although periods of non-Brownian confinement and directed transport are observed. The quantitative methods detailed in this paper can be broadly applied to the study of mRNA localization and the dynamics of diverse other biomolecules in a wide variety of cell types.

In vivo Three-Dimensional Superresolution Fluorescence Tracking using a Double-Helix Point Spread Function.

The point spread function (PSF) of a widefield fluorescence microscope is not suitable for three-dimensional super-resolution imaging. We characterize the localization precision of a unique method for 3D superresolution imaging featuring a double-helix point spread function (DH-PSF). The DH-PSF is designed to have two lobes that rotate about their midpoint in any transverse plane as a function of the axial position of the emitter. In effect, the PSF appears as a double helix in three dimensions. By comparing the Cramer-Rao bound of the DH-PSF with the standard PSF as a function of the axial position, we show that the DH-PSF has a higher and more uniform localization precision than the standard PSF throughout a 2 µm depth of field. Comparisons between the DH-PSF and other methods for 3D super-resolution are briefly discussed. We also illustrate the applicability of the DH-PSF for imaging weak emitters in biological systems by tracking the movement of quantum dots in glycerol and in live cells.

Localizing and tracking single nanoscale emitters in three dimensions with high spatiotemporal resolution using a double-helix point spread function.

Three-dimensional nanoscale localization and tracking of dim single emitters can be obtained with a widefield fluorescence microscope exhibiting a double-helix point spread function (DH-PSF). We describe in detail how the localization precision quantitatively depends upon the number of photons detected and the z position of the nanoscale emitter, thereby showing a approximately 10 nm localization capability along x, y, and z in the limit of weak emitters. Experimental measurements are compared to Fisher information calculations of the ultimate localization precision inherent in the DH-PSF. The DH-PSF, for the first time, is used to track single quantum dots in aqueous solution and a quantum dot-labeled structure inside a living cell in three dimensions.

Very-high-resolution tandem Fabry-Perot etalon cylindrical beam volume hologram spectrometer for diffuse source spectroscopy.

We demonstrate a compact and slitless spectrometer with high resolution formed by cascading a Fabry-Perot etalon (FPE) and a cylindrical beam volume hologram (CBVH). The most significant advantage of this combined spectrometer is that we can independently encode spectral information of a diffuse beam in a 2D plane. Also, we show that in this slitless configuration we can simultaneously benefit from the advantages of both elements: the high resolution of the FPE and the large spectral range of the CBVH. Here, we report on the experimental demonstration of a spectrometer with better than 0.2 nm resolution.

Model for efficient simulation of spatially incoherent light using the Wiener chaos expansion method.

We demonstrate a new and efficient technique for modeling and simulation of spatially incoherent sources using the Wiener chaos expansion method. By implementing this new model, we show that a practical-size photonic structure with a spatially incoherent input source can be analyzed more than 2 orders of magnitude faster compared with the conventional models without sacrificing the accuracy.

Investigation of physical mechanisms in coupling photonic crystal waveguiding structures.

We explain the fundamental physical mechanisms involved in coupling triangular lattice photonic crystal waveguides to conventional dielectric slab waveguides. We show that the two waveguides can be efficiently coupled outside the mode gap frequencies. We especially focus on the coupling of the two structures within the mode gap frequencies and show for the first time that the diffraction from the main photonic crystal structure plays an important role on the reflection of power back into the slab waveguide. The practical importance of this effect and possible strategies to modify it are also discussed.