Microfluidic devices

Malaria is an ancient lethargic disease that remains a global burden. It has been difficult to end the scourge of P. falciparum malaria because of the parasites’ drug resistance so early diagnosis of malaria is crucial. Microscopy remains the gold standard but has limited reliability in detecting malaria parasites. This study proffered a method towards detection of low parasitemia P. falciparum infected RBCs (Pf-RBCs) based on dielectrophoresis (DEP). A microfluidic device was designed for label-free cell sorting of Pf-RBCs from other whole blood in a continuous manner, based on the intrinsic electrical signatures of the cells. The design was validated by a finite element simulation using COMSOL Multiphysics. Simulations show the feasibility of the separation in a 9-mm long microfluidic channel under laminar flow conditions, using a low voltage supply of +/-10 V at 50 kHz.

Exosomes have gained recognition in cancer diagnostics and therapeutics. Most exosome isolation methods are time-consuming, costly and require bulky equipment, rendering them unsuitable for point-of-care (POC) settings. Microfluidics can be the key to solving these challenges. Here, we employ the development of a double filtration microfluidic device that can rapidly isolate exosomes in POC settings. The device can efficiently isolate exosomes from just 100 uL of plasma within 50 minutes. The device was compared against Polyethylene glycol (PEG) based precipitation, and findings show that both methods yield comparable exosome sizes and purity, but the device can detect exosomal miRNA earlier than PEG. Finally, a comparative analysis of membrane filters with exosomes collected from pore sizes 15 nm and 30 nm showed a similarity in exosome size and miRNA expressions, with significantly increased sample purity. These findings suggest that this device has potential in POC settings.

The ultimate challenge for assisted reproductive technologies (ARTs) is to select the most competent sperm population from a semen sample in an efficient way. In this thesis, we report on an effective sperm sorting microfluidic device that exploits the rheotaxis of sperm and investigates the sperm quality sorted under various flow conditions. Rheotaxis is the ability of a sperm cell to orient itself in the direction of the flow and swim against it. We developed a novel passively driven pumping system that provides a steady flow rate while it requires no external power source. We have also developed another rheotaxis-based microfluidic device that washes out the raw semen sample from any dead or less motile sperm. The device consists of a collection and waste chamber. To evaluate the effect of the shape and height of the collection chamber, we measured the sperm motility and velocity parameters after sorting using varying the shape and height of the collection chamber. We demonstrated that sperm selected with all devices have higher motility, normal morphology, and a fewer degree of DNA fragmentation compared to a control group.

Zika virus (ZIKV) is an emerging flavivirus transmitted to humans by Aedes mosquitos. ZIKV can be transmitted from mother to fetus during pregnancy and can cause microcephaly and other birth defects. Effective vaccines for Zika are yet to be approved. Detection of the ZIKV is based on serological testing that often shows cross-reactivity with the Dengue virus (DENV) and other flaviviruses. Currently, identification of ZIKV infection is usually done by i) testing the patient’s serum sample to detect ZIKV RNA using reverse transcriptase-polymerase chain reaction (RT-PCR), ii) testing patient’s serum sample for the presence of the NS1 protein antigen or iii) serological assays to determine the presence of virus-specific immunoglobin antibodies (IgG and IgM) by the use of ELISA assay. But ELISA-based assays show cross-reactivity and poor sensitivity. The gold standard for ZIKV RNA detection is RT-PCR, involves expensive medical facilities and skillful technicians. However, the plaque reduction neutralization test are executed to quantity neutralizing antibodies of the virus-but show high accuracy only after day 7 of the disease onset. Therefore, the development of POC assays which has the ASSURED (affordability, sensitivity, specificity, user-friendly, rapid and robust, equipment-free and deliverable) criteria defined by the World Health Organization are topmost priority. The core objective of this thesis is to find inexpensive, sensitive, precise, and fast assays for the specific diagnosis of ZIKV suitable for resource-constrained settings.

This thesis aims to address the challenges of the development of cost-effective and rapid assays for the accurate counting of CD4+ T cells and quantification of HIV-1 viral load for resource-constrained settings. The lack of such assays has severely affected people living in disease prevalent areas. CD4+ T cells count information plays a vital role in the effective management of HIV-1 disease. Here, we present a flow-free magnetic actuation platform that uses antibody-coated magnetic beads to efficiently capture CD4+ T cells from a 30 μL drop of whole blood. On-chip cell lysate electrical impedance spectroscopy has been utilized to quantify the isolated CD4 cells. The developed assay has a limit of detection of 25 cells per μL and provides accurate CD4 counts in the range of 25–800 cells per μL. The whole immunoassay along with the enumeration process is very rapid and provides CD4 quantification results within 5 min time frame. The assay does not require off-chip sample preparation steps and minimizes human involvement to a greater extent. The developed impedance-based immunoassay has the potential to significantly improve the CD4 enumeration process especially for POC settings.

Droplet microfluidics generates and manipulates microdroplets in microfluidic devices at high manufacturing efficiency and controllability. Microdroplets have proven effective in biomedical applications such as single-cell analysis, DNA sequencing, protein partitioning and drug delivery. Conventionally, a series of aqueous microdroplets containing biosamples is generated and controlled in an oil environment. One of the critical challenges in this system is that recovery of the aqueous samples from the oil phase is very difficult and often requires expensive and cumbersome post-processing. Also, the low Reynolds (Re) number characteristic of this system results in low throughput of droplet generation. To circumvent challenges and fully utilize microdroplets for practical clinical applications, this research aims to unpack the fundamental physics that governs droplet generation in oil-free systems including an aqueous two-phase system (ATPS) and a high inertial liquid-gas system.

This thesis was about finding a recovery method for TiO2, using a TiO2 recovery technology, which was high enough to be economical ($10 - $15 per 1,000 gallons) to be adopted by wastewater treatment plants. When comparing recovery technologies, the top three which were investigated further through experimentation were a centrifuge, sedimentation tank, and microfilter membrane. Upon experimentation and research, the TiO2 recovery efficiencies of these technologies were 99.5%, 92.5%, and 96.3%, respectively. When doing economic analysis on these technologies comparing TiO2 efficiencies and capital and operational costs, the centrifuge was the most preferred economic option. Also, its cost did were in the economical range ($10 - $15/1,000 gallons) which makes even this technology economical. Besides that, important and valuable information about TiO2: settling behavior, particle size and zeta potential, interactions with COD, and filter operations (particle characterization) were discovered for future research and future testing on this issue.