Fatih Inci

Advances in Plasmonic Technologies for Point of Care Applications

Demand for accessible and affordable healthcare for infectious and chronic diseases present significant challenges for providing high-value and effective healthcare. Traditional approaches are expanding to include point-of-care (POC) diagnostics, bedside testing, and community-based approaches to respond to these challenges.1 Innovative solutions utilizing recent advances in mobile technologies, nanotechnology, imaging systems, and microfluidic technologies are envisioned to assist this transformation. Infectious diseases have considerable economic and societal impact on developing settings. For instance, malaria is observed more commonly in sub-Saharan Africa and India.2 The societal impact of acquired immune deficiency syndrome (AIDS) and tuberculosis is high, through targeting adults in villages and leaving behind declining populations.3 In resource-constrained settings, it is estimated that about 32% of the disease burden is from communicable diseases such as respiratory infections, AIDS, and malaria, while 43% of the burden is from noncommunicable diseases, such as cardiovascular diseases, neuropsychiatric conditions, and cancer.4 Developing diagnostic platforms that are affordable, robust, and rapid-targeting infectious diseases is one of the top priorities for improving healthcare delivery in the developing world.5 The early detection and monitoring of infectious diseases and cancer through affordable and accessible healthcare will significantly reduce the disease burden and help preserve the social fabric of these communities. Further, improved diagnostics and disease monitoring technologies have potential to enhance foreign investment, trade, and mobility in the developing countries.6 Highly sensitive and specific lab assays such as cell culture methods, polymerase chain reaction (PCR), and enzyme-linked immunosorbent assay (ELISA) are available for diagnosis of infectious diseases in the developed world. They require sample transportation, manual preparation steps, and skilled and well-trained technicians. These clinical conventional methods provide results in several hours to days, precluding rapid detection and response at the primary care settings. Another diagnostic challenge is identifying multiple pathogens. Since common symptoms like sore throat and fever can be caused by multiple infectious agents (e.g., bacteria and viruses), it is important to accurately identify the responsible agent for targeted treatment. Therefore, high-throughput sensors for multiplexed identification would help improve patient care.7 Medical instruments in centrally located institutions in the developed world rely on uninterrupted electricity and running water and require controlled environmental conditions. It may not be viable to satisfy some of these criteria in some POC settings, where well-trained healthcare personnel are not available and clean water access is unreliable.7,8 Further, in remote settings without infrastructure, rain and dust can act as contaminants.7 Diagnostic devices for POC testing in these settings are identified by the World Health Organization to be affordable, sensitive, user-friendly, specific to biological agents, and providing rapid response to small sample volumes.9 Optical biosensor devices are emerging as powerful biologic agent detection platforms satisfying these considerations.10 Optical sensing platforms employ various methods, including refractive index change monitoring, absorption, and spectroscopic-based measurements.11 Optical sensors that are based on refractive index monitoring cover a range of technologies, including photonic crystal fibers, nano/microring resonator structures, interferometric devices, plasmonic nano/micro arrays, and surface plasmon resonance (SPR)-based platforms.11,12 The latter two are plasmonic-based technologies. Plasmonics is an enabling optical technology with applications in disease monitoring, diagnostics, homeland security, food safety, and biological imaging applications. The plasmonic-based biosensor platforms along with the underlying technologies are illustrated in the Figure ​Figure1.1. Here, we reviewed SPR, localized surface plasmon resonance (LSPR), and large-scale plasmonic arrays (e.g., nanohole arrays). Figure 1 Plasmonic-based technologies for versatile biosensor applications. SPR stands for surface plasmon resonance, LSPR for localized surface plasmon resonance, SPRi for surface plasmon resonance imaging, and SERS for surface-enhanced Raman scattering.

Nanoplasmonic quantitative detection of intact viruses from unprocessed whole blood.

Infectious diseases such as HIV and hepatitis B pose an omnipresent threat to global health. Reliable, fast, accurate, and sensitive platforms that can be deployed at the point-of-care (POC) in multiple settings, such as airports and offices, for detection of infectious pathogens are essential for the management of epidemics and possible biological attacks. To the best of our knowledge, no viral load technology adaptable to the POC settings exists today due to critical technical and biological challenges. Here, we present for the first time a broadly applicable technology for quantitative, nanoplasmonic-based intact virus detection at clinically relevant concentrations. The sensing platform is based on unique nanoplasmonic properties of nanoparticles utilizing immobilized antibodies to selectively capture rapidly evolving viral subtypes. We demonstrate the capture, detection, and quantification of multiple HIV subtypes (A, B, C, D, E, G, and subtype panel) with high repeatability, sensitivity, and specificity down to 98 ± 39 copies/mL (i.e., HIV subtype D) using spiked whole blood samples and clinical discarded HIV-infected patient whole blood samples validated by the gold standard, i.e., RT-qPCR. This platform technology offers an assay time of 1 h and 10 min (1 h for capture, 10 min for detection and data analysis). The presented platform is also able to capture intact viruses at high efficiency using immuno-surface chemistry approaches directly from whole blood samples without any sample preprocessing steps such as spin-down or sorting. Evidence is presented showing the system to be accurate, repeatable, and reliable. Additionally, the presented platform technology can be broadly adapted to detect other pathogens having reasonably well-described biomarkers by adapting the surface chemistry. Thus, this broadly applicable detection platform holds great promise to be implemented at POC settings, hospitals, and primary care settings.

Paper and Flexible Substrates as Materials for Biosensing Platforms to Detect Multiple Biotargets

The need for sensitive, robust, portable, and inexpensive biosensing platforms is of significant interest in clinical applications for disease diagnosis and treatment monitoring at the point-of-care (POC) settings. Rapid, accurate POC diagnostic assays play a crucial role in developing countries, where there are limited laboratory infrastructure, trained personnel, and financial support. However, current diagnostic assays commonly require long assay time, sophisticated infrastructure and expensive reagents that are not compatible with resource-constrained settings. Although paper and flexible material-based platform technologies provide alternative approaches to develop POC diagnostic assays for broad applications in medicine, they have technical challenges integrating to different detection modalities. Here, we address the limited capability of current paper and flexible material-based platforms by integrating cellulose paper and flexible polyester films as diagnostic biosensing materials with various detection modalities through the development and validation of new widely applicable electrical and optical sensing mechanisms using antibodies and peptides. By incorporating these different detection modalities, we present selective and accurate capture and detection of multiple biotargets including viruses (Human Immunodeficieny Virus-1), bacteria (Escherichia coli and Staphylococcus aureus), and cells (CD4+ T lymphocytes) from fingerprick volume equivalent of multiple biological specimens such as whole blood, plasma, and peritoneal dialysis effluent with clinically relevant detection and sensitivity.

Point-of-care assays for tuberculosis: Role of nanotechnology/microfluidics

Tuberculosis (TB) remains one of the most devastating infectious diseases and its eradication is still unattainable given the limitations of current technologies for diagnosis, treatment and prevention. The World Health Organizations goal to eliminate TB globally by 2050 remains an ongoing challenge as delayed diagnosis and misdiagnosis of TB continue to fuel the worldwide epidemic. Despite considerable improvements in diagnostics for the last few decades, a simple and effective point-of-care TB diagnostic test is yet not available. Here, we review the current assays used for TB diagnosis, and highlight the recent advances in nanotechnology and microfluidics that potentially enable new approaches for TB diagnosis in resource-constrained settings.

Portable microfluidic chip for detection of Escherichia coli in produce and blood

Pathogenic agents can lead to severe clinical outcomes such as food poisoning, infection of open wounds, particularly in burn injuries and sepsis. Rapid detection of these pathogens can monitor these infections in a timely manner improving clinical outcomes. Conventional bacterial detection methods, such as agar plate culture or polymerase chain reaction, are time-consuming and dependent on complex and expensive instruments, which are not suitable for point-of-care (POC) settings. Therefore, there is an unmet need to develop a simple, rapid method for detection of pathogens such as Escherichia coli. Here, we present an immunobased microchip technology that can rapidly detect and quantify bacterial presence in various sources including physiologically relevant buffer solution (phosphate buffered saline [PBS]), blood, milk, and spinach. The microchip showed reliable capture of E. coli in PBS with an efficiency of 71.8% ± 5% at concentrations ranging from 50 to 4,000 CFUs/mL via lipopolysaccharide binding protein. The limits of detection of the microchip for PBS, blood, milk, and spinach samples were 50, 50, 50, and 500 CFUs/mL, respectively. The presented technology can be broadly applied to other pathogens at the POC, enabling various applications including surveillance of food supply and monitoring of bacteriology in patients with burn wounds.

Nanostructured optical photonic crystal biosensor for HIV viral load measurement

Detecting and quantifying biomarkers and viruses in biological samples have broad applications in early disease diagnosis and treatment monitoring. We have demonstrated a label-free optical sensing mechanism using nanostructured photonic crystals (PC) to capture and quantify intact viruses (HIV-1) from biologically relevant samples. The nanostructured surface of the PC biosensor resonantly reflects a narrow wavelength band during illumination with a broadband light source. Surface-adsorbed biotarget induces a shift in the resonant Peak Wavelength Value (PWV) that is detectable with <10 pm wavelength resolution, enabling detection of both biomolecular layers and small number of viruses that sparsely populate the transducer surface. We have successfully captured and detected HIV-1 in serum and phosphate buffered saline (PBS) samples with viral loads ranging from 104 to 108 copies/mL. The surface density of immobilized biomolecular layers used in the sensor functionalization process, including 3-mercaptopropyltrimethoxysilane (3-MPS), N-gamma-Maleimidobutyryl-oxysuccinimide ester (GMBS), NeutrAvidin, anti-gp120, and bovine serum albumin (BSA) were also quantified by the PC biosensor.

Multiscale assembly for tissue engineering and regenerative medicine

Our understanding of cell biology and its integration with materials science has led to technological innovations in the bioengineering of tissue-mimicking grafts that can be utilized in clinical and pharmaceutical applications. Bioengineering of native-like multiscale building blocks provides refined control over the cellular microenvironment, thus enabling functional tissues. In this review, we focus on assembling building blocks from the biomolecular level to the millimeter scale. We also provide an overview of techniques for assembling molecules, cells, spheroids, and microgels and achieving bottom-up tissue engineering. Additionally, we discuss driving mechanisms for self- and guided assembly to create micro-to-macro scale tissue structures.

Towards artificial tissue models: past, present, and future of 3D bioprinting.

Regenerative medicine and tissue engineering have seen unprecedented growth in the past decade, driving the field of artificial tissue models towards a revolution in future medicine. Major progress has been achieved through the development of innovative biomanufacturing strategies to pattern and assemble cells and extracellular matrix (ECM) in three-dimensions (3D) to create functional tissue constructs. Bioprinting has emerged as a promising 3D biomanufacturing technology, enabling precise control over spatial and temporal distribution of cells and ECM. Bioprinting technology can be used to engineer artificial tissues and organs by producing scaffolds with controlled spatial heterogeneity of physical properties, cellular composition, and ECM organization. This innovative approach is increasingly utilized in biomedicine, and has potential to create artificial functional constructs for drug screening and toxicology research, as well as tissue and organ transplantation. Herein, we review the recent advances in bioprinting technologies and discuss current markets, approaches, and biomedical applications. We also present current challenges and provide future directions for bioprinting research.

Acute On‐Chip HIV Detection Through Label‐Free Electrical Sensing of Viral Nano‐Lysate

Development of portable biosensors has broad applications in environmental monitoring, clinical diagnosis, public health, and homeland security. There is an unmet need for pathogen detection at the point-of-care (POC) using a fast, sensitive, inexpensive, and easy-to-use method that does not require complex infrastructure and well-trained technicians. For instance, detection of Human Immunodeficiency Virus (HIV-1) at acute infection stage has been challenging, since current antibody-based POC technologies are not effective due to low concentration of antibodies. In this study, we demonstrated for the first time a label-free electrical sensing method that can detect lysed viruses, i.e. viral nano-lysate, through impedance analysis, offering an alternative technology to the antibody-based methods such as dipsticks and Enzyme-linked Immunosorbent Assay (ELISA). The presented method is a broadly applicable platform technology that can potentially be adapted to detect multiple pathogens utilizing impedance spectroscopy for other infectious diseases including herpes, influenza, hepatitis, pox, malaria, and tuberculosis. The presented method offers a rapid and portable tool that can be used as a detection technology at the POC in resource-constrained settings, as well as hospital and primary care settings.

Efficient on-chip isolation of HIV subtypes

HIV has caused a global pandemic over the last three decades. There is an unmet need to develop point-of-care (POC) viral load diagnostics to initiate and monitor antiretroviral treatment in resource-constrained settings. Particularly, geographical distribution of HIV subtypes poses significant challenges for POC immunoassays. Here, we demonstrated a microfluidic device that can effectively capture various subtypes of HIV particles through anti-gp120 antibodies, which were immobilized on the microchannel surface. We first optimized an antibody immobilization process using fluorescent antibodies, quantum dot staining and AFM studies. The results showed that anti-gp120 antibodies were immobilized on the microchannel surface with an elevated antibody density and uniform antibody orientation using a Protein G-based surface chemistry. Further, RT-qPCR analysis showed that HIV particles of subtypes A, B and C were captured repeatably with high efficiencies of 77.2 ± 13.2%, 82.1 ± 18.8, and 80.9 ± 14.0% from culture supernatant, and 73.2 ± 13.6, 74.4 ± 14.6 and 78.3 ± 13.3% from spiked whole blood at a viral load of 1000 copies per mL, respectively. HIV particles of subtypes A, B and C were captured with high efficiencies of 81.8 ± 9.4%, 72.5 ± 18.7, and 87.8 ± 3.2% from culture supernatant, and 74.6 ± 12.9, 75.5 ± 6.7 and 69.7 ± 9.5% from spiked whole blood at a viral load of 10,000 copies per mL, respectively. The presented immuno-sensing device enables the development of POC on-chip technologies to monitor viral load and guide antiretroviral treatment (ART) in resource-constrained settings.