Rashid Bashir

Nanopore sensors for nucleic acid analysis

Nanopore analysis is an emerging technique that involves using a voltage to drive molecules through a nanoscale pore in a membrane between two electrolytes, and monitoring how the ionic current through the nanopore changes as single molecules pass through it. This approach allows charged polymers (including single-stranded DNA, double-stranded DNA and RNA) to be analysed with subnanometre resolution and without the need for labels or amplification. Recent advances suggest that nanopore-based sensors could be competitive with other third-generation DNA sequencing technologies, and may be able to rapidly and reliably sequence the human genome for under

Single virus particle mass detection using microresonators with nanoscale thickness

1,000. In this article we review the use of nanopore technology in DNA sequencing, genetics and medical diagnostics.

Micromechanical cantilever as an ultrasensitive pH microsensor

In this letter, we present the microfabrication and application of arrays of silicon cantilever beams as microresonator sensors with nanoscale thickness to detect the mass of individual virus particles. The dimensions of the fabricated cantilever beams were in the range of 4–5 μm in length, 1–2 μm in width and 20–30 nm in thickness. The virus particles we used in the study were vaccinia virus, which is a member of the Poxviridae family and forms the basis of the smallpox vaccine. The frequency spectra of the cantilever beams, due to thermal and ambient noise, were measured using a laser Doppler vibrometer under ambient conditions. The change in resonant frequency as a function of the virus particle mass binding on the cantilever beam surface forms the basis of the detection scheme. We have demonstrated the detection of a single vaccinia virus particle with an average mass of 9.5 fg. These devices can be very useful as components of biosensors for the detection of airborne virus particles.

Dielectrophoretic separation and manipulation of live and heat-treated cells of Listeria on microfabricated devices with interdigitated electrodes

We report on a pH sensor with ultrahigh sensitivity based on a microcantilever structure with a lithographically-defined crosslinked copolymeric hydrogel. Silicon-on-insulator wafers were used to fabricate cantilevers on which a polymer consisting of poly(methacrylic acid) (PMAA) with poly(ethylene glycol) dimethacrylate was patterned using free-radical UV polymerization. As the pH around the cantilever was increased above the pKa of PMAA, the polymer network expanded and resulted in a reversible change in surface stress causing the microcantilever to bend. Excellent mechanical amplification of polymer swelling as a function of pH change within the dynamic range was obtained, with a maximum deflection sensitivity of 1 nm/5×10−5 ΔpH.

3D Biofabrication Strategies for Tissue Engineering and Regenerative Medicine

Abstract Dielectrophoresis, the movement of particles in non-uniform ac electric field, was used to separate live and heat-treated Listeria innocua cells with great efficiency on the micro-fabricated devices with interdigitated electrodes by utilizing the difference of dielectric properties between alive and dead cells. Both live and dead cells are found to be only able to collect either at the centers of the electrodes in negative dielectrophoresis or at the electrode edges in positive dielectrophoresis due to the dielectrophoretic force and electrohydrodynamic force. Cell viability was verified by a rapid method using epifluorescence staining. The dependency of the applied ac signal’s frequency on the dielectrophoretic properties of Listeria cells is studied and discussed. This on-electrode manipulation and separation of cells can prove to be useful in micro-scale sample preparation and diagnostic applications in biochips.

Optical measurement of cycle-dependent cell growth

Over the past several decades, there has been an ever-increasing demand for organ transplants. However, there is a severe shortage of donor organs, and as a result of the increasing demand, the gap between supply and demand continues to widen. A potential solution to this problem is to grow or fabricate organs using biomaterial scaffolds and a persons own cells. Although the realization of this solution has been limited, the development of new biofabrication approaches has made it more realistic. This review provides an overview of natural and synthetic biomaterials that have been used for organ/tissue development. It then discusses past and current biofabrication techniques, with a brief explanation of the state of the art. Finally, the review highlights the need for combining vascularization strategies with current biofabrication techniques. Given the multitude of applications of biofabrication technologies, from organ/tissue development to drug discovery/screening to development of complex in vitro models of human diseases, these manufacturing technologies can have a significant impact on the future of medicine and health care.

Stacked Graphene-Al2O3 Nanopore Sensors for Sensitive Detection of DNA and DNA–Protein Complexes

Determining the growth patterns of single cells offers answers to some of the most elusive questions in contemporary cell biology: how cell growth is regulated and how cell size distributions are maintained. For example, a linear growth in time implies that there is no regulation required to maintain homeostasis; an exponential pattern indicates the opposite. Recently, there has been great effort to measure single cells using microelectromechanical systems technology, and several important questions have been explored. However, a unified, easy-to-use methodology to measure the growth rate of individual adherent cells of various sizes has been lacking. Here we demonstrate that a newly developed optical interferometric technique, known as spatial light interference microscopy, can measure the cell dry mass of many individual adherent cells in various conditions, over spatial scales from micrometers to millimeters, temporal scales ranging from seconds to days, and cell types ranging from bacteria to mammalian cells. We found evidence of exponential growth in Escherichia coli, which agrees very well with other recent reports. Perhaps most importantly, combining spatial light interference microscopy with fluorescence imaging provides a unique method for studying cell cycle-dependent growth. Thus, by using a fluorescent reporter for the S phase, we measured single cell growth over each phase of the cell cycle in human osteosarcoma U2OS cells and found that the G2 phase exhibits the highest growth rate, which is mass-dependent and can be approximated by an exponential.

Microfluidic Biochip for Impedance Spectroscopy of Biological Species

We report the development of a multilayered graphene-Al(2)O(3) nanopore platform for the sensitive detection of DNA and DNA-protein complexes. Graphene-Al(2)O(3) nanolaminate membranes are formed by sequentially depositing layers of graphene and Al(2)O(3), with nanopores being formed in these membranes using an electron-beam sculpting process. The resulting nanopores are highly robust, exhibit low electrical noise (significantly lower than nanopores in pure graphene), are highly sensitive to electrolyte pH at low KCl concentrations (attributed to the high buffer capacity of Al(2)O(3)), and permit the electrical biasing of the embedded graphene electrode, thereby allowing for three terminal nanopore measurements. In proof-of-principle biomolecule sensing experiments, the folded and unfolded transport of single DNA molecules and RecA-coated DNA complexes could be discerned with high temporal resolution. The process described here also enables nanopore integration with new graphene-based structures, including nanoribbons and nanogaps, for single-molecule DNA sequencing and medical diagnostic applications.

Impedance microbiology-on-a-chip: microfluidic bioprocessor for rapid detection of bacterial metabolism

This paper describes the fabrication and characterization of a microelectronic device for the electrical interrogation and impedance spectroscopy of biological species. Key features of the device include an all top-side processing for the formation of fluidic channels, planar fluidic interface ports, integrated metal electrodes for impedance measurements, and a glass cover sealing the non-planar topography of the chip using spin-on-glass as an intermediate bonding layer. The total fluidic path volume in the device is on the order of 30 nl. Flow fields in the closed chip were mapped by particle image velocimetry. Electrical impedance measurements of suspensions of the live microorganism Listeria innocua injected into the chip demonstrate an easy method for detecting the viability of a few bacterial cells. By-products of the bacterial metabolism modify the ionic strength of a low conductivity suspension medium, significantly altering its electrical characteristics.

Ultrasensitive Biomems Sensors Based on Microcantilevers Patterned with Environmentally Responsive Hydrogels

Detection of a few live bacterial cells in many industrial or clinical samples is a very important technological problem. We have developed a microscale technique for concentrating bacterial cells from a dilute sample, by factors on the order of 10/sup 4/ to 10/sup 5/, and detecting their metabolic activity by purely electrical means. The technique was implemented on a silicon-based microfluidic chip where the cells are concentrated and incubated in a chamber with a volume of 400 pL. Concentration and capture are obtained by the use of dielectrophoresis on the bacterial cells, and metabolism detection is achieved by means of impedance measurements of the medium in which the bacteria are incubated. Performing impedance-based detection at the microscale results in drastically reduced detection times for dilute bacterial samples, thanks to the ability to efficiently concentrate and capture the cells in an extremely small volume. Such concentration eliminates the need to amplify the bacterial population by long culture steps. This detection technique can be used for a wide variety of applications.