Leyla Esfandiari

Sequence-specific DNA detection at 10 fM by electromechanical signal transduction.

Target DNA fragments at 10 fM concentration (approximately 6 × 105 molecules) were detected against a DNA background simulating the noncomplementary genomic DNA present in real samples using a simple, PCR-free, optics-free approach based on electromechanical signal transduction. The development of a rapid, sensitive, and cost-effective nucleic acid detection platform is highly desired for a range of diverse applications. We previously described a potentially low-cost device for sequence-specific nucleic acid detection based on conductance change measurement of a pore blocked by electrophoretically mobilized bead-(peptide nucleic acid probe) conjugates upon hybridization with target nucleic acid. Here, we demonstrate the operation of our device with longer DNA targets, and we describe the resulting improvement in the limit of detection (LOD). We investigated the detection of DNA oligomers of 110, 235, 419, and 1613 nucleotides at 1 pM to 1 fM and found that the LOD decreased as DNA length increased, with 419 and 1613 nucleotide oligomers detectable down to 10 fM. In addition, no false positive responses were obtained with noncomplementary, control DNA fragments of similar length. The 1613-base DNA oligomer is similar in size to 16S rRNA, which suggests that our device may be useful for detection of pathogenic bacteria at clinically relevant concentrations based on recognition of species-specific 16S rRNA sequences.

Initial studies of mechanical compression on neurogenesis with neonatal neural stem cells

UNLABELLED In this article we demonstrate the effect of mechanical compression on the behavior of cultured neural stem cells using a microelectromechanical system platform. Polydimethylsiloxane (PDMS)-based stretchable substrates were used on a neurosphere (NS) assay to investigate the role of mechanical forces on the formation of radial glial processes and neuronal migration. To induce mechanical compression on NS, the PDMS culturing substrate was patterned with micron-sized wells. NS were cultured on the prestretched device. After 48 hours, when the NS had grown to the size of the wells width, the stretched substrate was released. The experimental results showed that applied mechanical compression on neural stem cells could be a factor accelerating the radial glial formation, which is associated with neurogenesis and neuronal migration. FROM THE CLINICAL EDITOR This study demonstrates that mechanical compression on neural stem cells could be a factor accelerating the radial glial formation, which is associated with neurogenesis and neuronal migration.

Cortisol extraction through human skin by reverse iontophoresis

Continuous monitoring of cortisol at the surface of the skin would advance the diagnosis and treatment of cortisol-related diseases, or of elevated cortisol levels related to stress in otherwise healthy populations. Reliable and accurate detection of cortisol at the skin surface remains a limiting factor in real-time monitoring of cortisol. To address this limitation, cortisol extraction through excised human skin by reverse iontophoresis was studied in vitro in side-by-side diffusion cells using a radiolabeled probe. The skin was subjected to four direct current regimens (0, 28, 56, 113μAcm-2) with the anode in the donor chamber and the cumulative cortisol concentrations recorded in the receiver chamber. The 56 and 113μAcm-2 regimens significantly increased transport of 3H-cortisol through the skin, and current density correlated directly with transcutaneous transport of 3H-cortisol. The threshold of detection of electroosmotic versus passive diffusion of cortisol through the skin was between 28 and 56μAcm-2. The results of this study are significant in examining how lipophilic analytes found in the bloodstream respond to reverse iontophoresis across the skin. In addition, a device integration technique is presented which illustrates how continuous cortisol extraction and sensing could potentially be achieved in a conventional wearable format.

PCR-Independent Detection of Bacterial Species-Specific 16S rRNA at 10 fM by a Pore-Blockage Sensor

A PCR-free, optics-free device is used for the detection of Escherichia coli (E. coli) 16S rRNA at 10 fM, which corresponds to ~100–1000 colony forming units/mL (CFU/mL) depending on cellular rRNA levels. The development of a rapid, sensitive, and cost-effective nucleic acid detection platform is sought for the detection of pathogenic microbes in food, water and body fluids. Since 16S rRNA sequences are species specific and are present at high copy number in viable cells, these nucleic acids offer an attractive target for microbial pathogen detection schemes. Here, target 16S rRNA of E. coli at 10 fM concentration was detected against a total RNA background using a conceptually simple approach based on electromechanical signal transduction, whereby a step change reduction in ionic current through a pore indicates blockage by an electrophoretically mobilized bead-peptide nucleic acid probe conjugate hybridized to target nucleic acid. We investigated the concentration detection limit for bacterial species-specific 16S rRNA at 1 pM to 1 fM and found a limit of detection of 10 fM for our device, which is consistent with our previous finding with single-stranded DNA of similar length. In addition, no false positive responses were obtained with control RNA and no false negatives with target 16S rRNA present down to the limit of detection (LOD) of 10 fM. Thus, this detection scheme shows promise for integration into portable, low-cost systems for rapid detection of pathogenic microbes in food, water and body fluids.

Simultaneous detection of multiple charged particles using a borosilicate nanopore-based sensor

Nanopore sensing has been widely researched owing to its single molecule sensitivity. In this work, we have demonstrated the potential application of a relatively low-cost single borosilicate nanopore-based sensor for simultaneous detection of multiple charged particles with various diameters.

Quantitative estimation of electro-osmosis force on charged particles inside a borosilicate resistive-pulse sensor

Nano and micron-scale pore sensors have been widely used for biomolecular sensing application due to its sensitive, label-free and potentially cost-effective criteria. Electrophoretic and electroosmosis are major forces which play significant roles on the sensors performance. In this work, we have developed a mathematical model based on experimental and simulation results of negatively charged particles passing through a 2μm diameter solid-state borosilicate pore under a constant applied electric field. The mathematical model has estimated the ratio of electroosmosis force to electrophoretic force on particles to be 77.5%.

PDMS stretchable platforms for the studies of mechanical compression on neurogenesis

The use of MEMS to study the effect of mechanical compression on neurogenesis has been demonstrated. Polydimethylsiloxane — (PDMS)-based stretchable platforms were used on neurosphere assay to investigate the role of mechanical forces on the formation of radial glial processes, neuronal generation and migration. To induce mechanical compression on neurospheres, The PDMS culturing substrate is patterned with micron-sized wells. Neurospheres are cultured on the prestretched device. After 48 hours, when neurospheres are grown to the size of the wells width, the stretched substrate is released. The experimental results showed that applied mechanical compression on neural stem cells cultured as neurospheres could be a factor accelerating the radial glial formation, which is associated to neurogenesis and neuronal migration.