Tohid Fatanat Didar

Patterning Multiplex Protein Microarrays in a Single Microfluidic Channel

The development of versatile biofunctional surfaces is a fundamental prerequisite in designing Lab on a Chip (LOC) devices for applications in biosensing interfaces and microbioreactors. The current paper presents a rapid combinatorial approach to create multiplex protein patterns in a single microfluidic channel. This approach consists of coupling microcontact printing with microfluidic patterning, where microcontact printing is employed for silanization using (3-Aminopropyl) triethoxysilane (APTES), followed by microfluidic patterning of multiple antibodies. As a result, the biomolecules of choice could be covalently attached to the microchannel surface, thus creating a durable and highly resistant functional interface. Moreover, the experimental procedure was designed to create a microfluidic platform that maintains functionality at high flow rates. The functionalized surfaces were characterized using X-ray photoelectron spectroscopy (XPS) and monitored with fluorescence microscopy at each step of functionalization. To illustrate the possibility of patterning multiple biomolecules along the cross section of a single microfluidic channel, microarrays of five different primary antibodies were patterned onto a single channel and their functionality was evaluated accordingly through a multiplex immunoassay using secondary antibodies specific to each patterned primary antibody. The resulting patterns remained stable at shear stresses of up to 50 dyn/cm(2). The overall findings suggest that the developed multiplex functional interface on a single channel can successfully lead to highly resistant multiplex functional surfaces for high throughput biological assays.

Characterization and modeling of 2D-glass micro-machining by spark-assisted chemical engraving (SACE) with constant velocity

Spark-assisted chemical engraving (SACE) is an unconventional micro-machining technology based on electrochemical discharge used for micro-machining nonconductive materials. SACE 2D micro-machining with constant speed was used to machine micro-channels in glass. Parameters affecting the quality and geometry of the micro-channels machined by SACE technology with constant velocity were presented and the effect of each of the parameters was assessed. The effect of chemical etching on the geometry of micro-channels under different machining conditions has been studied, and a model is proposed for characterization of the micro-channels as a function of machining voltage and applied speed.

Separation of rare oligodendrocyte progenitor cells from brain using a high-throughput multilayer thermoplastic-based microfluidic device

Despite the advances made in the field of regenerative medicine, the progress in cutting-edge technologies for separating target therapeutic cells are still at early stage of development. These cells are often rare, such as stem cells or progenitor cells that their overall properties should be maintained during the separation process for their subsequent application in regenerative medicine. This work, presents separation of oligodendrocyte progenitor cells (OPCs) from rat brain primary cultures using an integrated thermoplastic elastomeric (TPE)- based multilayer microfluidic device fabricated using hot-embossing technology. OPCs are frequently used in recovery, repair and regeneration of central nervous system after injuries. Indeed, their ability to differentiate in vitro into myelinating oligodendrocytes, are extremely important for myelin repair. OPCs form 5-10% of the glial cells population. The traditional macroscale techniques for OPCs separation require pre-processing of cells and/or multiple time consuming steps with low efficiency leading very often to alteration of their properties. The proposed methodology implies to separate OPCs based on their smaller size compared to other cells from the brain tissue mixture. Using aforementioned microfluidic chip embedded with a 5 μm membrane pore size and micropumping system, a separation efficiency more than 99% was achieved. This microchip was able to operate at flow rates up to 100 μl/min, capable of separating OPCs from a confluent 75 cm(2) cell culture flask in less than 10 min, which provides us with a high-throughput and highly efficient separation expected from any cell sorting techniques.

An omniphobic lubricant-infused coating produced by chemical vapor deposition of hydrophobic organosilanes attenuates clotting on catheter surfaces

AbstarctCatheter associated thrombosis is an ongoing problem. Omniphobic coatings based on tethering biocompatible liquid lubricants on self-assembled monolayers of hydrophobic organosilanes attenuate clotting on surfaces. Herein we report an efficient, non-invasive and robust process for coating catheters with an antithrombotic, omniphobic lubricant-infused coating produced using chemical vapor deposition (CVD) of hydrophobic fluorine-based organosilanes. Compared with uncoated catheters, CVD coated catheters significantly attenuated thrombosis via the contact pathway of coagulation. When compared with the commonly used technique of liquid phase deposition (LPD) of fluorine-based organosilanes, the CVD method was more efficient and reproducible, resulted in less disruption of the outer polymeric layer of the catheters and produced greater antithrombotic activity. Therefore, omniphobic coating of catheters using the CVD method is a simple, straightforward and non-invasive procedure. This method has the potential to not only prevent catheter thrombosis, but also to prevent thrombosis on other blood-contacting medical devices.

Sentinel Wraps: Real-Time Monitoring of Food Contamination by Printing DNAzyme Probes on Food Packaging

Here, we report the development of a transparent, durable, and flexible sensing surface that generates a fluorescence signal in the presence of a specific target bacterium. This material can be used in packaging, and it is capable of monitoring microbial contamination in various types of food products in real time without having to remove the sample or the sensor from the package. The sensor was fabricated by covalently attaching picoliter-sized microarrays of an E. coli-specific RNA-cleaving fluorogenic DNAzyme probe (RFD-EC1) to a thin, flexible, and transparent cyclo-olefin polymer (COP) film. Our experimental results demonstrate that the developed (RFD-EC1)-COP surface is specific, stable for at least 14 days under various pH conditions (pH 3-9), and can detect E. coli in meat and apple juice at concentrations as low as 103 CFU/mL. Furthermore, we demonstrate that our sensor is capable of detecting bacteria while still attached to the food package, which eliminates the need to manipulate the sample. The developed biosensors are stable for at least the shelf life of perishable packaged food products and provide a packaging solution for real-time monitoring of pathogens. These sensors hold the potential to make a significant contribution to the ongoing efforts to mitigate the negative public-health-related impacts of food-borne illnesses.

A Broad-Spectrum Infection Diagnostic that Detects Pathogen-Associated Molecular Patterns (PAMPs) in Whole Blood

Background Blood cultures, and molecular diagnostic tests that directly detect pathogen DNA in blood, fail to detect bloodstream infections in most infected patients. Thus, there is a need for a rapid test that can diagnose the presence of infection to triage patients, guide therapy, and decrease the incidence of sepsis. Methods An Enzyme-Linked Lectin-Sorbent Assay (ELLecSA) that uses magnetic microbeads coated with an engineered version of the human opsonin, Mannose Binding Lectin, containing the Fc immunoglobulin domain linked to its carbohydrate recognition domain (FcMBL) was developed to quantify pathogen-associated molecular patterns (PAMPs) in whole blood. This assay was tested in rats and pigs to explore whether it can detect infections and monitor disease progression, and in prospectively enrolled, emergency room patients with suspected sepsis. These results were also compared with data obtained from non-infected patients with or without traumatic injuries. Results The FcMBL ELLecSA was able to detect PAMPS present on, or released by, 85% of clinical isolates representing 47 of 55 different pathogen species, including the most common causes of sepsis. The PAMP assay rapidly (< 1 h) detected the presence of active infection in animals, even when blood cultures were negative and bacteriocidal antibiotics were administered. In patients with suspected sepsis, the FcMBL ELLecSA detected infection in 55 of 67 patients with high sensitivity (> 81%), specificity (> 89%), and diagnostic accuracy of 0·87. It also distinguished infection from trauma-related inflammation in the same patient cohorts with a higher specificity than the clinical sepsis biomarker, C-reactive Protein. Conclusion The FcMBL ELLecSA-based PAMP assay offers a rapid, simple, sensitive and specific method for diagnosing infections, even when blood cultures are negative and antibiotic therapy has been initiated. It may help to triage patients with suspected systemic infections, and serve as a companion diagnostic to guide administration of emerging dialysis-like sepsis therapies.

A Miniaturized Multipurpose Platform for Rapid, Label‐Free, and Simultaneous Separation, Patterning, and In Vitro Culture of Primary and Rare Cells

Given that current cell isolation techniques are expensive, time consuming, yield low isolation purities, and/or alter target cell properties, a versatile, cost effective, and easy-to-operate microchip with the capability to simultaneously separate, capture, pattern, and culture rare and primary cells in vitro is developed. The platform is based on target cell adhesion onto the micro-fabricated interfaces produced by microcontact printing of cell-specific antibodies. Results show over 95% separation efficiency in less than 10 min for the separation of oligodendrocyte progenitor cells (OPCs) and cardiomyocytes from rat brain and heart mixtures, respectively. Target cell attachment and single cell spreading can be precisely controlled on the basis of the designed patterns. Both cell types can maintain their biofunctionality. Indeed, isolated OPCs can proliferate and differentiate into mature oligodendrocytes, while isolated cardiomyocytes retain their contractile properties on the separation platform. Successful separation of two dissimilar cell types present in varying concentrations in their respective cell mixtures and the demonstration of their integrity after separation open new avenues for time and cost-effective sorting of various cell types using the developed miniaturized platform.

Generating 2-dimensional concentration gradients of biomolecules using a simple microfluidic design.

This study reports a microfluidic device for generating 2-dimensional concentration gradients of biomolecules along the width and length of a chamber and conventional 1-dimensional gradients along the width of its lateral parallel channels. The gradient profile can be precisely controlled by the applied flow rate. The proposed design is simple and straightforward, has a small footprint size compared to previously reported devices such as tree-shape designs, and for the first time, provides capability of generating desired 2D and 1D gradients, simultaneously. The finite element simulation analysis proves the feasibility of the microfluidic device, and the fluorescently labelled IgG antibody is used to demonstrate generated chemical gradients. This simple microfluidic device can be implemented for a wide range of high-throughput concentration gradient applications such as chemotaxis, drug screening, and organs-on-chips.

Self-Cleaning Ceramic Tiles Produced via Stable Coating of TiO2 Nanoparticles

The high photocatalytic power of TiO2 nanoparticles has drawn great attention in environmental and medical applications. Coating surfaces with these particles enables us to benefit from self-cleaning properties and decomposition of pollutants. In this paper, two strategies have been introduced to coat ceramic tiles with TiO2 nanoparticles, and the self-cleaning effect of the surfaces on degradation of an organic dye under ultraviolent (UV) exposure is investigated. In the first approach, a simple one-step heat treatment method is introduced for coating, and different parameters of the heat treatment process are examined. In the second method, TiO2 nanoparticles are first aminosilanized using (3-Aminopropyl)triethoxysilane (APTES) treatment followed by their covalently attachment onto CO2 plasma treated ceramic tiles via N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) and N-Hydroxysuccinimide (NHS) chemistry. We monitor TiO2 nanoparticle sizes throughout the coating process using dynamic light scattering (DLS) and characterize developed surfaces using X-ray photoelectron spectroscopy (XPS). Moreover, hydrophilicity of the coated surfaces is quantified using a contact angle measurement. It is shown that applying a one-step heat treatment process with the optimum temperature of 200 °C for 5 h results in successful coating of nanoparticles and rapid degradation of dye in a short time. In the second strategy, the APTES treatment creates a stable covalent coating, while the photocatalytic capability of the particles is preserved. The results show that coated ceramic tiles are capable of fully degrading the added dyes under UV exposure in less than 24 h.

Lubricant-Infused Surfaces with Built-In Functional Biomolecules Exhibit Simultaneous Repellency and Tunable Cell Adhesion

Lubricant-infused omniphobic surfaces have exhibited outstanding effectiveness in inhibiting nonspecific adhesion and attenuating superimposed clot formation compared with other coated surfaces. However, such surfaces blindly thwart adhesion, which is troublesome for applications that rely on targeted adhesion. Here we introduce a new class of lubricant-infused surfaces that offer tunable bioactivity together with omniphobic properties by integrating biofunctional domains into the lubricant-infused layer. These novel surfaces promote targeted binding of desired species while simultaneously preventing nonspecific adhesion. To develop these surfaces, mixed self-assembled monolayers (SAMs) of aminosilanes and fluorosilanes were generated. Aminosilanes were utilized as coupling molecules for immobilizing capture ligands, and nonspecific adhesion of cells and proteins was prevented by infiltrating the fluorosilane molecules with a thin layer of a biocompatible fluorocarbon-based lubricant, thus generating biofunctional lubricant-infused surfaces. This method yields surfaces that (a) exhibit highly tunable binding of anti-CD34 and anti-CD144 antibodies and adhesion of endothelial cells, while repelling nonspecific adhesion of undesirable proteins and cells not only in buffer but also in human plasma or human whole blood, and (b) attenuate blood clot formation. Therefore, this straightforward and simple method creates biofunctional, nonsticky surfaces that can be used to optimize the performance of devices such as biomedical implants, extracorporeal circuits, and biosensors.