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2019

(109) Kseniia Boriachek, Mostafa Kamal Masud, Carlos Palma, Hoang-Phuong Phan, Yusuke Yamauchi, Md. Shahriar A. Hossain, Nam-Trung Nguyen, Carlos Salomon, and Muhammad J. A. Shiddiky (2019)

Avoiding Pre-Isolation Step in Exosome Analysis: Direct Isolation and Sensitive Detection of Exosomes Using Gold-Loaded Nanoporous Ferric Oxide Nanozymes

Analytical Chemistry, 2019. Accepted manuscript; DOI: 10.1021/acs.analchem.8b03619

Publication year: 2019

Abstract

Most of the current strategies for exosome analysis are time-consuming and largely dependent on pre-isolation step using a commercial extraction kit, which requires extensive sample manipulation, costly isolation, reagents, tedious procedures, and sophisticated equipment, and is prone to bias/artefacts. Herein we introduce a simple method for direct isolation and subsequent detection of a specific population of exosomes using an engineered superparamagnetic material with multi-functional properties, namely gold-loaded ferric oxide nanocubes (Au-NPFe2O3NC). In this method, the Au-NPFe2O3NC were initially functionalized with a generic tetraspanin (exosomes associated) antibody (i.e., CD63) and dispersed in sample fluids where they work as ‘dispersible nanocarriers’ to capture bulk population of exosomes. After magnetic collection and purification, Au-NPFe2O3NC-bound exosomes were transferred to the tissue-specific antibody-modified screen-printed electrode. As a proof of principle, we used a specific placental marker, placenta alkaline phosphatase (PLAP), to detect exosomes secreted from placental cells. The peroxidase-like activity of Au-NPFe2O3NC was then used to accomplish an ELISA-based sensing protocol for naked-eye observation along with UV-visible and electrochemical detection of PLAP-specific exosomes present in placental cell-conditioned media. We demonstrated excellent agreement in analytical performance for the detection of placental cell-derived exosomes (i.e., linear dynamic range, 103-107 exosomes /mL; limit of detection, 103 exosomes/mL; relative standard deviation (%RSD) of < 5.5 % for n=3) using with and without the pre-isolation step with commercial “total exosome isolation kit”. We envisage that this highly sensitive, rapid and inexpensive assay could be useful in quantifying specific populations of exosomes for various clinical applications, focusing on pregnancy complications.

(108) Dr. Shunsuke Tanaka, Mostafa Kamal Masud, Dr. Yusuf Valentino Kaneti, Dr. Muhammad J. A. Shiddiky, Prof. Amanullah Fatehmulla, Prof. Abdullah M. Aldhafiri, Prof. W. Aslam Farooq, Prof. Yoshio Bando, Dr. Md. Shahriar A. Hossain, Prof. Yusuke Yamauchi (2019)

Enhanced Peroxidase Mimetic Activity of Porous Iron OxideNanoflakes

ChemNnoMat, 2019. Accepted Manuscript; DOI:https://doi.org/10.1002/cnma.201800487

Publication year: 2019

Abstract
Porous nanomaterials with superior peroxidase mimetic activity (nanozyme) at room temperature have gained increasing attention as potential alternatives to natural peroxidase enzymes. Herein, we report the application of porous iron oxide nanoflakes (IONFs), synthesized using the combination of solvothermal method and high‐temperature calcination as peroxidase nanozyme for the oxidation of 3,3′,5,5′‐tetramethylbenzidine (TMB) in the presence of H2O2. The four IONF catalysts possess porous structures with a wide pore size distribution between 2–30 nm and high specific surface areas around to 200 m2 g−1. The increase of calcination temperature of the IONFs from 250 °C to 400 °C resulted in a gradual decrease in their specific surface area and Michaelis‐Menten constant (Km) for TMB oxidation. The optimum IONF sample showed a much lower Km at 0.24 mM (towards TMB) compared to natural enzyme horseradish peroxidase (HRP) at 0.434 mM, revealing the promising potential of the as‐prepared IONFs as alternatives to HRP for biosensing applications.

(107) Mostafa Kamal Masud Muhammad Umer Md. Shahriar A. Hossain Yusuke Yamauchi Nam-Trung Nguyen Muhammad J.A. Shiddiky (2019)

Nanoarchitecture Frameworks for Electrochemical miRNA Detection

Trends in Biochemical Science, 2019, Accepted Manuscript:
DOI:https://doi.org/10.1016/j.tibs.2018.11.012

Publication year: 2019

Abstract

With revolutionary advances in next-generation sequencing, the human transcriptome has been comprehensively interrogated. These discoveries have highlighted the emerging functional and regulatory roles of a large fraction of RNAs suggesting the potential they might hold as stable and minimally invasive disease biomarkers. Although a plethora of molecular-biology- and biosensor-based RNA-detection strategies have been developed, clinical application of most of these is yet to be realized. Multifunctional nanomaterials coupled with sensitive and robust electrochemical readouts may prove useful in these applications. Here, we summarize the major contributions of engineered nanomaterials-based electrochemical biosensing strategies for the analysis of miRNAs. With special emphasis on nanostructure-based detection, this review also chronicles the needs and challenges of miRNA detection and provides a future perspective on the presented strategies.

(106) Yusuf Kaneti, Ni Luh Wulan Septiani, Indra Saptiama, Xuchuan Jiang, Brian Yuliarto, Muhammad J A Shiddiky, Nobuyoshi Fukumitsu, Yong-Mook Kang, Dmitri GOLBERG and Yusuke Yamauchi (2019)

Self-Sacrificial Templated Synthesis of Three-Dimensional Hierarchical Macroporous Honeycomb-Like ZnO/ZnCo2O4 Hybrid for Carbon Monoxide Sensing

J. Mater. Chem. A, 2019, Accepted Manuscript; DOI:10.1039/C8TA11380G

Publication year: 2019

Abstract
his work reports the fabrication of three-dimensional (3D) zinc oxide/zinc cobaltite (ZnO/ZnCo2O4) hybrid with hierarchical macroporous honeycomb-like structure using highly uniform cobalt glycerate spheres as a self-sacrificial template. In the proposed method, the conversion of the template cobalt glycerate nanospheres into 3D hierarchical macroporous honeycomb-like ZnO/ZnCo2O4 hybrid is achieved via a facile room-temperature reaction with aqueous zinc nitrate solution, followed by calcination in air at 350 °C. The proposed method offers several benefits including: (i) the attainment of ZnO/ZnCo2O4 hybrid in one-step without additional or separate coating step, (ii) the achievement of a unique 3D hierarchical macroporous honeycomb-like structure with interconnecting nanosheets and macropores which are assembled of smaller mesopores, leading to higher surface area and good interparticle separation, (iii) the relatively low calcination temperature required to obtain the ZnO/ZnCo2O4 hybrid (350 °C) and (iv) potential generalization for the creation of other macroporous honeycomb-like cobalt-based oxide nanostructures (including Al-Co and Cu-Co systems). When evaluated as a sensing material for carbon monoxide (CO), the hierarchical honeycomb-like ZnO/ZnCo2O4 hybrid sensor displays a higher sensing response with enhanced selectivity and stability towards CO gas at 300 °C compared to both ZnO hierarchical spheres and ZnCo2O4 nanospheres. The enhanced sensing performance of the hierarchical honeycomb-like ZnO/ZnCo2O4 hybrid is derived from the synergistic cooperation of the formed p-n heterojunction, large surface area and hierarchical macroporous nature of the as-synthesized ZnO/ZnCo2O4 hybrid. It is expected that the proposed general method may open a new path for creating other hierarchical macroporous honeycomb-like oxide nanostructures with enhanced surface areas and improved functional performance.

2018

(105) Abu Ali Ibn Sina, Laura G. Carrascosa, Ziyu Liang, Yadveer S. Grewal, Andri Wardiana, Muhammad J. A. Shiddiky, Robert A. Gardiner, Hemamali Samaratunga, Maher K. Gandhi, Rodney J. Scott, Darren Korbie and Matt Trau (2018)

Epigenetically reprogrammed methylation landscape drives the DNA self-assembly and serves as a universal cancer biomarker

. 2018; 9: 4915.  DOI: 10.1038/s41467-018-07214-w

Publication year: 2018

Abstract

Epigenetic reprogramming in cancer genomes creates a distinct methylation landscape encompassing clustered methylation at regulatory regions separated by large intergenic tracks of hypomethylated regions. This methylation landscape that we referred to as Methylscape is displayed by most cancer types, thus may serve as a universal cancer biomarker. To-date most research has focused on the biological consequences of DNA Methylscape changes whereas its impact on DNA physicochemical properties remains unexplored. Herein, we examine the effect of levels and genomic distribution of methylcytosines on the physicochemical properties of DNA to detect the Methylscape biomarker. We find that DNA polymeric behaviour is strongly affected by differential patterning of methylcytosine, leading to fundamental differences in DNA solvation and DNA-gold affinity between cancerous and normal genomes. We exploit these Methylscape differences to develop simple, highly sensitive and selective electrochemical or colorimetric one-step assays for the detection of cancer. These assays are quick, i.e., analysis time≤10minutes, and require minimal sample preparation and small DNA inpu

(104) Shuaihua Zhang, Qian Yang, Xin Zhou, Zhi Li, Wenjin Wang, Xiaohuan Zang, Chun Wang, Muhammad J. A. Shiddiky, Adharvana Chari Murugulla, Nam-Trung Nguyen, Zhi Wang and Yusuke Yamauchi (2018)

Synthesis of nanoporous poly-melamine-formaldehyde (PMF) based on Schiff base chemistry as a highly efficient adsorbent

Analyst, 2018. Accepted Manuscript. DOI: 10.1039/c8an01623b

Publication year: 2018

Abstract

This study proposes the construction of nanoporous poly-melamine-formaldehyde (PMF) through the Schiff base condensation reaction of paraformaldehyde and melamine. The PMF nanoparticles showed a good adsorption capability to some benzene-ring-containing dyes including acid fuchsine, nigrosine, and methyl orange. Moreover, the as-prepared PMF nanoparticles were employed as the coating adsorbent for the solid-phase microextraction (SPME) of seven volatile fatty acids (VFAs) with high enrichment factors. A PMF-assisted SPME method was established for the enrichment of VFAs from environmental water samples with satisfactory recoveries (88.5%–102.0%) and acceptable precisions (relative standard deviations <10.9%). This contribution might furnish an advanced benchmark for the exploitation of new porous organic polymers as the effective adsorbents for SPME or other fields of utilization.

(103) Ripon Bhattacharjee, Sofia Moriam, Nam-Trung Nguyen and Muhammad J. A. Shiddiky (2018)

A bisulfite treatment and PCR-free global DNA methylation detection method using electrochemical enzymatic signal engagement

Biosensors and Bioelectronics, 2018, Accepted Manuscript. DOI: 10.1016/j.bios.2018.10.020

Publication year: 2018

Abstract

In this paper we report on a bisulfite treatment and PCR amplification-free method for sensitive and selective quantifying of global DNA methylation. Our method utilises a three-step strategy that involves (i) initial isolation and denaturation of global DNA using the standard isolation protocol and direct adsorption onto a bare gold electrode via gold-DNA affinity interaction, (ii) selective interrogation of methylation sites in adsorbed DNA via methylation-specific 5mC antibody, and (iii) subsequent signal enhancement using an electrochemical-enzymatic redox cycling reaction. In the redox cycling reaction, glucose oxidase (GOx) is used as an enzyme label, glucose as a substrate and ruthenium complex as a redox mediator. We initially investigated the enzymatic properties of GOx by varying glucose and ruthenium concentration to delineate the redox cyclic mechanism of our assay. Because of the fast electron transfer by ruthenium (Ru) complex and intrinsic signal amplification from GOx label, this method could detect as low as 5% methylation level in 50 ng of total DNA input. Moreover, the use of methylation-specific 5mC antibody conjugated GOx makes this assay relatively highly selective for DNA methylation analysis. The data obtained from the electrochemical response for different levels of methylation showed excellent interassay reproducibility of RSD (relative standard deviation) <5% for n = 3. We believe that this inexpensive, rapid, and sensitive assay will find high relevance as an alternative method for DNA methylation analysis both in research and clinical platforms.

(102) Ripon Bhattacharjee, Sofia Moriam, Muhammad Umer, Nam-Trung Nguyen and Muhammad J A Shiddiky (2018)

DNA methylation detection: recent developments in bisulfite free electrochemical and optical approaches

Analyst, 2018, Accepted Manuscript (Critical Review)

Publication year: 2018

Abstract

DNA methylation is one of the significant epigenetic modifications involved in mammalian development as well as the initiation and progression of various diseases like cancer. Over the past few decades, enormous research has been carried out for quantification of DNA methylation in the mammalian genome. Earlier, most of these methodologies used bisulfite treatment. However, low conversion, false reading, longer assay time and complex chemical reaction are the common limitations of this method that hinders their application to routine clinical screening. Thus, alternative to bisulfite conversion-based DNA methylation detection, numerous bisulfite-free methods have been proposed. In this regard, electrochemical biosensors have gained much attention in recent years for being highly sensitive yet cost effective, portable, and simple to operate. On the other hand, biosensors with optical readout enable direct real time detection of biological molecules and are easily adaptable to multiplexing. Incorporation of electrochemical and optical readouts to bisulfite free DNA methylation analysis is paving the way for translation of this important biomarker to standard patient care. In this review, we provide a critical overview of recent advances in the development of electrochemical and optical readout based bisulfite free DNA methylation assays.

(101) Ahmed Munaz, Muhammad J. A. Shiddiky, Nam-Trung Nguyen (2018)

Magnetophoretic separation of diamagnetic particles through parallel ferrofluid streams

Sensors and Actuators B: Chemical 275, 459-469.

https://doi.org/10.1016/j.snb.2018.07.176

Publication year: 2018

Abstract

Particle separation based on microfluidic technology offers a simple, reliable, and low-cost approach for the diagnosis of diseases. The separation concept can be extended to genetic engineering, cell transplantation, and immunology. This paper reports a simple microfluidic platform for the separation of diamagnetic particles of different sizes utilizing parallel ferrofluid streams. The ferrofluid streams with predefined concentrations of magnetic nanoparticles promote negative magnetophoresis and are able to separate a particle mixture with a subtle size variation. Numerical simulation was used to optimise the magnetic field gradient, e.g. the number and position of the external permanent magnets. The effect of flow rate ratio and the concentration distribution were analyzed by the simulation and validated by experiments. Furthermore, two-stream and three-stream ferrofluid configurations were evaluated to find the optimum separation performance. The experimental results show a maximum separation efficiency of 78% and 75% with three-stream configuration for 3.2-µm and 4.8-µm particles, respectively.

(100) Ripon Bhattacharjee, Shunsuke Tanaka, Sofia Moriam, Mostafa Kamal Masud, Jianjian Lin, Saad M. Alshehri, Tansir Ahamad, Rahul R Salunkhe, Nam-Trung Nguyen, Yusuke Yamauchi, Md Shahriar Hossain, Muhammad J A Shiddiky (2018)

Porous Nanozymes: Peroxidase-Mimetic Activity of Mesoporous Iron Oxide for Colorimetric and Electrochemical Detection of Global DNA Methylation

Journal of Materials Chemistry B, 2018, 6, 4783 – 4791

Publication year: 2018

Abstract

Nanomaterials (nanozymes) with peroxidase-mimetic activity have been widely used in biosensing platforms as low-cost, relatively stable and prevailing alternatives to natural enzymes. Herein, we report on the synthesis and application of the peroxidase-mimetic activity of mesoporous iron oxide (MIO) for the detection of global DNA methylation in colorectal cancer cell lines. The target DNA was extracted and denatured to get ssDNA followed by direct adsorption onto the surface of a bare screen-printed gold electrode (SPGE). A 5-methylcytosine antibody (5mC) functionalized nanomaterial (MIO-5mC) was then used to recognise the methylcytosine groups present on the SPGE. The MIO-5mC conjugates catalyse the TMB solution in the presence of hydrogen peroxide to give the colorimetric (i.e., naked-eye observation) and electrochemical detection of DNA methylation. The assay could successfully detect as low as 10% difference in the global DNA methylation level in synthetic samples and cell lines with good reproducibility and specificity (%RSD = <5%, for n = 3). This strategy avoids the use of natural enzyme horseradish peroxidase (HRP), traditional PCR based amplification and bisulfite treatment steps that are generally used in many conventional DNA methylation assays. We envisage that our assay could be a low-cost platform with great potential for genome-wide DNA methylation analysis in point-of-care applications.

Graphical abstract: Porous nanozymes: the peroxidase-mimetic activity of mesoporous iron oxide for the colorimetric and electrochemical detection of global DNA methylation

(99) Md. Nazmul Islam, Lena Gorgannezhad, Mostafa Kamal Masud, Shunsuke Tanaka, Md. Shahriar A. Hossain, Yusuke Yamauchi, Nam-Trung Nguyen, Muhammad J. A. Shiddiky (2018)

Novel Graphene Oxide‐loaded Iron Oxide Superparamagnetic Nanoparticles for Ultrasensitive Electrocatalytic Detection of MicroRNA

ChemElectroChem, First published: 12 June 2018

Publication year: 2018

Abstract

We report on the electrocatalytic activity of a new class of superparamagnetic nanoparticle graphene oxide‐ loaded iron oxide (GO/IO hybrid material) towards the reduction of ruthenium hexaammine(III) chloride (Ru(NH3)6]3, RuHex). Leveraging the electrocatalytic activity of GO/IO hybrid material, and signal enhancement capacity of [Ru(NH3)6]3+/[Fe(CN)6]3‐ electrocatalytic cycle, an ultrasensitive and specific electrochemical sensor was developed for the detection of cancer‐related microRNA (miRNA). Using direct affinity interaction between RNA and graphene oxide, magnetically isolated and purified target miRNA were directly adsorbed onto the GO/IO hybrid material‐ modified screen‐printed electrode. The detection was enabled by chronocoulometric (CC) readout of charge compensating [Ru(NH3)6]3+ followed by an enhancement in CC charge display via Ru(NH3)6]3+/[Fe(CN)6]3‐ system. We demonstrated an excellent LOD of 1.0 fM by accurately detecting miR‐21 in synthetic samples, and showcased its clinical utility in ovarian cancer cell lines with high sensitivity (10 cells) and good reproducibility (% RSD = < 5%, for n = 3).

(98) Ahmed Munaz, Muhammad J. A. Shiddiky, Nam-Trung Nguyen (2018)

Recent Advances and Current Challenges in Magnetophoresis Based Micro Magnetofluidics

Biomicrofluidics 12, 031501.

Publication year: 2018

Abstract

The combination of magnetism and microscale fluid flow has opened up a new era for handling and manipulation of samples in microfluidics. In particular, magnetophoresis, the migration of particles in a magnetic field, is extremely attractive for microfluidic handling due to its contactless nature, independence of ionic concentration, and lack of induced heating. The present paper focuses on recent advances and current challenges of magnetophoresis and highlights the key parameters affecting the manipulation of particles by magnetophoresis. The magnetic field is discussed according to their relative motion to the sample as stationary and dynamic fields. The migration of particles is categorized as positive and negative magnetophoresis. The applications of magnetophoresis are discussed according to the basic manipulation tasks such as mixing, separation, and trapping of particles or cells. Finally, the paper highlights the limitations of current approaches and provides the future perspective for this research area.

(97) Lena Gorgannezhad, Muhammad Umer, Mostafa Kamal Masud, Yusuke Yamauchi, Shahriar Al Hossain, Carlos Salomon, Richard Kline, Nam-Trung Nguyen, Muhammad J.A. Shiddiky (2018)

Detection of FGFR2: FAM76A Fusion Gene in Circulating Tumor RNA Based on Catalytic Signal Amplification of Graphene Oxide‐loaded Magnetic Nanoparticles

Electroanalysis 30, Available on line. 

DOI: 10.1002/elan.201800282

 

Publication year: 2018

Abstract

Circulating tumor nucleic acids (ctNAs) are promising biomarkers for minimally invasive cancer assessment. The FGFR2:FAM76A fusion gene is one of the highly promising ovarian cancer biomarkers detectable in ctNAs. Herein, we introduce a new amplification‐free electrochemical assay for the detection of FGFR2:FAM76A fusion gene in ctNAs extracted from ovarian cancer patients. The assay relies on the electrocatalytic activity of a new class of superparamagnetic graphene‐loaded iron oxide nanoparticles (GO‐NPFe2O3). After isolation and purification, the target RNA was directly adsorbed onto the GO‐NPFe2O3 surface through graphene‐RNA affinity interaction. The electrocatalytic signal was achieved by the reduction of surface‐attached ruthenium hexaammine(III) chloride which was further enhanced by using the ferricyanide redox system. Our assay depicted an excellent detection sensitivity down to 1.0 fM, high specificity and excellent reproducibility (% RSD=<5 %, for n=3). The analytical performance of our method was validated with standard qRT‐PCR analysis. We believe that this newly developed assay would be practically applicable in clinical research.

(96) Muhammad Umer, Ramanathan Vaidyanathan, Nam-Trung Nguyen, Muhammad JA Shiddiky (2018)

Circulating Tumor Microemboli: Progress in Molecular Understanding and Enrichment Technologies,

Biotechnology advances 36, 1367-1389.

 

Publication year: 2018

Abstract

Circulating tumor cells (CTCs) and their clusters, also known as circulating tumor microemboli (CTM), have emerged as valuable tool that can provide mechanistic insights into the tumor heterogeneity, clonal evolution, and stochastic events within the metastatic cascade. However, recent investigations have hinted that CTM may not be mere aggregates of tumor cells but cells comprising CTM exhibit distinct phenotypic and molecular characteristics in comparison to single CTCs. Moreover, in many cases CTM demonstrated higher metastatic potential and resistance to apoptosis as compared to their single cell counterparts. Thus, their evaluation and enumeration may provide a new dimension to our understanding of cancer biology and metastatic cancer spread as well as offer novel theranostic biomarkers. Most of the existing technologies for isolation of hematogenous tumor cells largely favor single CTCs, hence there is a need to devise new approaches, or re-configure the existing ones, for specific and efficient CTM isolation. Here we review existing knowledge and insights on CTM biology. Furthermore, a critical commentary on current and emerging trends in CTM enrichment and characterization along with recently developed ex-vivo CTC expansion methodologies is presented with the aim to facilitate researchers to identify further avenues of research and development.

(95) Md Nazmul Islam, Sofia Moriam, Muhammad Umer, Hoang-Phuong Phan, Carlos Salomon, Richard Kline, Nam-Trung Nguyen, Muhammad J. A. Shiddiky (2018)

Naked-eye and Electrochemical Detection of Isothermally Amplified HOTAIR Long non-coding RNA,

Analyst, Just acceptedDOI: 10.1039/C7AN02109G

Publication year: 2018

Abstract

An inexpensive, simple and rapid sensor platform capable of detecting cancer-related long non-coding RNA (lncRNA) with high accuracy is of great interest in the field of molecular diagnostics. Herein, we report on the development of a new colorimetric and electrochemical assay platform for long non-coding HOX transcript antisense intergenic RNA (HOTAIR) detection. Isothermal reverse transcription-recombinase polymerase amplification (RT-RPA) was performed to amplify HOTAIR sequences from a RNA pool extracted from a designated number of ovarian cancer cells and a small cohort of plasma samples derived from patients with ovarian cancer. During RT-RPA, biotinylated dUTPs were randomly incorporated in the amplified product. Subsequently, HOTAIR amplicons were magnetically purified and isolated followed by a horseradish peroxidase (HRP)-catalyzed colorimetric reaction in the presence of 3,3′,5,5′- tetramethylbenzidine (TMB)/ H2O2 system. We finally introduced three potential readout methods for HOTAIR detection – (i) naked-eye visualisation of the color change for a quick screening of the target, (ii) quantitative absorbance measurement by UV-vis, and (iii) amperometric quantification using the electrochemical properties of TMB. The assay has shown excellent reproducibility (% RSD = < 5%, for n = 3) and sensitivity (10 cells/ mL) while detecting HOTAIR in cancer cell lines and patient samples. Expression of HOTAIR in clinical samples was also verified with a standard RT-qPCR method. We believe that our proof of concept assay may find potential relevance for routine clinical screening of cancer- associated lncRNAs.