Global Medical Discovery features paper: Direct detection of endogenous MicroRNAs and their post-transcriptional modifications in cancer serum by capillary electrophoresis-mass spectrometry

Significance Statement

MicroRNAs (miRNAs) are a class of small, single-stranded, non-protein coding RNA molecules that regulate cellular messenger RNA and protein levels by binding to specific messenger RNAs. MicroRNAs play a crucial role in almost every aspect of cell biology, including developmental timing, proliferation, and apoptosis. Moreover, microRNAs  are involved in various cellular activities, such as insulin secretion, immune response, neurotransmitter synthesis, and viral replication. Thus, aberrant microRNA expression has an impact on those critical processes, and consequently, leads to a number of pathological and malignant conditions.

It was observed that selective groups of miRNAs are commonly down-regulated or up-regulated in different types of human cancers and were frequently associated with cytogenetic abnormalities. For example consistent up-regulation of miR-17 and miR-21 was observed in prostate, colon, stomach, lung and pancreatic tumors and miR-155 was identified to be up-regulated in lung, breast and colon cancer. On the contrary, it was reported that miR-29 was down-regulated in chronic lymphocytic leukemia (CLL), acute myeloid leukemia, mantle cell lymphoma, breast, lung and liver cancer. Additionally miR-15a and miR16-1-3p was identified to be down-regulated in CLL, prostate and pituitary adenomas and let-7 family members were found to be down-regulated in breast, lung, colon, ovarian and stomach cancer.

According to the latest miRBase release (www.mirbase.org), more than 30,000 mature microRNA sequences are listed, with ~2500 human microRNAs identified up to date, which can target more than 30% of the human genome. Given the role miRNA plays in human diseases, recent studies have shown that microRNA expression profiles can serve for diagnostic tests on a molecular level for diseases, as well as bases for novel therapeutics. Most importantly, recent identification of circulating microRNAs, have shown great potential of their use as biomarkers since they are readily available in blood samples. MicroRNA analysis, however, presents many challenges due to their low abundance, small size and sequence similarity between miRNA family members. Their small size makes their analysis more difficult compared to messenger RNAs, particularly with conventional molecular biology methods, such as polymerase chain reaction (PCR) and hybridization-based assays. The small size of probes that are used greatly affects the efficiency of these methods because of a very low melting temperature.

Since the development of massively parallel/next-generation sequencing (NGS) of nucleic acids, there has been an increase of microRNA identification and discovery. NGS involves preparation of a complemented DNA library from the RNA sample, followed by the sequencing of millions of individual molecules. Obtained sequence reads undergo a bioinformatical analysis to identify and quantify (relative abundance) both known and novel microRNAs using application tools such as miRDeep. Although NGS is a high-throughput assay for miRNA expression profiling, disadvantages of this technique includes the high cost, the high amount of RNA used, and the sequence-specific biases due to enzymatic steps in complemented DNA library preparation. Nonetheless, NGS has been recently used for differential expression analysis of miRNAs in different diseases, including ovarian cancer and Huntington’s disease, suggesting the usefulness of this technique for diagnostics and early detection.

It has been reported that microRNAs are found in body fluids, such as blood, saliva and urine, of both diseased and healthy people. These extracellular circulating microRNAs exist in a stable form and are resistant to endogenous RNAse activity, as well as extreme pHs and temperatures. The stability of circulating microRNAs are proven to be due to the fact that they are found to be packaged in the micro vesicles, such as micro particles and exosomes, or associated with RNA binding proteins, including Argonaute 2 (AGO2) and nucleophosmin (NPM1). Aberrant expression of circulating miRNAs in different diseases, such as stroke, cardiovascular diseases, breast cancer, ovarian cancer, gastric cancer, lung cancer, colorectal cancer, diabetes, hepatocellular carcinoma and drug induced liver injury has been reported. Blood-based biomarkers are attractive for cancer screening due to their minimal invasiveness, relatively low cost and ease of reproducibility.

MicroRNAs are also modified through a series of processing events after transcription like 5´-end phosphorylation, 3´- end adenylation or uridylation, terminal nucleotide deletion. The problem is that existing bioanalytical methods such as microarrays and a quantitative polymerase chain reaction are sensitive, but not capable of identifying the posttranscriptional modifications of microRNA. Thus, there is a need for a miRNA detection technique, which is direct and multiplexed, requiring minimal sample preparation and can provide qualitative information regarding these modifications.

In this publication, the research laboratory lead by Professor Maxim Berezovski at University of Ottawa reported a multiplexed microRNA detection technique based on capillary electrophoresis – electrospray ionization – mass spectrometry (CE-ESI-MS) that offers a convenient platform for label-free, direct analysis of miRNA from biological samples. CE is highly efficient, and versatile, CE separations are fast, relatively inexpensive, and robust, requiring small amounts of sample and reagents. Coupling CE with MS makes it a powerful method for analysis of biomolecules as it combines high-resolution separations with high detection selectivity and sensitivity. Electrospray ionization is a soft ionization technique used in mass spectrometry for biomolecules. miRNAs can be directly observed without any amplification by mass spectrometry. On-line sample pre-concentration with desalting prior to CE-ESI-MS improves concentration sensitivity for detection of very low amounts of miRNA in complex biological samples without ionization suppression. Due to the size of mature miRNA molecules (21-23 nucleotides), the effect of nucleotide chain fragmentation can potentially be minimized. Therefore, CE-ESI-MS represents a promising method for endogenous miRNA expression and sequence analyses. The results for the CE-MS study were validated by conventional SYBR green-based quantitative reverse transcription PCR.

Using the CE-MS method, the researchers detected two endogenous human circulating microRNAs, a 23-nucleotide long 5´-phosporylated microRNA with 3´-uridylation (iso-miR-16-5p) and a 22-nucleotide long 5´-phosporylated microRNA (miR-21-5p) isolated from B-cell chronic lymphocytic leukemia serum. The CE separation and following MS analysis provides label-free quantitation and reveals modifications of microRNAs.  MicroRNA profiling of serum samples with CE-MS has the potential to be a versatile and minimally invasive bioassay that could lead to better clinical diagnostics and disease treatment.

 

About The Author

Dr. Nasrin Khan is an NSERC Visiting Research Fellow in the department of Environmental Health Science and Research Bureau, Health Canada. Her current research is focused on the study of gene expression and epigenetics (DNA methylation, histone modifications, and microRNA expression) in mammals upon exposure to environmental chemicals. 

About The Author

Dr. Gleb Mironov is a mass spectrometrist at the University of Ottawa. His major research interests lie in the area of mass spectrometry assisted biomarker discovery and characterization.  

About The Author

Dr. Maxim V. Berezovski is an Associate Professor of Chemistry at the University of Ottawa in Canada and a Director of Imaging and Proteomics core facilities. He won a 2015 University of Ottawa’s Young Researcher Award and was the recipient of an Early Research Award from Ministry of Research and Innovation in 2012. His research focuses on the study of affinity interactions and conformational dynamics of biomolecules by kinetic capillary electrophoresis and mass spectrometry and on the development of DNA aptamers for sensing pathogens and cancer cells.  

Journal Reference

Anal Bioanal Chem. 2016 Apr;408(11):2891-9. 

Khan N¹, Mironov G¹, Berezovski MV².

Show Affiliations

1. Department of Chemistry and Biomolecular Sciences, University of Ottawa, 10 Marie Curie, Ottawa, Ontario, K1N 6 N5, Canada.
2. Department of Chemistry and Biomolecular Sciences, University of Ottawa, 10 Marie Curie, Ottawa, Ontario, K1N 6 N5, Canada. Maxim.Berezovski@uOttawa.ca.

 

Abstract

MicroRNA molecules (miRNAs) are a class of small, single-stranded, non-coding RNA molecules that regulate cellular messenger RNA and their corresponding proteins. Extracellular miRNAs circulate in the bloodstream inside exosomes or in complexes with proteins and lipoproteins. The miRNA sequences and their quantitative levels are used as unique signatures associated with cancer diagnosis and prognosis after anticancer treatment. MicroRNAs are modified through a series of processing events after transcription like 5′-end phosphorylation, 3′- end adenylation or uridylation, terminal nucleotide deletion. The problem is that existing bioanalytical methods such as microarrays and a quantitative polymerase chain reaction are sensitive, but not capable of identifying the post-transcriptional modifications of miRNA. Here we report a capillary electrophoresis-mass spectrometry (CE-MS) method, which performs a multiplex, direct analysis of miRNAs from biological samples. Using the CE-MS method, we detected two endogenous human circulating miRNAs, a 23-nucleotide long 5′-phosporylated miRNA with 3′-uridylation (iso-miR-16-5p), and a 22-nucleotide long 5′-phosporylated miRNA (miR-21-5p) isolated from B-cell chronic lymphocytic leukemia serum. The CE separation and following MS analysis provides label-free quantitation and reveals modifications of miRNAs. MicroRNA profiling of serum samples with CE-MS has the potential to be a versatile and minimally invasive bioassay that could lead to better clinical diagnostics and disease treatment.

Go To Anal Bioanal Chem

 

Global Medical Discovery features paper: Biomarkers of liver fibrosis detecting with electrochemical immunosensor on clinical serum

Significance Statement

Chronic hepatic disease causes high morbidity and mortality worldwide, which can lead to liver fibrosis and the subsequent development of cirrhosis and even hepatocellular carcinoma. For chronic liver injuries of many etiologies, including viral hepatitis, alcohol abuse, metabolic diseases, autoimmune diseases, and cholestatic liver diseases, they could produce fibrosis as a result of deregulation of the normal healing process with massive accumulation of extracellular matrix (ECM). Therefore, the early diagnosis of liver fibrosis is vital for therapeutic decisions and prognostic evaluations. Among the various diagnostic approaches, needle biopsy is considered as the ‘gold standard’. However, it was invasive, confounded by high sampling heterogeneity and carried a finite risk of complications. Especially, it is not suit for frequent evaluations of this chronic disease.

In the research, an electrochemical immunosensor was established to detect representative biomarkers of liver fibrosis, such as hyaluronate acid (HA) and transforming growth factor beta 1 (TGF1). Through a self-assembled monolayer of polyethylene glycol (PEG), anti-bodies against HA and TGF1 were successfully immobilized on interdigitated electrodes. It produced a robust and sensitive membrane by improving the uniformity, density, and distribution of the antibodies for the biomarkers. Based on impedance sensing, HA and TGF1 were sensitively detected in the ranges of 1-1000 ng/ml. The detection limits of HA and TGF1 reached 0.586 ng/ml and 0.570 ng/ml, respectively. In addition, for the detection of clinical serum samples, the results were in excellent agreement with the tests of HA, type III pre-collagen (PCIII), IV collagen (IV-C), and laminin (LN) that conducted by radio immunoassay for liver fibrosis. It indicated that the approach provided a valuable, universal, and label-free strategy in evaluating liver fibrosis and other chronic diseases for point-of-care diagnostics. Compared to the common immunoassays, electrochemical sensing had attracted more interests due to their inherent advantages, including high sensitivity, time-saving, and eases of operating. It exhibited great potential in point-of-care diagnostics for early detection of various diseases. Through sensitively detecting the serum biomarkers, the electrochemical immunosensor may be useful in detecting the presence of or severe, and probably moderate, fibrosis. The precise diagnostic for a disease is essential for successful treatment and recovery of patients. Therefore, the combination of different biomarkers, serum levels of HA, TGF-β1, would aid clinicians in diagnosing fibrosis during the early stages, eliminating the need for liver biopsy and allowing early treatment, thereby preventing fibrosis progression.

Figure Legend: A label-free immunosensor in detecting representative biomarkers of liver fibrosis, hyaluronate acid (HA) and transforming growth factor beta 1 (TGF1) through electrochemical impedance.

About The Author

Qingjun Liu received his Ph.D. degree in biomedical engineering from Zhejiang University, PR China in 2006. He is currently a professor in Biosensor National Special Lab, Zhejiang University. He is also a visiting scholar in the Micro and Nanotechnology Laboratory (MNTL) at the University of Illinois at Urbana-Champaign (UIUC). He received Nomination Award of the Excellent PhD Dissertation of China, in 2008. He published the book of Cell-Based Biosensors: Principles and Applications, by Artech House Publishers USA in October 2009. And, the book of Biomedical Sensors and Measurement published by Zhejiang University Press and Springer-Verlag GmbH Berlin Heidelberg, 2011. His research interests concentrate on the biosensors (e.g. living cell sensor, DNA sensor and protein sensor) and BioMEMS system. 

Journal Reference

Sensors and Actuators B: Chemical, Volume 222, 2016, Pages 127–132.

Yao Yao¹, Jianfeng Bao², Yanli Lu¹,Diming Zhang¹,Senbiao Luo³,Xing Cheng¹,Qian Zhang¹, Shuang i¹,Qingjun Liu¹

Show Affiliations

1. Biosensor National Special Laboratory, Key Laboratory for Biomedical Engineering of Education Ministry, Department of Biomedical Engineering, Zhejiang University, Hangzhou 310027, PR China
2. Xixi Hospital of Hangzhou, Hangzhou 310023, PR China
3. Shangyu People’s Hospital of Zhejiang Province, Shangyu 312000, PR China

Abstract

Diagnosing hepatic fibrosis at an early stage with sensitive and specific monitoring approach is crucial for patient therapeutics and survival. In this study, an electrochemical immunosensor was established to detect representative biomarkers of liver fibrosis, such as hyaluronate acid (HA) and transforming growth factor beta 1 (TGFβ1). Through a self-assembled monolayer of polyethylene glycol (PEG), antibodies against HA and TGFβ1 were successfully immobilized on interdigitated electrodes. It produced a robust and sensitive membrane by improving the uniformity, density, and distribution of the antibodies for the biomarkers. Based on impedance sensing, HA and TGFβ1 were sensitively detected in the ranges of 1–1000 ng/ml. The detection limits of HA and TGFβ1 reached 0.586 ng/ml and 0.570 ng/ml, respectively. In addition, for the detection of clinical serum samples, the results were in excellent agreement with the tests of HA, type III pre-collagen (PCIII), IV collagen (IV-C), and laminin (LN) that conducted by radioimmunoassay for liver fibrosis. The research indicated that the approach provided a valuable, universal, and label-free strategy in evaluating liver fibrosis and other chronic diseases for point-of-care diagnostics.

Go To Sensors and Actuators B: Chemical

 

Global Medical Discovery features paper: Synergism of Water Shock and a Biocompatible Block Copolymer Potentiates the Antibacterial Activity of Graphene Oxide

Significance Statement

There is an urgent need to find alternative routes to control the spreading of antibiotics-resistant bacteria. Graphene and its water-soluble form graphene oxide (GO) were previously shown to be promising antibacterial agents. However, their antibacterial activity was relatively low and they show toxicity to human cells at high dose of usage. This study demonstrated a new strategy for significantly enhancing the antibacterial activity of graphene oxide. It showed that graphene oxide can kill 99% of bacteria when mixed with a human-friendly detergent in salt-reduced water. Further, the mixture of graphene oxide and Pluronic displays over 50% lower toxicity to human skin cells than the effect of graphene oxide alone. The developed strategy relies on the combination of two effects. The first effect is coming from the water itself. Bacteria inevitably undergo swelling when faced with salt-reduced water which puts a stress on bacterial envelope due to occurrence of microscopic damages. The second effect comes from pluronic; a bio-friendly polymer widely used in drug formulations and food applications. Pluronic increases the stability of graphene oxide in solution and also helps graphene oxide to interact with bacteria better by surrounding the bacterial cells more effectively. Water and detergents are undoubtedly the most common cleaning agents. This study may open the door for graphene oxide to be a widespread cleaning agent for fighting bacteria.

 

About The Author

Yuan Chen received his BEng in Chemical Engineering and MEng in Biochemical Engineering from Tsinghua University. He obtained his PhD in Chemical Engineering at Yale University in 2005. He is currently a Professor at School of Chemical and Biomolecular Engineering (CBE), The University of Sydney. He joined the School of Chemical and Biomedical Engineering in Nanyang Technological University (NTU) in Singapore as an Assistant Professor in 2005. He was promoted to a tenured Associate Professor in 2010. He was visiting Associate Professor at Brown University in 2010 and visiting Chair Professor at Tianjin University of Technology in China 2011-2015. He served as Head of CBE Division in NTU from July 2011 to June 2014. He received an Excellence in Review Award from CARBON in 2015, a Young Scientist Award from the Singapore National Academy of Science in 2011, a Tan Chin Tuan Exchange Fellowship in Engineering in 2010, and JSPS exchange award in 2009. He is currently associate editor for Carbon (Elsevier), editorial board member for Nanomaterials, and Heliyon (Elsevier).  His research focuses on developing scalable chemical processes to synthesize carbon nanomaterials with well-defined nanoscale structures, assembling nanoscale carbon nanomaterials into functional macroscale systems, and utilizing these novel materials for sustainable energy, environmental and biomedical applications.

About The Author

Enis Karahan earned BSc double-degree in Chemical Engineering and Molecular Biology & Genetics from İstanbul Technical University. He received his MSc in Materials Science and Engineering at Koç University. Holding Singapore International Graduate Award (SINGA), he currently pursues his PhD studies in Bioengineering at Nanyang Technological University and Singapore Institute of Manufacturing Technologies of A*STAR. In addition to his academic experience, he also has an industrial background majorly gained in an R&D department of a textile company focused on polymer technologies. His expertise covers a broad spectrum of materials science including self-assembly of multilayered surface coatings, noncovalent functionalization of nanoparticles, and antibacterial, biomedical, and environmental applications of carbon-based nanocomposites to name a few. To date, he has over ten publications in peer-reviewed journals on his account. 

 

Journal Reference

Small. 2016 Feb;12(7):951-62.

Karahan HE^1,2, Wei L¹, Goh K¹, Wiraja C¹, Liu Z¹, Xu C^1,3, Jiang R¹, Wei J², Chen Y^1,4.

Show Affiliations

1. School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore, 637459, Singapore.
2. Singapore Institute of Manufacturing Technology (SIMTech), Singapore, 638075, Singapore.
3. NTU-Northwestern Institute of Nanomedicine, Nanyang Technological University, 50 Nanyang Avenue, Singapore, 639798, Singapore.
4. School of Chemical and Biomolecular Engineering, The University of Sydney, Sydney, 2006, Australia.

 

Abstract

Graphene oxide (GO) is promising in the fight against pathogenic bacteria. However, the antibacterial activity of pristine GO is relatively low and concern over human cytotoxicity further limits its potential. This study demonstrates a general approach to address both issues. The developed approach synergistically combines the water shock treatment (i.e., a sudden decrease in environmental salinity) and the use of a biocompatible block copolymer (Pluronic F-127) as a synergist co-agent. Hypoosmotic stress induced by water shock makes gram-negative pathogens more susceptible to GO. Pluronic forms highly stable nanoassemblies with GO (Pluronic-GO) that can populate around bacterial envelopes favoring the interactions between GO and bacteria. The antibacterial activity of GO at a low concentration (50 μg mL(-1) ) increases from <30% to virtually complete killing (>99%) when complemented with water shock and Pluronic (5 mg mL(-1) ) at ≈2-2.5 h of exposure. Results suggest that the enhanced dispersion of GO and the osmotic pressure generated on bacterial envelopes by polymers together potentiate GO. Pluronic also significantly suppresses the toxicity of GO toward human fibroblast cells. Fundamentally, the results highlight the crucial role of physicochemical milieu in the antibacterial activity of GO. The demonstrated strategy has potentials for daily-life bacterial disinfection applications, as hypotonic Pluronic-GO mixture is both safe and effective.

© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Go To Small.

 

Global Medical Discovery features paper: Label-free mapping of single bacterial cells using surface-enhanced Raman spectroscopy

Significance Statement

In this paper we presented a simple, rapid and label-free surface-enhanced Raman spectroscopy (SERS) based mapping method for the detection and discrimination of Salmonella enterica and Escherichia coli on silver dendrites. The results show the developed mapping method is able to detect single bacterial cells adsorbed on the surface of silver dendrites with a limit of detection as low as 10⁴ CFU/mL, which is two orders of magnitude lower than that of the traditional SERS method under the same experimental condition. The time needed for collecting a 225 points map was merely 24 minutes. Moreover, the developed SERS mapping method can realize simultaneous detection and identification of Salmonella enterica subsp enterica BAA1045 and Escherichia coli BL21 from a mixture sample. The surface-enhanced Raman spectroscopy spectrum of an unknown bacterial cell can be used as fingerprints to identify this cell using a prediction model based on principle component analysis results of the standard spectra. The plot of the detected bacterial signal number versus the log value of the bacterial concentration demonstrates the potential quantitative capacity of the developed method at certain range. To the best of our knowledge, this is the first study that a label-free surface-enhanced Raman spectroscopy mapping method has been developed for simultaneous detection and identification of a bacterial mixture. Our results demonstrate the great potential of the label-free surface-enhanced Raman spectroscopy mapping method to detect, identify and quantify bacteria and bacterial mixtures simultaneously.

 

About The Author

Panxue Wang is a visiting PhD student in Dr. Lili He’s Lab (University of Massachusetts, Amherst). Panxue Wang obtained her bachelor’s degree of Food Science and Engineering in 2012 from Northwest A&F University, and she became a master combined with PhD student (Major: Food science; Advisor: Mingtao Fan) of the Northwest A&F University since 2012. She got the scholarship from the China Scholarship Council (CSC) to support her study in the United States and started to work with Dr. Lili He from September 2014. She will finish this program in August 2016. During this time, she focused on developing rapid bacteria detection, identification and characterization methods using an advanced analytical technique, surface enhanced Raman spectroscopy (SERS). Her research interests comprise many aspects of bacteria, such as rapid detection and identification of food pathogens using SERS mapping, interaction of nanoparticles with bacteria cells, malolactic fermentation process in wine production, and antibiotic resistance in lactic acid bacteria. 

 

Journal Reference

Analyst. 2016 Feb 8;141(4):1356-62. 

Wang P¹, Pang S², Chen J², McLandsborough L², Nugen SR², Fan M³, He L².

Show Affiliations

1. Department of Food Science, University of Massachusetts, Amherst, MA 01003, USA. lilihe@foodsci.umass.edu and College of Food Science and Engineering, Northwest A&F University, Yangling 712100, Shaanxi, P. R. China.
2. Department of Food Science, University of Massachusetts, Amherst, MA 01003, USA. lilihe@foodsci.umass.edu.
3. College of Food Science and Engineering, Northwest A&F University, Yangling 712100, Shaanxi, P. R. China.

Abstract

Here we presented a simple, rapid and label-free surface-enhanced Raman spectroscopy (SERS) based mapping method for the detection and discrimination of Salmonella enterica and Escherichia coli on silver dendrites. The sample preparation was first optimized to maximize sensitivity. The mapping method was then used to scan through the bacterial cells adsorbed on the surface of silver dendrites. The intrinsic and distinct SERS signals of bacterial cells were used as the basis for label-free detection and discrimination. The results show the developed method is able to detectsingle bacterial cells adsorbed on the silver dendrites with a limit of detection as low as 10(4) CFU mL(-1), which is two orders of magnitude lower than the traditional SERS method under the same experimental condition. The time needed for collecting a 225 points map was approximately 24 minutes. Moreover, the developed SERS mapping method can realize simultaneous detection and identification of Salmonella enterica subsp. enterica BAA1045 and Escherichia coli BL21 from a mixture sample using principle component analysis. Our results demonstrate the great potential of the label-free SERS mapping method to detect, identify and quantify bacteria and bacterial mixtures simultaneously.

Go To Analyst

 

Global Medical Discovery features paper: A Smart Antibacterial Surface for the On-Demand Killing and Releasing of Bacteria

Significance Statement

A novel smart antibacterial surface with on-demand switchable functionalities was developed by exploring the synergistic effects of combining stimuli-responsive polymers and nanomaterials with unique topographies. This surface is based on silicon nanowire arrays modified with a pH-responsive polymer, poly(methacrylic acid) (SiNWAs-PMAA). The SiNWAs-PMAA surface could be regarded as an effective dynamic antibacterial reservoir that not only exhibited a remarkably high capacity for binding lysozyme at an acidic pH (pH 4) but also could release a majority of the adsorbed lysozyme when the pH was increased to a neutral value (pH 7). The released lysozyme molecules maintained their enzymatic activity and thus served as biocides to kill bacteria both suspended in solution and attached to the surface. More importantly, after the killing process, the dead bacteria and debris attached to the SiNWAs-PMAA surface could be readily removed by further increasing the pH to a basic value (pH 10); the “cleaned” surface could then be used to load new lysozyme for repeated applications. Specifically, the functionality of the surface (loading biocides, killing bacteria, releasing bacteria) could be simply switched via step-wise modification of the environmental pH and could be effectively maintained after several kill-release cycles.

Considering that bacterial contamination is one of the main underlying causes of tissue infections and inflammation as well as device failure and endowing surfaces with antibacterial properties have attracted great research interests, it is believed that this work contributes remarkably to this subject area by providing a new strategy to engineer multifunctional surfaces with advanced antibacterial capability of both maintaining long-term antibacterial effects and keeping the surfaces free of the accumulation of dead bacteria and debris. Moreover, this approach constitutes a design platform, not limited to the silicon nanowire arrays and poly(methacrylic acid) pair, but applicable to other combinations of porous nanomaterials and stimuli-responsive polymers.

 

About The Author

Qian Yu is an associate professor at the College of Chemistry, Chemical Engineering and Materials Science at Soochow University, Suzhou, China. He received his PhD degree in Materials Science from Wuhan University of Technology in 2011, and then worked in the Department of Biomedical Engineering in Duke University, as a postdoctoral associate (2011-2014). In 2014, he joined Soochow University and was appointed as an associate professor. His research interests include development of stimuli-responsive polymers for applications in biomedical and biotechnology fields, nanostructural biointerfaces, and anti-biofouling surfaces. He has published published more than 40 research articles in various peer reviewed journals with a total citiaton of more than 750 times (please refer to google scholar: http://ift.tt/1Z7JquS).  

About The Author

Hong Chen is a professor at the College of Chemistry, Chemical Engineering and Materials Science at Soochow University, Suzhou, China. She earned her Ph.D. degree from Nanjing University in 2001 and worked as a postdoctral fellow at McMaster University (2001-2004). After returning back to China, she held a full professor position at Wuhan University of Technology from 2004 to 2009. In 2010, her research group moved to Soochow University, where she established Macromolecules and Biointerface Laboratory (MacBio) of Soochow University. She has been the PI of more than 10 national research projects funded by the Ministry of Science and Technology of China, the Ministry of Education of China and the National Natural Science Foundation of China, including a Major International Joint Research Project. She is the winner of National Science Fund for Distinguished Young Scholars (2011). She has been admitted as a Fellow of the Royal Society of Chemistry (FRSC) in 2014. She was a guest editor for the special issue: “Biointerfaces in China” published in 2011 and became an editor of Colloids and Surface B: Biointerfaces since 2013, after three years she became the associate editor of Polymer Chemistry. Her research interests include: surface modification and functionalization of biomaterials, interaction of protein/cell and biomaterials, hemocompatibility of biomaterials, and biological detection. She has published more than 100 research articles in various peer reviewed journals. For more detailed information, please refer to her website: macbio.suda.edu.cn. 

Journal Reference

Adv Healthc Mater. 2016 Feb;5(4):449-56.

Wei T¹, Yu Q¹, Zhan W¹, Chen H¹.

College of Chemistry, Chemical Engineering and Materials Science, Soochow University, Suzhou, Jiangsu, 215123, P. R. China.

Abstract

For various human healthcare and industrial applications, endowing surfaces with the capability to not only efficiently kill bacteria but also release dead bacteria in a rapid and repeatable fashion is a promising but challenging effort. In this work, the synergistic effects of combining stimuli-responsive polymers and nanomaterials with unique topographies to achieve smart antibacterial surfaces with on-demand switchable functionalities are explored. Silicon nanowire arrays are modified with a pH-responsive polymer, poly(methacrylic acid), which serves as both a dynamic reservoir for the controllable loading and release of a natural antimicrobial lysozyme and a self-cleaning platform for the release of dead bacteria and the reloading of new lysozyme for repeatable applications. The functionality of the surface can be simply switched via step-wise modification of the environmental pH and can be effectively maintained after several kill-release cycles. These results offer a new methodology for the engineering of surfaces with switchable functionalities for a variety of practical applications in the biomedical and biotechnology fields.

© 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Go To Adv Healthc Mater

 

Global Medical Discovery features paper: Preparation of photoluminescent enzymatic nanosensors for glucose sensing

Significance Statement

Diabetes mellitus is a worldwide problem because many people are diseased. Its main characteristic the glucose level, is chronically raised. Rigorous controlling of glucose level can decelerate long-term complications such as microangiopathy, kidney or nerve damages which are attributed to diabetes. Hence, numerous sensors were developed for fast monitoring of glucose levels in physiological fluids.

In the present work, a simple and steady fluorescent nanoparticle NPs-GOx was prepared by coupling GOx to nanoparticle doped with two fluorescence dyes for detecting glucose. The ensembles comprises oxygen sensitived ratiometric fluorescence nanoparticles (NPs) and glucose oxidase (GOx), which can be used as glucose biosensor for its quantitative analysis. The large surface area of these nanoparticles resulted in a high enzyme loading. Specifically, NPs are prepared by a one-step reprecipitation-encapsulation method for sensing dissolved oxygen. To endow the NPs with glucose-detecting capability, GOx was successfully immobilized onto the surface of PLL shell via a glutaraldehyde-mediated Schiff-base reaction. Thus, the nanoprobe NPs-GOx for glucose detecting is successfully constructed. The nanoprobe possess excellent fluorescence emission (at 520nm and 650nm) and high stability. Glucose calibration was performed with ratiometric photoluminescence and time-resolved fluorescence (TRF) respectively, and a series of calibration plots were constructed according to determination time. In comparison, the ratiometric method resulted in wide dynamic range (e.g. 2–10 mM) and high limit of detection (∼1–2 mM), while the TRF mode gave narrow dynamic range (e.g. 1–6 mM) with low detection limit (∼0.1–0.2 mM). Finally the enzymatic glucose nanosensors were tested in human serum samples with a TRF microplate reader.

About The Author

Prof. Hong-shang Peng received his PhD degree in Optics from Beijing Jiaotong University in 2007. After postdoctoral research in the group of Prof. Otto S. Wolfbeis at University of Regensburg (Germany) as an Alexander von Humboldt fellow, he joined the faculty of Beijing Jiaotong University in 2009. From 2013 to 2014, he visited the group of Prof. Daniel T. Chiu at the University of Washington (Seattle). In Nov. 2015, he transferred to Minzu University of China as a Professor. His research interest focused on fluorescent nanoparticles as biosensors and bio-imaging agents. 

About The Author

Shao-wei Gao is a graduate student of Beijing Jiaotong University in China. She obtains her bachelor’s degree from Beijing Jiaotong University. Her research focuses on fluorescent glucose and hydrogen peroxide nanosensors. 

Journal Reference

Sensors and Actuators B: Chemical, Volume 222, January 2016, Pages 638–644. 

Shao-wei Gao¹,Hong-shang Peng¹, Xiao-hui Wang¹,Fang-tian You¹,Feng Teng¹,Hong-xia Wang²

Show Affiliations

1. Key Laboratory of Luminescence and Optical Information, Ministry of Education, Institute of Optoelectronic Technology, Beijing Jiaotong University, Beijing 100044, PR China
2. Department of Neurology, Zhongguancun Hospital, Beijing 100190, PR China

Abstract

A novel photoluminescent glucose nanosensor was facilely prepared by coupling glucose oxidase (GOx) with poly-l-lysine coated oxygen nanosensors via a glutaraldehyde-mediated Schiff-base reaction. The GOx molecules residing on particle surface catalyzed glucose with the expense of oxygen, which was detected by the sensing particle core incorporated with the reference dye coumarin 6 and oxygen probe Pt(II)-meso-tetra(pentafluorophenyl)porphine. The proposed glucose nanosensors (∼150 nm in hydrodynamic diameter) had a quick response time varied from less than 2 min to 4 min. Glucose calibration was performed with ratiometric photoluminescence and time-resolved fluorescence (TRF) respectively, and a series of calibration plots were constructed according to determination time. In comparison, the ratiometric method resulted in wide dynamic range (e.g. 2–10 mM) and high limit of detection (∼1–2 mM), while the TRF mode gave narrow dynamic range (e.g. 1–6 mM) with low detection limit (∼0.1–0.2 mM). Finally the enzymatic glucose nanosensors were tested in human serum samples with a TRF microplate reader.

Go To Sensors and Actuators B: Chemical

 

Global Medical Discovery features paper: Development of localized surface plasmon resonance biosensors for the detection of Brettanomyces bruxellensis in wine

About The Author

Marisa Manzano was born in Udine, Italy. She got the degree in Natural Sciences at University of Padua. She was researcher in Food Microbiology at the University of Udine at the Department of Food Science from 1990 till 2005. Since 2005 Associate Professor at University of Udine teaching: Molecular Biology Techniques, Biotechnology of Microorganisms, Genetic of Microorganisms, Selection and Use of Enological Yeasts in the degree course of Food Science and Technology and Viticulture and Enology. She is Member of Committees: Academic Collegiate of Doctorate Research, Didactic Committee, State Exam Commission; member of the Committee “Quality System in the  laboratory of Microbiology” (UNICHIM). Co-author of a patent – detection of Listeria monocytogenes by PCR, 1996, C12Q. She has long experience in Microbiology and Molecular Biology. Now working on the development of DNA probes for the construction of biosensors for the detection of pathogens. She was invited speaker in various international conferences with talks about rapid detection of pathogens using molecular methods or biosensors. Author of  271 publications including papers/book chapter/abstracts and posters.

About The Author

Priya Vizzini was born in India in 1985. She obtained bachelor’s degree in Biomedical Laboratory Technician at the University of Udine (Italy) in 2011. She won an Erasmus grant to spend 5 months at Troyes University of technology (France) working  on biological applications of  QCM and Localized Surface Plasmon Resonance biosensor under the supervision of the Prof. Rodica Elena Ionescu. She got the master’s degree in Plant and Animal Biotechnology in 2014 at the University of Udine (Italy). In december 2015 she was a selected speaker at the “2^th PARMA NANO-DAY”scientific workshop for the session agribusiness, ambient and biotechnology at Parma University. At present she collaborates in the Department of Agricultural, Food, Environmental and Animal science  in University of Udine. She works on the development of molecular biology methods (PCR, Nested-PCR, DGGE/ TTGE and blotting techniques) for the detection of pathogens in food-beverage and on the evaluation of sterilization systems for private companies. 

About The Author

Kun Jia received his B.S. and M.S. degree in Chemistry from University of Electronic Science and Technology of China (UESTC) in 2007 and 2010, respectively. He obtained his Ph.D. degree from Laboratory of Nanotechnology and Instrumentation Optics (LNIO) at University of Technology of Troyes (UTT) in France under the supervision of Prof. Elena Rodica IONESCU at the end of 2013. In March 2014, Dr. JIA joined the School of Microelectronics and Solid State Electronics of UESTC as an associate professor in Chemistry. His current research interests lie in the field of plasmonic controlled polymer fluorescence, optical (bio)chemical sensors on flexible substrate, and synthesizing of advanced optofunctional polymeric nanocomposites.    

About The Author

Pierre-Michel Adam has obtained his PhD in Physics in 1995 (Université de Bourgogne), and is at present Full Professor at the Université de Technologie de Troyes (ICD-LNIO). Title of his PhD is “Photon Scanning Tunneling Microscope (PSTM) with a polychromatic incoherent light source. Near-field investigation of test samples and surface plasmons.” In january 1995, P.M. Adam has joined the Universite de Technologie de Troyes as an Assistant Professor and has been appointed Full professor in February 2003. His fields of research are near-field microscopy and spectroscopy, surface plasmons, surface enhanced Raman scattering. He is currently in charge of a research group “Nanospectroscopy” at the ICD-LNIO. He is vice-president of a European network COST action MP1302 “Nanospectroscopy”. He is author/coauthor of 76 publications in international journals. He is/has been the supervisor of 16 PhD thesis. 

About The Author

Rodica Elena Ionescu earned one Ph.D. degree from the Ben-Gurion University of Negev, Israel in Biotechnological Engineering in 2004 and a second Ph.D. degree in Chemistry, from the University of Bucharest, in 2007. She has performed three post-doctoral positions in France in the field of electrochemical biosensors. Between February-October 2008, Dr. Ionescu was a researcher fellow at the National Nanotechnology Laboratory, Lecce (Italy) working on impedimetric cell-chips based interdigitated microlectrodes. In November 1^st 2008, Dr. Ionescu joined the Université de Technologie de Troyes (UTT) as an Assistant Professor, becoming a member of the Laboratoire de Nanotechnologie et d’Instrumentation Optique (LNIO). In December 4^th 2009, Dr. Ionescu obtained the Habilitation Title for conducting independent research (HDR) equivalent to an Associate Professor position. Thanks to the obtained research grants, Dr. Ionescu developed novel acoustic and optical platforms for ultrasensitive detection of biomolecules and pesticides. In 2012, Dr. Ionescu was awarded with a national OSEO Innovation grant. Between April 1^st 2014 to March 30, 2015 Dr. Ionescu was the research manager of a Proof-of Concept (POC) Grant-NRF-POC 002-026 supported by the Singapore National Research Foundation concerning the Electrochemical lateral flow biosensor detection and quantification of Dengue virus in whole blood at the Nanyang Technological University, School of Materials Science and Engineering under an international CREATE-NTU-HUJ-BGU program. Her current research activities include the development of multi-analyte biosensing platforms, specific (bio)functionalization of surfaces, atomic force microscopy nanopipette applications, controllable synthesis of nanoparticle, evaluation of nanoparticles and water pollutants toxicity to living cells and microorganisms. She has published over 40 pre-reviewed articles, 4 book chapters and 7 patents with 9 extensions. 

Journal Reference

Sensors and Actuators B: Chemical, Volume 223, February 2016, Pages 295–300 .

Marisa Manzano¹, Priya Vizzini¹,Kun Jia^2,3,Pierre-Michel Adam²,Rodica Elena Ionescu² 

Show Affiliations

1. Department of Food Science, University of Udine, via Sondrio 2/A, 33100 Udine, Italy
2. Laboratoire de Nanotechnologie et d’Instrumentation Optique, Institute Charles Delaunay, Universite’ de Technologie de Troyes, UMR CNRS 6281, 12 Rue Marie-Curie, CS 42060, 10004 Troyes Cedex, France
3. School of Microelectronics and Solid-State Electronics, University of Electronic Science and Technology of China, 610054 Chengdu, China

Abstract

Incident light interacting with noble-metal nanoparticles with smaller sizes than the wavelength of the incident light induces localized surface plasmon resonance (LSPR). In this work a gold nanostructured surface was used for the immobilization of a 5′ end Thiol modified DNA probe to develop a LSPR nanobiosensor for the detection of the spoiler wine yeast Brettanomyces bruxellensis. Gold was evaporated to obtain a gold thickness of 4 nm. DNA (2 μL) from the target microorganism and the negative control at various concentrations were used to test the specificity and sensitivity of the LSPR technique. Changes in the optical properties of the nanoparticles due to DNA-probe binding are reflected in the shift of LSPR extinction maximum (λ_max). The results obtained using as target microorganism B. bruxellensis, and as negative control Saccharomyces cerevisiae demonstrated the specificity of both the DNA-probe and the protocol. The LSPR spectrophotometry technique detects 0.1 ng/μL DNA target confirming the possibility to utilize this system for the detection of pathogen microorganisms present in low amount in food and beverage samples.

Go To Sensors and Actuators B: Chemical

 

Global Medical Discovery features paper: Ischemic Injury of the Papillomacular Bundle Is a Predictive Marker of Poor Vision in Eyes With Branch Retinal Artery Occlusion

Significance Statement

Retinal artery occlusion (RAO) is a relatively common, visually disabling, ocular vascular occlusive disorder. Although most of central retinal artery occlusion showed poor visual prognosis, branch retinal artery occlusion (BRAO) is recognized as a disease with a relatively favorable outcome. However, in clinical situation, we often see patients who present with severely deteriorated central vision and this can sometimes not be explained with a classic diagnostic approach, fundus photography (FP) and fluorescein angiography (FA). Accordingly, using spectral-domain optical coherence tomography (SD-OCT), we investigated the retinal structural changes in eyes with macula-involving branch retinal artery occlusion and searched for anatomic factors that could predict the visual prognosis.

This study is designed as retrospective comparative case study and reviewed medical records of 66 eyes from 66 patients with acute branch retinal artery occlusion who visited Seoul National University Bundang Hospital between January 2009 and October 2014. Initial FP, FA, and SD-OCT images were compared with those obtained at the final images which was at least 3months after the initial visit.

 Through analysis of FP, FA, and SD-OCT images, we noticed that ischemic injury of papillomacular bundle is a determining factor of poor vision and it has a degree-dependent association with the visual prognosis of branch retinal artery occlusion. Interestingly, edema or morphological change of central fovea were absent in all patients in this study. The central region with the highest visual acuity in the retina is known as the fovea. In the central fovea, there is a depression, known as the foveal pit, which results from a lack of overlying inter-neurons. The central fovea is known to be composed of outer retinal structures lacking an inner retinal layer. Therefore, the papillomacular bundle does not include the central fovea. Importantly, in our study we found that even the normal structures of both the central fovea and the optic disc can show poor vision if accompanied by ischemic changes of the papillomacular bundle that connects the fovea and the optic disc. This implies that the crucial factor affecting vision loss in branch retinal artery occlusion is not the state of the central fovea, but the degree of inner-retinal ischemia in the papillomacular bundle area.

Despite a few limitations inherent to a retrospective study design, the present study, with a relatively large number of analyzed cases, demonstrated for the first time that the integrity of papillomacular bundle is the key factor for visual outcome of branch retinal artery occlusion.

In conclusion, there is a degree-dependent association between ischemic injury of the papillomacular bundle and visual prognosis in eyes with macula-involving branch retinal artery occlusion. Our findings emphasize the importance of a detailed papillomacular bundle area evaluation using SD-OCT in the acute stage of branch retinal artery occlusion, because the degree of papillomacular bundle invasion shows significant association with visual prognosis.

Figure legend: Representative photographic images of the fundus and horizontal foveal scan of spectral domain-optical coherence tomography at the initial visit in the Improvement group (Top row) and the Non-Improvement group in the Poor Vision group (Second row) in patients with branch retinal artery occlusion. Dashed eclipses indicate morphological changes to the inner-retinal layer of the papillomacular bundle area. The Improvement group showed only focal hyperreflectivity change, however, the Non-improvement group showed increased retinal thickness, hyper-reflectivity, and loss of layer by layer integrity in the papillomacular bundle. In the three-dimensional images of Good Vision (Third row) and Poor Vision–Non improvement group (Bottom row), the yellow line indicates an imaginary line of the papillomacular bundle and the white line indicates the ischemic retinal area. The invasion of the papillomacular bundle area by ischemic retinal injury can be seen in the Poor Vision-Non improvement group, but was absent in the Good Vision group.

About The Author

Se Joon Woo, MD, is an associate professor of Seoul National University Bundang Hospital, Seongnam, South Korea. His research interests include development of new drugs and biomarkers for age-related macular degeneration, ocular drug delivery, vascular biology, and clinical research on retinal artery occlusion, diabetic retinopathy, myopia, central serous chorioretinopathy, polypoidal choroidal vasculopathy, vitreoretinal surgery, genetics, and epidemiology. 

About The Author

Kwan Hyuk Cho, MD, is a clinical fellow at Seoul National University Bundang Hospital, Seongnam, South Korea. His current clinical research interests include retinal artery occlusion, diabetic retinopathy, various macular diseases including epiretinal membrane and vitreoretinal surgery. 

Journal Reference

Am J Ophthalmol. 2016 Feb;162:107-120.e2.

Cho KH¹, Ahn SJ², Jung C³, Han MK⁴, Park KH¹, Woo SJ⁵.

Show Affiliations

1. Department of Ophthalmology, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea.
2. Department of Ophthalmology, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea; Department of Ophthalmology, Armed Forces Capital Hospital, Seongnam, South Korea.
3. Department of Radiology, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea.
4. Department of Neurology, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea.
5. Department of Ophthalmology, Seoul National University College of Medicine, Seoul National University Bundang Hospital, Seongnam, South Korea. Electronic address: sejoon1@snu.ac.kr.

Abstract

PURPOSE:

To propose a novel prognostic feature of spectral-domain optical coherence tomography (SDOCT) in macula-involving branch retinal artery occlusion (BRAO).

DESIGN:

Retrospective comparative case study.

METHODS:

We analyzed 66 eyes diagnosed with acute branch retinal artery occlusion involving the macula from our hospital retinal artery occlusion registry. At presentation, a detailed ophthalmic and medical history was obtained from all patients, and all underwent a comprehensive ophthalmic evaluation, which included visual acuity examination, fundus photography, fluorescein angiography, and SDOCT. This evaluation was performed at each follow-up visit.

RESULTS:

The 66 eyes diagnosed with acute branch retinal artery occlusion involving the macula were divided into 2 groups according to initial vision: Good Vision (≥20/40, 29 eyes, 44%) and Poor Vision (<20/40, 37 eyes, 56%). The Poor Vision group was further divided into Improvement (18 eyes, 27%) and Nonimprovement (19 eyes, 28%) groups, according to visual recovery at the final examination. Among multiple OCT parameters, the involvement ofpapillomacular bundle, but not that of the central fovea, was consistently observed in the Poor Vision group (P < .001) and more significantly in the Nonimprovement group (P < .001). Papillomacular bundle involvement features included signs of inner retinal ischemia, including inner retinalthickening, inner retinal hyperreflectivity, and loss of layer-by-layer integrity. Loss of layer-by-layer integrity was seen consistently in the Nonimprovement group. Quantitative analysis of inner retinal thickness also supported this association.

CONCLUSION:

In eyes with macula-involving branch retinal artery occlusion, ischemic injury of the papillomacular bundle at the acute stage, as seen on OCT, correlates closely with poor vision and can explain the poor visual prognosis.

Copyright © 2016 Elsevier Inc. All rights reserved.

Go To Am J Ophthalmol