Global Medical Discovery features paper: Inkjet-Print Micromagnet Array on Glass Slides for Immunomagnetic Enrichment of Circulating Tumor Cells

Significance Statement

To advance cancer studies and enable oncologists to make more accurate diagnosis, prognosis, and develop personalized therapies, Prof. John X.J. Zhang and Dr. Peng Chen led a team of bioengineers from the Thayer School of Engineering at Dartmouth and the University of Texas at Austin in developing an innovative system that combines inkjet-printing, microfluidic technology, and immunoassay for capturing and manipulating circulating tumor cells (CTCs). This miniaturized bio-analytical system can be further integrated with fluorescent microscope for cancer cell imaging, with fluorescent in-situ hybridization (FISH) and PCR for molecular level cancer studies. It opens up a great many possibilities to optimize cancer diagnostic and tumor management.

“This work provides a new angle, other than traditional tissue biopsy and medical imaging, to approach cancer. A fast, simple and less painful blood test may eventually provide equivalently accurate disease information about individual cancer patient in a timely manner, ” said Zhang. circulating tumor cells detach from primary tumor site, shed into the blood circulation system, and may initiate the deadly cancer metastasis process. Through efficient detection of these cells, especially at an early stage, physicians will be able to design the best diagnosis and treatment strategy for each individual cancer patient.

Immunomagnetic assay has been successfully used to separate rare circulating tumor cells from blood. However, traditional immunomagnetic assay is often limited in the low magnetic field gradient and low density of effective magnetic traps. In this work, Dr. Chen developed a novel method using inkjet-printing technology to fabricate microscale magnetic structures that can be easily deposited on an arbitrary substrate.

This inkjet-printing technology is a versatile but cost-effective approach for rapid prototyping with high accuracy and flexibility. Such microscale magnets, when placed in an external magnetic field, largely enhance the magnetic field and the attractive force applied on the target circulating tumor cells, and hence facilitates the detection. In our experiments with COLO205 (a human colorectal cancer cell line) as the separation target, the inkjet-printed micro-magnets integrated assay increased the system sensitivity by 26% compared with using normal glass slide as the substrate.

Dr. Chen’s research is focused on developing miniaturized high-performance bio-analytical systems for point-of-care and globally relevant medical diagnostic applications. “This project is to implement advanced engineering fabrication, sensing, and manipulating techniques with multiplexing immunoassay to separate these rare circulating tumor cells from whole blood.” Zhang said. “The ultimate goal is to deliver a highly sensitive, reliable, affordable, and portable cancer screening platform with high throughput. The next step – bringing the technology from lab bench to clinics is a big challenging step, but is also the most rewarding step.

This work has the potential to revolutionize cancer risk assessment, cancer management, and increase the cure rate for cancers such as breast cancer, lung cancer, and prostate cancer.” This paper is published on Annals of Biomedical Engineering (DOI: 10.1007/s10439-015-1427-z).

      

About The Author

Dr. Peng Chen received his PhD degree in Biomedical Engineering from the University of Texas at Austin. He is now working as a postdoc research associate at the Center for Applied NanoBioscience and Medicine at the University of Arizona. His research interests include (1) miniaturized microfluidic system for detection and analysis of rare circulating tumor cells (CTCs), and (2) vertical flow paper based microfluidic device as point-of-need multiplex diagnostic and surveillance tool for bio-threat detection. He has published 10 peer-reviewed journal papers and 10 conference proceedings. He now serves the editorial board of Scientific Reports (Nature publishing group). Email: pengchen@email.arizona.edu

About The Author

Prof. John Zhang, Ph.D., is a Professor at Thayer School of Engineering at Dartmouth, and a Fellow of American Institute for Medical and Biological Engineering (AIMBE). He received his Ph.D. from Stanford University, and was a Research Scientist at MIT. His key contribution is in developing miniature medical systems to improve global health, through innovations in bio-inspired nanomaterials, lab-on-chip design, and advanced nanofabrication technologies for probing complex biological networks critical to human development and diseases such as cancer. He received the Wallace Coulter Foundation Early Career Award for developing handheld microphotonic imaging scanners and microsystems for early oral cancer detection; NSF CAREER award for the invention of plasmonic scanning probes design for controlled perturbation and imaging at sub-cellular level; and DARPA Young Faculty Award for patterning plasmonic surface on MEMS for biomarker sensing applications. He has published over 120 peer reviewed papers and proceedings, presented over 45 invited seminars worldwide, and filed over 60 US and international patents. He is an alumnus of NAE Frontiers of Engineering programs, an Associate Editor for Biomedical Microdevices, IEEE/ASME Journal of Microelectromechanical Systems, and has published a textbook for undergraduates titled “Molecular Sensors and Nanodevices: Principles, Designs and Applications in Biomedical Engineering”. Email: john.zhang@dartmouth.edu

Reference

Ann Biomed Eng. 2016;44(5):1710-20.

Peng Chen¹, Yu-Yen Huang², Gauri Bhave¹, Kazunori Hoshino³, Xiaojing Zhang^4,5

Show Affiliations

¹Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, 78712, USA.

²Thayer School of Engineering, Dartmouth College, Hanover, NH, 03755, USA.

³Department of Biomedical Engineering, University of Connecticut, Storrs, CT, 06269, USA.

⁴Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, 78712, USA.

⁵Thayer School of Engineering, Dartmouth College, Hanover, NH, 03755, USA. john.zhang@dartmouth.edu.

We report an inkjet-printed microscale magnetic structure that can be integrated on regular glass slides for the immunomagnetic screening of rare circulating tumor cells . circulating tumor cells detach from the primary tumor site, circulate with the bloodstream, and initiate the cancer metastasis process. Therefore, a liquid biopsy in the form of capturing and analyzing circulating tumor cells may provide key information for cancer prognosis and diagnosis. Inkjet printing technology provides a non-contact, layer-by-layer and mask-less approach to deposit defined magnetic patterns on an arbitrary substrate. Such thin film patterns, when placed in an external magnetic field, significantly enhance the attractive force in the near-field close to the circulating tumor cells to facilitate the separation. We demonstrated the efficacy of the inkjet-print micromagnet array integrated immunomagnetic assay in separating COLO205 (human colorectal cancer cell line) from whole blood samples. The micromagnets increased the capture efficiency by 26% compared with using plain glass slide as the substrate.

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Global Medical Discovery features paper: Highly specific quantification of microRNA by coupling probe-rolling circle amplification and Förster resonance energy transfer

Significance Statement

The first microRNA (miRNA) was discovered in C. elegans in 1993. Subsequently, miRNAs have been identified as a class of small non-coding RNAs containing around 21 nucleotides and virtually exist in all mammalian cells. Currently, miRNAs have been regarded as a class of potential diagnostic markers or therapeutic targets for cancer, cardiovascular and others diseases.

The short length, the similarity of nucleotide sequence and extremely low level in living cells have seriously limited the detection of miRNAs with ultimate sensitivity, high specificity and superior accuracy. Up to date, such a limitation has not been adequately overcome by reported methods. In this paper, the authors presented a novel approach (pRCA-FRET) for quantitative detection of miRNA based on a sequential combination of padlock probe-rolling circle amplification (p-RCA) and förster resonance energy transfer (FRET). First, the padlock probe is specifically ligated and circularized with the target miRNA to produce a long ssDNA by p-RCA in the presence of a DNA polymerase. In this step, p-RCA can superiorly distinguish the mismatch between miRNA and padlock probe and increase the specificity towards target miRNAs. Then, FRET probes labeled with florescent groups Cy3 and Cy5 are used to hybridize the ssDNA from p-RCA. After the excitation of the donor fluorophore Cy3, detectable readouts from the fluorescence emission of Cy5 are generated by FRET. Consequently, the target miRNA can be quantified by analyzing the intensity of fluorescent emission. Compared to previous methods, the sensitivity and operational simplicity have obviously been improved in the step of FRET without complex PCR amplification, accompanying a further enhancement of specificity for miRNA detection.

Using pRCA-FRET, the detection limit of miRNA quantification is markedly reduced from fM level to 103 aM, and remarkable specificity is exemplified to differentiate single-base mismatch between target miRNA and similar miRNAs. Accordingly, this pRCA-FRET has the potential to quantify low amount of miRNA with excellent sensitivity and specificity for the exploration of the biological functions of miRNAs and their clinical applications.  

About The Author

Prof. Dr. Yijun Chen received a B.S. degree in pharmaceutical science at China Pharmaceutical University, Nanjing, China, in 1982 and completed his Ph.D. degree in medicinal and natural products chemistry at University of Iowa, Iowa, USA, in 1996 under the guidance of Professor John P. N. Rosazza. After his postdoctoral research with Professor Edward A. Dennis at University of California, San Diego, he worked as a Senior Scientist at MicroGenomics, Inc., California, USA from 1998 to 1999. From 1999 to 2006, he served as a Senior Research Investigator at Lead Discovery and Enzyme Technology Departments of Bristol-Myers Squibb Pharmaceutical Research Institute in New Jersey, USA. Since 2007, he has been appointed as an Endowed Professor and Director of Laboratory of Chemical Biology at China Pharmaceutical University, and he presently is also an Adjunct Professor of Department of Chemical Biology, Rutgers University, USA. Currently, he has published over 80 peer-reviewed papers and invented more than 40 patents. The research of Dr. Chen’s group focuses on discovery and validation of novel antitumor targets using chemical proteomics approach, elucidation and manipulation of biosynthetic pathways for microbial secondary metabolites and development of biocatalytic routes for chiral drug intermediates. 

Journal Reference

Anal Biochem. 2016 Jun 1;502:16-23.

Wu X¹, Zhu S¹, Huang P¹, Chen Y².

Show Affiliations

1. State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu Province, 210009, People’s Republic of China.
2. State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University, Nanjing, Jiangsu Province, 210009, People’s Republic of China. Electronic address: yjchen@cpu.edu.cn. 

Abstract

MicroRNA (miRNA) plays vital roles in various biological processes. In general, sensitivity and specificity are the major parameters for the quantification of miRNA. In this study, padlock probe-rolling circle amplification and Förster resonance energy transfer (pRCA-FRET) were coupled for specific and quantitative detection of miRNA. pRCA-FRET showed superior specificity to differentiate single-base mismatch and excellent sensitivity with a detection limit of 103 aM. The current method has the potential to quantify low amounts of miRNA in the same family for studies on their biological functions.

Copyright © 2016 Elsevier Inc. All rights reserved.

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Global Medical Discovery features paper: Diagnostic Utility of Flow Cytometry Analysis of Reactive T Cells in Nodular Lymphocyte-Predominant Hodgkin Lymphoma

About The Author

James Huang, M.D.

Oakland University William Beaumont School of Medicine, Rochester, MI, USA

James Huang undertook his early medical studies at Hunan Medical University, Changsha, China, completing a research fellowship at Harvard Medical School, a pathology residency at Dartmouth-Hitchcock Medical Center, a hematopathology fellowship in University of Nebraska Medical Center. He is currently an Associate Professor of Pathology, Co-Director of Diagnostic Medicine Clerkship, and Director of Clinical Pathology Elective at Oakland University William Beaumont School of Medicine. He is also a senior staff hematopathologist at Beaumont Health.

Dr. Huang is on the Editorial Board of a number of journals including International Journal of Pathology and Clinical Research, the Clinics in Oncology, and Pathology- Remedy Open Access.  

About The Author

James David, M.D.

Oakland University William Beaumont School of Medicine, Rochester, MI, USA

James David graduated from Oakland University William Beaumont School of Medicine. This research was his capstone project for which he received a competitive scholarship award form Oakland University William Beaumont School of Medicine. He is currently in residency program in ophthalmology at Louisiana State University-Ochsner, Louisiana.  

Journal Reference

Am J Clin Pathol. 2016 Jan;145(1):107-15.

David JA¹, Huang JZ².

Show Affiliations

1. From the Department of Pathology, Oakland University William Beaumont School of Medicine, Rochester, MI;
2. From the Department of Pathology, Oakland University William Beaumont School of Medicine, Rochester, MI; Department of Clinical Pathology, William Beaumont Hospital, Royal Oak, MI. james.huang@beaumont.edu.

Abstract

OBJECTIVES:

This study aims to define the diagnostic utility of flow cytometric features of T cells in  nodular  lymphocyte- predominant Hodgkin lymphoma (NLPHL).

METHODS:

Cases were retrospectively identified based on diagnosis with NLPHL (n = 30 samples), classic  Hodgkin lymphoma (CHL; n = 33), and reactive lymphoid hyperplasia (RLH; n = 43). Pathology slides were reviewed. Flow cytometry list mode data were reanalyzed.

RESULTS:

The mean proportion of CD4 + CD8 + T cells (8.4%) in cases of NLPHL was significantly higher than seen in CHL (1.0%) or RLH (0.6%). Of the T cells, 28.4% were CD57 + in NLPHL, significantly higher (P < .05) than in CHL (3.2%) or RLH (3.2%). Based on receiver operating characteristic curve analysis, when using a cutoff of 3.0% of CD4 + CD8 + T cells, the diagnostic sensitivity for NLPHL is 83.3% with a specificity of 97.4%. The diagnostic sensitivity was 96.7% with a specificity of 98.7% when using a cutoff of 12% for CD57 + T cells.

CONCLUSIONS:

Increased portions of CD57 + T cells and CD4 + CD8 + T cells are highly suggestive of the possibility of NLPHL. In addition, NLPHL diagnosis appears unlikely if neither CD57 + T cells nor CD4 + CD8 + T cells are increased. Future prospective studies including cases of progressive transformation of germinal center and T-cell/histiocyte-rich large B-cell lymphoma will further define the  utility  of  flow cytometry of T cells in NLPHL.

© American Society for Clinical Pathology, 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

 

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Global Medical Discovery features paper: Development of neuraminidase detection using gold nanoparticles boron-doped diamond electrodes

Significance Statement

Neuraminidase detection is substantial in controlling the disease caused by related virus and microbes since Neuraminidase is an important enzyme in the pathogenic viruses and microbes spreading. Common detection methods of neuraminidase, such as RT-PCR, ELISA, and enzymatic reaction require several types of chemical reagents, specific instruments, and highly skilled operators. Therefore, a more simple and practical method is urgently required. In this work, a sensor for Neuraminidase has been developed using gold nanoparticles modified boron-doped diamond (AuNPs-BDD) electrodes. The detection method was performed based on the difference of electrochemical responses of Zanamivir at gold surface before and after the reaction with neuraminidase in phosphate buffer solution (PBS) pH 5.5.

In order to have a stable response, the modified electrodes was prepared through a self-assembly deposition of AuNPs at an amine-terminated boron-doped diamond. Cyclic voltammetry was used for the method. A linear calibration curve for Zanamivir in 0.1 M PBS in the absence of neuraminidase was achieved in the concentration range of 1 x 10^-6 – 1 x 10^-5 M (R² = 0.99), with the estimated limit of detection (LOD) of 2.29 x 10^-6 M.

Furthermore, using its reaction with 1.00 x 10^-5 M Zanamivir a linear calibration curve of neuraminidase can be obtained in the concentration range of 0 – 12 mU (R² = 0.99) with an estimated LOD of 0.12 mU. High reproducibility was shown with an RSD of 1.14 % (n=30). These performances could be maintained when the detection was performed in mucin matrix. Comparison made using gold-modified BDD (Au-BDD) electrodes suggested that the good performance of the detection method is due to the stability of the gold particles position at the BDD surface.

About The Author

Dr. Tribidasari A. Ivandini is a member of the teaching and research staff at Department of Chemistry, Faculty of Mathematics and Science, Universitas Indonesia, since 1997. She was born in Jakarta, Jakarta, on January 29th, 1970. She received her magister degrees in 1996 from Universitas Indonesia. Then, in 2003 she accomplished her doctoral degree at The University of Tokyo, Tokyo, Japan. Between 2003 and 2007 she performed post-doctoral research at the Department of Chemistry, Keio University, Tokyo, Japan with JSPS fellowship during 2004-2006. Up to the present time she is still performing research collaboration with Keio University.

Her research interests are mostly concerning electrochemistry of diamond, including its development for sensors and biosensors, electrocatalytics, and waste-water treatments. She has published about 50 scientific papers in highly respectable journals and conference proceedings, such as Analytical Chemistry, Physical Chemistry Chemical Physics, Sensors Actuators B: Chemicals, and Diamond Related Materials. She also holds some patents under European and Japan Patents. Ivandini can be reached at the Department of Chemistry, Faculty of Mathematics and Science, Universitas Indonesia, Indonesia.E-mail : ivandini.tri@sci.ui.ac.id   

About The Author

Dr. Yasuaki Einaga is a professor in the Department of Chemistry at Keio University, Japan.  He received his BS (1994) , MS (1996) , and PhD (1999) degree from The University of Tokyo. After 2 years as a research associate at The University of Tokyo, he started a faculty career as an assistant professor in Keio University in 2001. He was promoted to associate professor in 2003, and to professor in 2011.  He was also a research director of JST-CREST (2011-2014), and JST-ACCEL (2014-present). His research interests include functional materials science, photochemistry, electrochemistry, and diamond electrodes. Einaga can be reached at the Department of Chemistry, Keio University, Yokohama, Japan. E-mail einaga@chem.keio.ac.jp  

Journal Reference

Anal Biochem. 2016 Mar 15;497:68-75.

Wahyuni WT¹, Ivandini TA², Saepudin E¹, Einaga Y³.

Show Affiliations

1. Department of Chemistry, Faculty of Mathematics and Sciences, University of Indonesia, Kampus UI Depok, Jakarta 16424, Indonesia.
2. Department of Chemistry, Faculty of Mathematics and Sciences, University of Indonesia, Kampus UI Depok, Jakarta 16424, Indonesia. Electronic address: ivandini.tri@ui.ac.id.
3. Department of Chemistry, Faculty of Science and Technology, Keio University, Yokohama 223-8522, Japan; JST ACCEL, Yokohama 223-8522, Japan. Electronic address: einaga@chem.keio.ac.jp.

Abstract

Gold nanoparticles-modified boron-doped diamond (AuNPs-BDD) electrodes, which were prepared with a self-assembly deposition of AuNPs at amine-terminated boron-doped diamond, were examined for voltammetric detection of neuraminidase (NA). The detection method was performed based on the difference of electrochemical responses of zanamivir at gold surface before and after the reaction with neuraminidase in phosphate buffer solution (PBS, pH 5.5). A linear calibration curve for zanamivir in 0.1 M PBS in the absence of NA was achieved in the concentration range of 1 × 10(-6) to 1 × 10(-5) M (R(2) = 0.99) with an estimated limit of detection (LOD) of 2.29 × 10(-6) M. Furthermore, using its reaction with 1.00 × 10(-5) M zanamivir, a linear calibration curve of neuraminidase can be obtained in the concentration range of 0-12mU (R(2) = 0.99) with an estimated LOD of 0.12mU. High reproducibility was shown with a relative standard deviation (RSD) of 1.14% (n = 30). These performances could be maintained when the detection was performed in mucin matrix. Comparison performed using gold-modified BDD (Au-BDD) electrodes suggested that the good performance of the detection method is due to the stability of the gold particles position at the BDD surface.

Copyright © 2016 Elsevier Inc. All rights reserved.

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Global Medical Discovery features paper: Digital PCR for direct quantification of viruses without DNA extraction

Significance Statement

 DNA extraction before amplification is considered an essential step for the measurement of any DNA targets, as it releases the DNA from cells, viruses and bacteria and eliminates any PCR-inhibitory substances that are initially present in the matrix. However, different extraction procedures have different DNA yields and efficiency of inhibitory substances removal, consequently reducing quantification accuracy and hampering inter-laboratory comparability of data. In the past some viruses and bacteria have already been directly quantified using qPCR without DNA extraction, however drawbacks related to the qPCR formats such as susceptibility to inhibitory substances and dependence on the calibration material probably hampered the precise and reliable measurements of the actual concentrations therefore direct quantification using dPCR has a potential to overcome these.

To investigate the potential of digital PCR for direct quantification of viruses, human cytomegalovirus (HCMV) has been quantified without prior extraction using two digital PCR platforms and two standard materials. Up to 50% higher concentrations of DNA targets were measured using direct quantification compared to concentrations measured after the extraction step. Results of direct quantification were equally or even more repeatable, than that of the extracted DNA.

Direct quantification can be used to determine absolute DNA extraction efficiencies and for the evaluation of the influence of matrix on the extraction yield and therefore supports laboratories in decisions on fit for purpose extraction methods as well as in comparability of results especially in diagnostics.

Direct quantification is also a valuable approach for the characterization of reference materials and materials for external quality assurance schemes that underpin quality of results in diagnostics of human pathogens. This approach is of particular importance in the field of plant pathogen diagnostics, where no reference materials are commercially available as it enables preparation of well characterized in-house reference materials. 

About The Author

Jernej Pavšič is a PhD student at the National Institute of Biology in Ljubljana, Slovenia.

 His research is focused on evaluation of novel nucleic-acid based techniques to standardize and improve diagnostics of human pathogens, enhance characterisation of reference materials and facilitate cancer diagnostics. He is especially interested is exploring capabilities of digital PCR and high-throughput qPCR.

About The Author

Jana Žel is head of the genetically modified organisms group at the National Institute of Biology, Slovenia. She has been working for several years on cutting-edge approaches and methods for GMO testing and also the establishment of an internationally harmonized and metrologically sound system for molecular analyses.

About The Author

Mojca Milavec is deputy head of GMO detection at the National Institute of Biology in Ljubljana, Slovenia, where she is responsible for introduction of new methods, in house validations and staff training for the National Reference Laboratory for GMO detection. Her research is focused on challenges associated with established and novel technologies for nucleic acid measurements as well as standardization and quality control of these technologies.

Journal Reference

Anal Bioanal Chem. 2016;408(1):67-75.

Pavšič J^1,2, Žel J³, Milavec M³.

Show Affiliations

1. Department of Biotechnology and Systems Biology, National Institute of Biology, Večna pot 111, 1000, Ljubljana, Slovenia. jernej.pavsic@nib.si.
2. Jožef Stefan International Postgraduate School, Jamova cesta 39, 1000, Ljubljana, Slovenia. jernej.pavsic@nib.si.
3. Department of Biotechnology and Systems Biology, National Institute of Biology, Večna pot 111, 1000, Ljubljana, Slovenia.

 

Abstract

DNA extraction before amplification is considered an essential step for quantification of viral DNA using real-time PCR (qPCR). However, this can directly affect the final measurements due to variable DNA yields and removal of inhibitors, which leads to increased inter-laboratory variability of qPCR measurements and reduced agreement on viral loads. Digital PCR (dPCR) might be an advantageous methodology for the measurement of virus concentrations, as it does not depend on any calibration material and it has higher tolerance to inhibitors.  DNA quantification without an extraction step (i.e. direct quantification) was performed here using digital PCR and two different human cytomegalo virus whole-virus materials. Two digital PCR platforms were used for this direct quantification of the viral DNA, and these were compared with quantification of the extracted viral DNA in terms of yield and variability. Direct quantification of both whole-virus materials present in simple matrices like cell lysate or Tris-HCl buffer provided repeatable measurements of virus concentrations that were probably in closer agreement with the actual viral load than when estimated through quantification of the extracted DNA. Direct digital PCR quantification of other viruses, reference materials and clinically relevant matrices is now needed to show the full versatility of this very promising and cost-efficient development in virus quantification.

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Global Medical Discovery features paper: Development of green to near-infrared turn-on fluorescent probes for the multicolour imaging of nitroxyl in living systems

Journal Reference

Mater. Chem. B, 2016,4, 1263-1269.

Baoli Dong¹ , Kaibo Zheng² , Yonghe Tang¹ , Weiying Lin^1,2

Show Affiliations

1. Institute of Fluorescent Probes for Biological Imaging, School of Chemistry and Chemical Engineering, School of Biological Science, University of Jinan, Jinan, P. R. China
   E-mail: weiyinglin2013@163.com
2. State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan university, Changsha, P. R. China

Abstract

Nitroxyl (HNO) is one of the important reactive nitrogen species (RNS) and show significant biological activities with significant therapeutic potential. Herein, three novel turn-on probes (NP-1–3) based on structurally related dyes with different emission colors as fluorescent scaffolds have been developed for detecting HNO in biological systems. The probes exhibit high sensitivity, excellent selectivity, desirable performance at physiological pH and low cytotoxicity. By incubating living cells with these probes simultaneously, we demonstrate the multicolor imaging of HNO with emission colors in the range of green to near-infrared (NIR) in living systems for the first time. Furthermore, probe NP-3 responds to HNO with a significant turn-on NIR fluorescence signal upon excitation in the NIR region, and it is successfully applied for sensing HNO in living mice.

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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.

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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

 

Trace analysis of three fungicides in animal origin foods with a modified QuEChERS method and liquid chromatography-tandem mass spectrometry

Significance Statement

Most people ignore the risk of pesticide residues in biological materials of animal origin, which makes people more vulnerable to some potential epidemics. However, plant materials, such as straws, grains and leaves, which used for feeding livestock may be contaminated, and, consequently, pesticides may be consumed by humans via animal feed. Due to the lipophilic nature of some pesticides, they are accumulated in milk, meat or animal offal and enter the human body through the food chain, and then may cause serious health problems. Increasing attention has been paid to the risks posed to consumers by pesticide residues in animal foods. However, as far as we know, few articles referred to the pesticide detection in animal origin foods. Amongst commercial pesticides, propiconazole , pyraclostrobin and isopyrazam , three of most widely used fungicides in the world, deserves special attention because of its high effectiveness against diseases affecting cereals, fruits, and vegetables. The key of our study is to use the same method to achieve simultaneous detection for the three fungicides in seven animal origin foods, which will save labor, time and cost significantly and make it more suitable for the detection of large-scale real samples.

Journal Reference

Anal Bioanal Chem. 2016 Feb;408(5):1515-22.

Mu Z¹, Feng X¹, Zhang Y¹, Zhang H².

Show Affiliations

1. College of Science, China Agricultural University, Beijing, 100193, China.
2. College of Science, China Agricultural University, Beijing, 100193, China. hongyan@cau.edu.cn.

Abstract

A multi-residue method based on modified QuEChERS (quick, easy, cheap, effective, rugged, and safe) sample preparation, followed by liquid chromatography tandem mass spectrometry (LC-MS/MS), was developed and validated for the determination of three selected fungicides (propiconazole, pyraclostrobin, and isopyrazam) in seven animal origin foods. The overall recoveries at the three spiking levels of 0.005, 0.05, and 0.5 mg kg(-1) spanned between 72.3 and 101.4 % with relative standard deviation (RSD) values between 0.7 and 14.9 %. The method shows good linearity in the concentrations between 0.001 and 1 mg L(-1) with the coefficient of determination (R (2)) value >0.99 for each target analyte. The limit of detections (LODs) for target analytes were between 0.04 and 1.26 μg kg(-1), and the limit of quantifications (LOQs) were between 0.13 and 4.20 μg kg(-1). The matrix effect for each individual compound was evaluated through the study of ratios of the areas obtained in solvent and matrix standards. The optimized method provided a negligible matrix effect for propiconazole within 20 %, whereas for pyraclostrobin and isopyrazam, the matrix effect was relatively significant with a maximum value of 49.8 %. The developed method has been successfully applied to the analysis of 210 animal origin samples obtained from 16 provinces of China. The results suggested that the developed method was satisfactory for trace analysis of three fungicides  in animal  origin foods. Graphical abstract Schematic representation of the proposed residue analytical  method for  animal  origin foods using  QuEChERS and LC-MS/MS.

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Serum α-klotho concentrations during preimplantation can predict aging or quality of human oocytes and clinical pregnancy rates

Significance Statement

 Many researches in the world are very interested in the aging or quality of human oocytes. However, a simple biomarker to predict the aging or quality of human oocytes is not discovered. On the other hand, α-klotho is known as an anti-aging molecule since 1997 (Kuro-o M, et al., Nature). Therefore, increases in α-klotho concentrations in human serum positively may improve the aging and quality of human oocytes.

The present research (a prospective cohort study) is the first study reporting that the serum α-klotho concentrations during preimplantation can predict the aging or quality of human oocytes and clinical pregnancy rates.

Furthermore, the present research suggests that the serum α-klotho concentrations during preimplantation may be improved by vitamin D supplementation. Therefore, the aging or quality of human oocytes may be improved by vitamin D supplementation, although clinical trials will be needed in the near future.  In conclusion, the present research is a breakthrough study in reproductive medicine.

Journal Reference

Springerplus. 2016 Jan 20;5:53.

Takemura T, Okabe M.

Reproductive Medicine Institute Japan, Nakano-ku, Chuo, 3-37-12, Tokyo, 164-0011 Japan.

Abstract

BACKGROUND:

To discover simple biomarkers to evaluate the aging or quality of human oocytes and clinical pregnancy rates is needed. However, the association among serum α-klotho concentrations during preimplantation, the aging  or quality of human oocytes and clinical pregnancy rates has not been investigated.

FINDINGS:

The serum α-klotho concentrations during preimplantation decreased due to aging (p < 0.001), whereas the maturation rates of human oocytes (p < 0.001) and the fertilization rates (p < 0.001) improved in association with increased serum α-klotho concentrations. Furthermore, multiple logistic regression analysis showed that the clinical pregnancy rates were influenced by serum α-klotho concentrations during preimplantation (p < 0.001), the patient’s age (p = 0.003), maturation rates of  human oocytes (p < 0.001), fertilization rates (p < 0.001) and the serum 25 (OH) D levels (p < 0.001) regardless of race (p = 0.29) and BMI (p = 0.96).

CONCLUSION:

The serum α-klotho concentrations during preimplantation would be a simple biomarker in order to  predict the aging or quality of human oocytes and clinical pregnancy rates.

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A new strategy to develop the disposable label-free immunosensor with electrochemiluminescent probing

Journal Reference

Journal of Electroanalytical Chemistry, Volume 747, 2015, Pages 136–142.

Lin Jiang¹, Ya Yang^1,2 , Yifeng Tu^1,3

Show Affiliations

1. Institute of Analytical Chemistry, Soochow University, Dushu Lake Campus of Soochow University, Suzhou Industrial Park, 215123 Suzhou, PR China
2. Department of Forensic Medicine, Soochow University, Dushu Lake Campus of Soochow University, Suzhou Industrial Park, 215123 Suzhou, PR China
3. The Key Lab of Health Chemistry and Molecular Diagnosis of Suzhou, Dushu Lake Campus of Soochow University, Suzhou Industrial Park, 215123 Suzhou, PR China

Abstract

In this paper, a new strategy to prepare the label-free immunosensor signaling with the electrochemiluminescence (ECL) of luminol will be reported. The gold nano-particles (AuNPs) functionalized indium tin oxide coated glass is used as the substrate for antibody immobilization. These AuNPs also improve the ECL behavior of luminol. This strategy for immunosensor construction is simple, fast, cheap, and with no requirements of expensive apparatus. For validating the practicability of this new strategy, with methamphetamine (MA) as target model, an methamphetamine sensor was built by immobilizing the methamphetamine-antibody onto the surface of previously mentioned substrate. The ECL intensity on resultant sensor, after the direct immuno-interaction, decreased proportional to the concentration of methamphetamine in a wide linear range from 2 to 500 ng mL⁻¹ with a detection limit of 0.3 ng mL⁻¹. This proposed sensor is sensitive, specific, stable and reliable. It has been successfully applied to detect the methamphetamine in spiked human serums.

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Comparative Tissue Proteomics of Microdissected Specimens Reveals Novel Candidate Biomarkers of Bladder Cancer

Significance Statement

This manuscript is focused on the biomarker discovery from clinical tissue specimens from bladder cancer patients as well as biomarker verification using several strategies. The results were integrated with a series of papers published by this group of urine proteomes [1-3] to select potential biomarker for further verification studies. Transgelin-2 (TAGLN2) and stathmin 1 (STMN1) proteins were successful verified with higher concentrations in bladder cancer tissue and urine specimens. The integrated results suggest that up-regulated TAGLN2 in bladder tumor cells may be secreted into the urine through microparticles, which in turn causes the higher concentration of urinary TAGLN2.

References:

1. Chen, Y.T., et al., Discovery of novel bladder cancer biomarkers by comparative urine proteomics using iTRAQ technology. J. Proteome Res., 2010. 9(11): p. 5803-15.
2. Chen, C.L., et al., Comparative and targeted proteomic analyses of urinary microparticles from bladder cancer and hernia patients. J. Proteome Res., 2012. 11(12): p. 5611-29.
3. Chen, C.L., et al., Identification of potential bladder cancer markers in urine by abundant-protein depletion coupled with quantitative proteomics. J. Proteomics, 2013. 85C: p. 28-43.

 

About The Author

Dr. Yi-Ting Chen received her Ph.D. degree in Department of Chemistry from the National Tsing Hua University, Taiwan. Before graduation, she went to University of Alberta, Canada as a visiting student in 2001. Following the graduation in 2001, she worked as a research scientist in the Industrial Technology Research Institute, Taiwan to establish a new mass spectrometry lab for numerous proteomic projects. From September 2007, she worked at Molecular Medicine Research Center of Chang Gung University (CGU) as an associated research scientist, with focusing on urinary protein biomarker discovery in bladder cancer. Being an assistant professor in Department of Biomedical Sciences of CGU in 2013, Dr. Chen’s research interests focus on proteomics, metabolomics and systems biology of urological diseases and advanced method development for detection of biomolecules, primarily based on mass spectrometry.

Journal Reference

Mol Cell Proteomics. 2015;14(9):2466-78.

Chien-Lun Chen¹, Ting Chung², Chih-Ching Wu³, Kwai-Fong Ng⁴, Jau-Song Yu ⁵, Cheng-Han Tsai ⁶, Yu-Sun Chang⁵, Ying Liang ², Ke-Hung Tsui ¹, Yi-Ting Chen⁷

Show Affiliations

1. From the ‡Department of Urology, Chang Gung Memorial Hospital, Taoyuan, Taiwan; §School of Medicine, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
2. Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan;
3. Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; ‖Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
4. Department of Pathology, Chang Gung Memorial Hospital, Taoyuan, Taiwan;
5. Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
6. Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan;
7. Molecular Medicine Research Center, Chang Gung University, Taoyuan, Taiwan; Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan; Department of Biomedical Sciences, College of Medicine, Chang Gung University, Taoyuan, Taiwan. ytchen@mail.cgu.edu.tw.

Abstract

More than 380,000 new cases of bladder cancer are diagnosed worldwide, accounting for ∼150,200 deaths each year. To discover potential biomarkers of bladder cancer, we employed a strategy combining laser microdissection, isobaric tags for relative and absolute quantitation labeling, and liquid chromatography-tandem MS (LC-MS/MS) analysis to profile proteomic changes in fresh-frozen bladder tumor specimens. Cellular proteins from four pairs of surgically resected primary bladder cancer tumor and adjacent nontumorous tissue were extracted for use in two batches of isobaric tags for relative and absolute quantitation experiments, which identified a total of 3220 proteins. A DAVID (database for annotation, visualization and integrated discovery) analysis of dysregulated proteins revealed that the three top-ranking biological processes were extracellular matrix organization, extracellular structure organization, and oxidation-reduction. Biological processes including response to organic substances, response to metal ions, and response to inorganic substances were highlighted by up-expressed proteins in bladder cancer. Seven differentially expressed proteins were selected as potential bladder cancer biomarkers for further verification. Immunohistochemical analyses showed significantly elevated levels of three proteins-SLC3A2, STMN1, and TAGLN2-in tumor cells compared with noncancerous bladder epithelial cells, and suggested that TAGLN2 could be a useful tumor tissue marker for diagnosis (AUC = 0.999) and evaluating lymph node metastasis in bladder cancer patients. ELISA results revealed significantly increased urinary levels of both STMN1 and TAGLN2 in bladder cancer subgroups compared with control groups. In comparisons with age-matched hernia urine specimens, urinary TAGLN2 in bladder cancer samples showed the largest fold change (7.13-fold), with an area-under-the-curve value of 0.70 (p < 0.001, n = 205). Overall, TAGLN2 showed the most significant overexpression in individual bladder cancer tissues and urine specimens, and thus represents a potential biomarker for noninvasive screening for bladder cancer. Our findings highlight the value of bladder tissue proteome in providing valuable information for future validation studies of potential biomarkers in urothelial carcinoma.

© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Go To Mol Cell Proteomics. 

Methodologic quality assessment of red blood cell transfusion guidelines and the evidence base of more restrictive transfusion thresholds

Significance Statement

The World Health Organization indicates that every second someone in the world needs a blood transfusion. Blood transfusions therefore save many lives every year, and patients can die when they don’t receive a transfusion when necessary. However, transfusions also carry certain risks, and therefore should not be performed unless really necessary.

In recent years, a lot of emphasis has been put on this second consideration, which has been more broadly rebranded as “patient blood management” (PBM). PBM is a worthwhile effort, but should be evidence-based and not based on other considerations, be it religious (e.g., refusing of an allogenic blood transfusion as a Jehovah Witness) or commercial (e.g., receiving sponsorships by commercial companies who promote RBC replacing products).

It is therefore important that transfusion guidelines are of high quality, and based on the best scientific evidence. Our analysis of the quality of the most widely used transfusion guidelines, using an internationally used tool (AGREE II), indicates that guidelines recommending red blood cell transfusion as of Hb levels of 7 to 8 g/dL are based on high quality evidence, whereas the more recent guidelines recommending red blood cell transfusion only at Hb thresholds of ≤6 g/dL are not based on solid evidence.

The methodological robustness of the published transfusion guidelines is variable and has room for improvement. Furthermore, guideline developers and reviewers should always be transparent about conflicts of interest, whether religious or commercial, which was, based on other available information, not always the case in these guidelines.

About The Author

Hans Van Remoortel, PT/PhD, physiotherapist, is staff member in the Centre for Evidence-Based Practice of the Belgian Red Cross-Flanders. After obtaining a Master in Rehabilitation Sciences and Physiotherapy in 2005, Hans worked for three years (2005-2008) as a clinical physiotherapist in cardiac rehabilitation programs in 2 Belgian hospitals (University Hospital of Leuven and Imelda Hospital Bonheiden). After working in a clinical setting, he obtained his PhD degree at the Faculty of Rehabilitation Sciences and Kinesiology at the University of Leuven (Belgium) with a project entitled ‘Physical activity and comorbidities in patients with chronic obstructive pulmonary disease’ (2009-2013).

He has published about 20 articles in peer reviewed journals, mainly in the field of exercise, physical activity, pulmonology and evidence-based medicine. In his current job, Hans is a methodologist working on the development of systematic reviews and guidelines within the relevant themes of the Belgian Red Cross, i.e. from humanitarian aid to blood supply. In his spare moments, Hans enjoys to spend time with his family, is a recreational sporter (squash, soccer, running, cycling) and enjoys to attend live music concerts.

About The Author

Philippe Vandekerckhove, M.D./PhD, pathologist, is the CEO of the Belgian Red Cross-Flanders.

Prior to this position Philippe worked as Clinical Director of the University Hospital Leuven from where he also obtained his M.D./PhD and Pathology degree. His clinical and pathology training was further carried out in South Africa (Baragwanath – University of Johannesburg, and Groote Schuur Hospital – University of Cape Town), the US (Woods Hole Marine Biology Laboratory, University of Hawaii, New York University), and The Netherlands (Erasmus University, Rotterdam).

In addition, Philippe studied healthcare management at INSEAD (France) and general management at Harvard Business School. He has published about 60 articles in peer reviewed journals, and 5 chapters in textbooks, mainly in the field of immunology, hematology, blood banking and evidence-based medicine.

Philippe is associate professor at the Faculties of Medicine of the University of Leuven and the University of Ghen. He holds non-executive positions as president of the European Blood Alliance, president of GAP, member of the governing board of the International Federation of Red Cross and Red Crescent Societies, and of the investment committee of Flanders’ Care Invest (Flemish government). In his spare moments, he enjoys jogging, natural horsemanship and time and travel spent in natural surroundings.

Journal Reference

Transfusion. 2016 Feb;56(2):472-80.

Van Remoortel H¹, De Buck E¹, Dieltjens T¹, Pauwels NS¹, Compernolle V^1,2, Vandekerckhove P^1,2,3.

Show Affiliations

1. BelgianRed Cross-Flanders, Mechelen, Belgium.
2. Faculty of Medicine, University of Ghent, Ghent, Belgium.
3. Department of Public Health and Primary Care, Faculty of Medicine, Catholic University of Leuven, Leuven, Belgium.

Abstract

BACKGROUND:

Recent literature suggests that more restrictive red blood cell (RBC) transfusion practices are equivalent or better than more liberal transfusion practices. The methodologic quality of guidelines recommending more restrictive transfusion thresholds and their underlying scientific evidence is unclear. Therefore, we aimed to evaluate the quality of the development process of red blood cell transfusion guidelines and to investigate the underlying evidence of guidelines recommending a more restrictive hemoglobin (Hb) threshold.

STUDY DESIGN AND METHODS:

Via systematic literature screening of relevant databases (NGC, GIN, Medline, and Embase), red blood cell transfusion guidelines recommending a more restrictive Hb level (<6, <7, or <8 g/dL) were included. Four assessors independently evaluated the methodologic quality by scoring the rigor of development domain (AGREE II checklist). The level of evidence served as a reference for the quality of the underlying evidence.

RESULTS:

The methodologic quality of 13 red blood cell transfusion guidelines was variable (18%-72%) but highest for those developed by Advancing Transfusion and Cellular Therapies Worldwide (72%), the Task Force of Advanced Bleeding Care in Trauma (70%), and the Dutch Institute for Healthcare Improvement (61%). A Hb level of less than 7 g/dL (intensive care unit patients) or less than 8 g/dL (postoperative patients) were the only thresholds based on high-quality evidence. Only four of 32 recommendations had a high-quality evidence base.

CONCLUSION:

Methodologic quality should be guaranteed in future RBC transfusion guideline development to ensure that the best available evidence is captured when recommending restrictive transfusion strategies. More high-quality trials are needed to provide a stronger scientific basis for RBC transfusion guidelines that recommend more restrictive transfusion thresholds.

© 2015 AABB.

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Effective use of the TSPY gene-specific copy number in determining fetal DNA in the maternal blood of cynomolgus monkeys

Journal Reference

Anim Sci J. 2015 Sep 30.

Yasmin L, Takano JI, Sankai T.

Tsukuba Primate Research Center, National Institutes of Biomedical Innovation, Health and Nutrition, Tsukuba, Japan.

Abstract

Since the available concentration of single-copy fetal genes in maternal blood DNA is sometimes lower than detection limits by PCR methods, the development of specific and quantitative PCR detection methods for fetal DNA in maternal blood is anticipated, which may broaden the methods that can be used to monitor pregnancy. We used the TaqMan qPCR amplification for DYS14 multi-copy sequence and the SRY gene in maternal blood plasma (cell-free DNA) and fractional precipitated blood cells (cellular DNA) from individual cynomolgus monkeys at 22 weeks of pregnancy. The availability of cell-free fetal DNA was higher in maternal blood plasma than that of cellular DNA from fractional precipitated blood cells. There was a significantly higher (P < 0.001) mean copy number of fetal male DYS14 from maternal plasma (4.4 × 10⁴ copies/mL) than that of detected fetal cellular DNA from fractional blood cell pellets. The sensitivity of the DYS14 PCR assay was found to be higher than that of the SRY assay for the detection of fetal DNA when its presence was at a minimum. The DYS14 assay is an improved method for quantifying male fetal DNA in circulating maternal blood in the primate model.

© 2015 Japanese Society of Animal Science.

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The influence of printing parameters on cell survival rate and printability in microextrusion-based 3D cell printing technology

Significance Statement

High cell survival rate and good printability, commonly conflicts with each other, has always been the pursuit in the practice of cell 3D printing technology. Since both can be influenced by many process parameters, Yao and her coworkers from Tsinghua University, China and Drexel University, USA focused on the establishment of an applicable protocol for microextrusion-based 3D cell printing technology to achieve both high cell survival rate and good printability. Their studies demonstrated that even though the bioink composition and concentration, holding temperature and holding time would all influence bioink rheological property in the printing period, bioink storage modulus was the decisive factor for both cell survival rate and printability. Different process parameter combinations would result in the similar storage modulus and thus showed similar cell survival rate after 3D bioprinting process. When the bioink storage modulus was in the range of 154 Pa to 382 Pa, 3D printing of A549 cell-laden gelatin/alginate construct with both high cell survival rate (>90%) and good printability could be achieved. Researchers in the bioprinting field may find this protocol useful for adjusting the printing process quickly. And the methodology and the protocol in this study may also be useful for the design and development of new temperature-sensitive bioinks for 3D cell printing. Future work will be focused on revealing the detailed mechanism and mathematical modelling of cell survival and printability in microextrusion-based 3D cell printing technology and optimizing the parameter ranges for bioprinting other types of cells, e.g. pluripotent stem cells.

About The Author

Dr. Rui Yao is Assistant Professor of Biomanufacturing Research Center, Department of Mechanical Engineering, Tsinghua University, Beijing, China. She has interdisciplinary background of tissue engineering, biomaterials, biofabrication and stem cells from Tsinghua University, MIT and Peking Union Medical University. Her main focus is Stem Cell-based Biofabrication and applications in Tissue Engineering, 3D Physiology/Pathological Model and Drug Testing Model. She has published over 40 journal papers and conference abstracts, filed over 10 patents and conducted over 10 oral presentations in international conferences in the field of her research.

 

Journal Reference

Biofabrication. 2015 Nov 2;7(4):045002.

Zhao Y, Li Y, Mao S, Sun W, Yao R.

Biomanufacturing Center, Department of Mechanical Engineering, Tsinghua University, Beijing, People’s Republic of China. Biomanufacturing and Rapid Forming Technology Key Laboratory of Beijing, Beijing, People’s Republic of China.

Abstract

Three-dimensional (3D) cell printing technology has provided a versatile methodology to fabricate cell-laden tissue-like constructs and in vitro tissue/pathological models for tissue engineering, drug testing and screening applications. However, it still remains a challenge to print bioinks with high viscoelasticity to achieve long-term stable structure and maintain high cell survival rate after printing at the same time. In this study, we systematically investigated the influence  of 3D cell printing parameters, i.e. composition and concentration of bioink, holding temperature and holding time, on the printability  and  cell survival rate in microextrusion-based 3D cell printing technology. Rheological measurements were utilized to characterize the viscoelasticity of gelatin-based bioinks. Results demonstrated that the bioink viscoelasticity was increased when increasing the bioink concentration, increasing holding time and decreasing holding temperature below gelation temperature. The decline of cell survival rate after 3D cell printing process was observed when increasing the viscoelasticity of the gelatin-based bioinks. However, different process parameter combinations would result in the similar rheological characteristics and thus showed similar cell survival rate after 3D bioprinting process. On the other hand, bioink viscoelasticity should also reach a certain point to ensure good printability and shape fidelity. At last, we proposed a protocol for3D bioprinting of temperature-sensitive gelatin-based hydrogel bioinks with both high cell survival rate and good printability. This research would be useful for biofabrication researchers to adjust the 3D bioprinting process parameters quickly and as a referable template for designing new bioinks.

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Development and validation of a HILIC- MS/MS multi-targeted method for metabolomics applications

Significance Statement

Metabolomics is a newly emerging field of “omics” research concerned with the comprehensive characterization of the small molecule basic polar metabolites in biological systems. The overall goal of metabolomics is to link the metabolite concentration information with clinical or physiology data and to assess the multi-parametric response of a living system to pathophysiological stimuli or genetic modification. Metabolites may describe phenomena which often are not successfully described or predicted by genomics or transcriptomics, such as the effects of xenobiotics or gut microbiome or the positive response or no response to pharmaceutical intervention.

The analysis of polar metabolites represents a challenging task for contemporary science. LC-MS presents a growing popularity as the platform for metabolomic studies due to its high throughput, robustness and wide coverage of metabolites. The success of a LC-MS-based metabolomic study often depends on multiple experimental steps. The comprehensive analysis of complex samples still requires meticulous method development due to lack of analyte free matrix and experience on both LC and MS methods.

To address this challenge we developed a HILIC-MS/MS profiling method for the quantification of ca 100 metabolites of high significance in central biosynthetic routes. The proposed method provides the identification and the generation of definite quantitative data. The method has been optimised and fully validated to accurately quantify polar metabolites in a single injection of 40 min. The use of a single analytical station enhances assay/practitioner control, reporting and inter-laboratory testing.

The developed method was applied first to the analysis of IVF extracellular culture medium of IVF procedure and subsequently to other biological matrices. The results from the analysis provided information on the metabolic activity reflecting from embryos with different growth potential. Differences in the profiles of media deriving from chromosomally normal embryos that resulted in blastocyst and abnormal that stayed in arrested stage could be detected.  Multivariate statistical analysis showed that alanine, glucose, lactic acid and creatine revealed to be the most significant to the differentiation of embryos with different growth potential.

Additional results showed that the developed method can sensitively detect the metabolic content of different matrices and could be used as a tool for detecting differences in metabolic signature and for different purposes in life sciences (diagnostic, nutritional assessment and other). Although the development of such an analytical tool may require time, hard work and dedication it is in our opinion definitely worth the effort.

Journal Reference

Electrophoresis. 2015:Volume 36: issue 18:2215-2225.

Virgiliou C¹, Sampsonidis I¹, Gika HG², Raikos N³, Theodoridis GA¹.

Show Affiliations

1. Department of Chemistry, Aristotle University, Thessaloniki, 54124, Greece.
2. Department of Chemical Engineering, Aristotle University, Thessaloniki, 54124, Greece.
3. Department of Medicine, Aristotle University, Thessaloniki, 54124, Greece.

Abstract

The paper reports the development of a multi-analyte method and its application in metabolic profiling of biological fluids. The initial aim of the method was the quantification of metabolites existing in cell culture medium used in in-vitro fertilisation (IVF) and in other biological fluids related to embryo growth. Since most of these analytes are polar primary metabolites a hydrophilic interaction liquid chromatography (HILIC) system was selected. The analytical system comprised Ultra (High) performance liquid chromatography (UHPLC) with detection on a triple quadrupole mass spectrometer operating in both positive and negative modes. Mobile phase and gradient elution conditions were studied with the aim to achieve the highest coverage of metabolic space in a single injection namely the largest number of analytes that could be detected and quantified. The developed method provides absolute quantitation of ca. 100 metabolites belonging to key metabolite classes such as sugars, amino acids, nucleotides, organic acids and amines. Following validation, the method was applied for the metabolic profiling of hundreds of samples of spent culture medium originating from human IVF procedures and several hundreds of biological samples such as amniotic fluid, human urine and blood serum from pregnant women. The bioanalytical end-point was to provide assistance in the process of embryo transfer and improving IVF success rates but also to provide insight in complications related to the subsequent embryo growth during pregnancy. This article is protected by copyright. All rights reserved.

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