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Analytical Biochemistry

Jenny Wang, Daniel Stenzel, Ally Liu, Dengfeng Liu, Darren Brown, Alexandre Ambrogelly
Therapeutic vaccines represent an emerging class of immune-modulatory treatments for cancer, infections, and chronic diseases. One such vaccine was designed as an immune stimulator of the T cell response against HBV antigens to eliminate HBV infected cells and offer a therapeutic avenue to treat patients suffering from chronic hepatitis B infection. Whole deactivated Saccharomyces cerevisiae cells expressing a recombinant fusion of HBV X, S and Core antigens elicit T cell responses in mice and activate human T cells linked with viral clearance...
January 12, 2018: Analytical Biochemistry
Kamfai Chan, Pui-Yan Wong, Chaitanya Parikh, Season Wong
Traditionally, the majority of nucleic acid amplification-based molecular diagnostic tests are done in centralized settings. In recent years, point-of-care tests have been developed for use in low-resource settings away from central laboratories. While most experts agree that point-of-care molecular tests are greatly needed, their availability as cost-effective and easy-to-operate tests remains an unmet goal. In this article, we discuss our efforts to develop a recombinase polymerase amplification reaction-based test that will meet these criteria...
January 12, 2018: Analytical Biochemistry
Sajid Rauf, Geetesh K Mishra, Jahanzaib Azhar, Rupesh K Mishra, K Yugender Goud, Muhammad Azhar Hayat Nawaz, Jean Louis Marty, Akhtar Hayat
In this work, we have developed for the first time a carboxylic group riched graphene oxide based disposable electrochemical immunosensor for cancer biomarker detection using methylene blue (MB). The developed immunosensor is highly sensitive for detection of biomarker Mucin1 (MUC1) in human serum samples. Development of this disposable electrochemical immunosensor was premeditated by applying specific monoclonal antibodies against MUC1. In this method, we explored highly conductive surface of carboxylic group (-COOH-) rich graphene oxide (GO) on screen-printed carbon electrodes (SPCE)...
January 12, 2018: Analytical Biochemistry
T A Kurgina, R O Anarbaev, M V Sukhanova, O I Lavrik
Poly(ADP-ribose) polymerase 1 (PARP1) is a key enzyme that regulates important cellular processes, including DNA repair. PARP1 binds to a DNA damage site and synthesizes poly(ADP-ribose) chains (PARs), which serve as a signal of DNA damage for other DNA repair enzymes. PARP1 is a recognized target for the development of anti-cancer drugs. In this work, a method is developed that makes it possible to investigate the complex formation of PARP1 with DNA as well as its dissociation by detecting the fluorescence anisotropy of this complex during the poly(ADP-ribose) synthesis...
January 8, 2018: Analytical Biochemistry
Anthony Maus, Rudolph Mignon, Franco Basile
The use of offline liquid chromatography-matrix assisted laser desorption/ionization (LC-MALDI) tandem mass spectrometry (MS/MS) for bottom-up proteomics offers advantages in terms of cost, ease of use, and the time-decoupled nature of the separation step and the mass analysis. A method was developed to improve the capabilities of LC-MALDI-MS/MS in terms of protein identification in a bottom-up proteomic workflow. Enhanced protein identification is achieved by an increase in the MALDI signal intensity of the precursor peptides brought about by coating the MALDI plate with a thin film of graphite powder...
January 8, 2018: Analytical Biochemistry
Garik V Mkrtchyan, Anastasia Graf, Lidia Trofimova, Alexander Ksenofontov, Ludmila Baratova, Victoria Bunik
Glutamate is a key metabolite and major excitatory neurotransmitter, degraded through transformation to 2-oxoglutarate which is further catabolized by 2-oxoglutarate dehydrogenase complex (OGDHC). Both the glutamate excitotoxicity and impaired OGDHC activity are hallmarks of neurodegeneration. This work quantifies a relationship between the brain OGDHC activity and glutamate levels, assessing its diagnostic value to characterize (patho)physiology. A moderate to strong positive correlation of the two parameters determined under varied physiological settings (brain regions, seasons, gender, pregnancy, rat line), is revealed...
January 8, 2018: Analytical Biochemistry
Mayur Katariya, Dwayne Chung Kim Chung, Tristan Minife, Harshit Gupta, Alifa Afiah Ahmad Zahidi, Oi Wah Liew, Tuck Wah Ng
Autonomous systems for sample transport to the laboratory for analysis can be improved in terms of timeliness, cost and error mitigation in the pre-analytical testing phase. Drones have been reported for outdoor sample transport but incorporating devices on them to attain homogenous mixing of reagents during flight to enhance sample processing timeliness is limited by payload issues. It is shown here that flipping maneuvers conducted with quadcopters are able to facilitate complete and gentle mixing. This capability incorporated during automated sample transport serves to address an important factor contributing to pre-analytical variability which ultimately impacts on test result reliability...
January 4, 2018: Analytical Biochemistry
Luciana Assis Gobo, Leandro Machado de Carvalho, Fernanda Temp, Carine Viana, Carlos Fernando Mello
An analytical method utilizing liquid chromatography coupled to mass spectrometry with electrospray ionization has been developed for the identification of prostaglandins (PGs) in cerebral tissues. The five compounds identified (thromboxane B2, prostaglandin E2, prostaglandin D2, 6-keto-prostaglandin F1 alpha and prostaglandin F2 alpha) are cellular mediators of inflammation and are involved in a variety of physiological and pathological processes by acting on membrane receptors on the surfaces of target cells...
January 3, 2018: Analytical Biochemistry
I L G Law, J F C Loo, H C Kwok, H Y Yeung, C C H Leung, M Hui, S Y Wu, H S Chan, Y W Kwan, H P Ho, S K Kong
With the emergence of multi- and extensive-drug (MDR/XDR) resistant Mycobacterium tuberculosis (M. tb), tuberculosis (TB) persists as one of the world's leading causes of death. Recently, isothermal DNA amplification methods received much attention due to their ease of translation onto portable point-of-care (POC) devices for TB diagnosis. In this study, we aimed to devise a simple yet robust detection method for M. tb. Amongst the numerous up-and-coming isothermal techniques, Recombinase Polymerase Amplification (RPA) was chosen for a real-time detection of TB with or without MDR...
January 2, 2018: Analytical Biochemistry
Xiaoren Tang, Bushra Aljahdali, Mansour Alasiri, Abdullah Bamashmous, Feng Cao, Serge Dibart, Erdjan Salih
Adherent cells such as mouse RAW cells or human cancer U2OS cells are beneficial to DNA transfection, with 20%-60% transfection efficiency. However, this DNA transfection is rarely used on suspension cells due to its low transfection efficiency (≤5%). We recently found a new DNA transfection method to increase the efficiency up to 13.5% in suspension cells without PMA treatment. We also found that DNA transfection of human TNFAIP1 or CXCL1 recombinant plasmid DNA in THP-1 cells induces a high level of TNF-α protein...
January 2, 2018: Analytical Biochemistry
Dao Thi Thuy Nguyen, Eun Yeong Lee, Bonhan Koo, Choong Eun Jin, Tae Yoon Lee, Yong Shin
Rapid and sensitive detection of low amounts of pathogen in large samples is needed for early diagnosis and treatment of patients and surveillance of pathogen. In this study, we report a microfluidic platform for detection of low pathogen levels in a large sample volume that couples an Magainin 1 based microfluidic platform for pathogen enrichment and a recombinase polymerase amplification (RPA) sensor for simultaneous pathogenic DNA amplification and detection in a label-free and real-time manner. Magainin 1 is used as a pathogen enrichment agent with a herringbone microfluidic chip...
December 28, 2017: Analytical Biochemistry
Tsutomu Sugihara, Masayasu Mie, Eiry Kobatake
Nanoparticles are small size-controlled particles from 1 to 100 nm diameters and characterized by their structure, base material and functional units displayed on their surfaces. In this study, protein-based nanoparticles composed of a hydrophobic elastin-like peptide unit, a hydrophilic aspartic acid-rich peptide unit and displaying antibody binding domains on their surfaces, were designed and genetically synthesized. The constituent fusion proteins, termed ELP-D-C, were found to exist in monomeric form (ELP-D-C/monomer) at low temperature...
December 25, 2017: Analytical Biochemistry
Franziska Pögel Neé Steinicke, Imke Oltmann-Norden, Hermann Waetzig
Surface Plasmon Resonance Biosensors (SPR) are one of the most powerful tools to characterize protein binding, e.g. for drug discovery, like target identification, ligand fishing, assay development, lead selection and manufacturing quality control. However, there is increasing concern about its reproducibility in the light of the reproducibility crisis. Therefore an appropriate analytical instrument qualification (AIQ) is required for quality assurance of SPR instruments. AIQ is a prerequisite for analytical method validation and it is consisting of four parts, Design Qualification (DQ), Installation Qualification (IQ), Operational Qualification (OQ) and Performance Qualification (PQ)...
December 25, 2017: Analytical Biochemistry
Laura Trapiella-Alfonso, Sylvain Broussy, Wang-Qing Liu, Michel Vidal, Edouard Lecarpentier, Vassilis Tsatsaris, Nathalie Gagey-Eilstein
Angiogenesis and its involved proteins, particularly Vascular Endothelial Growth Factor (VEGF) family and VEGF receptors (VEGFRs), have been considered as a target of therapeutic interest for numerous inflammatory and vascular diseases. Acting on this biological process through interaction with VEGFs or VEGFRs has received considerable attention. Indeed, VEGFs and VEGFRs are currently targeted by drugs such as monoclonal antibodies. The feasibility of a therapeutic strategy based on blocking the VEGF/VEGFR interaction by using ligands "other-than-biologics" is also explored...
December 25, 2017: Analytical Biochemistry
Morten M C H van Schie, Kourosh Honarmand Ebrahimi, Wilfred R Hagen, Peter-Leon Hagedoorn
Recent developments in microfluidic and nanofluidic technologies have resulted in development of new chip-based microfluidic calorimeters with potential use in different fields. One application would be the accurate high-throughput measurement of enzyme activity. Calorimetry is a generic way to measure activity of enzymes, but unlike conventional calorimeters, chip-based calorimeters can be easily automated and implemented in high-throughput screening platforms. However, application of chip-based microfluidic calorimeters to measure enzyme activity has been limited due to problems associated with miniaturization such as incomplete mixing and a decrease in volumetric heat generated...
December 20, 2017: Analytical Biochemistry
Miki Tanaka-Okamoto, Ken Hanzawa, Mikio Mukai, Hidenori Takahashi, Masayuki Ohue, Yasuhide Miyamoto
We previously identified four glycan tumor marker candidates using a HPLC-based method. One candidate was sialyl Tn (STN), NeuAcα2-6-GalNAc. In this study, glycans were prepared from sera by hydrazine treatment followed by fluorescent labeling with aminopyridine. Pyridylaminated-STN levels of 147 gastric cancer, 85 pancreatic cancer and 10 cholangiocarcinoma patients together with 102 normal controls were accurately quantified using HPLC separation followed by selected reaction monitoring (SRM) assay, which used a stable isotope, tetradeuterium-labeled pyridylamino glycan as an internal standard...
December 19, 2017: Analytical Biochemistry
Jonas Kissenkötter, Sören Hansen, Susanne Böhlken-Fascher, George Ademowo, Oladapo Elijah Oyinloye, Adeleye Solomon Bakarey, Gerhard Dobler, Dennis Tappe, Pranav Patel, Claus-Peter Czerny, Ahmed Abd El Wahed
Rickettsioses are zoonotic vector-transmitted bacterial infections leading to flu-like symptoms and can progress to severe illness in humans. The gold standard for diagnosis of rickettsial infections is the indirect immunofluorescence assay, a serological method which is not suitable for pathogen identification during the acute phase of the disease. Therefore, several real-time PCR assays were developed. These assays are very sensitive, but require high-equipped laboratories and well-trained personnel. Hence, in this study, a rapid point-of-need detection method was developed to detect all Rickettsia species...
December 16, 2017: Analytical Biochemistry
Joanna Smajdor, Robert Piech, Martyna Ławrywianiec, Beata Paczosa-Bator
A voltammetric method for fast and sensitive estradiol determination using carbon black modified glassy carbon electrode (CBGC) is proposed. The use of carbon black as a modifying layer led to obtain low detection limit (9.2·10-8 mol L-1 for a preconcentration time of 60 s) and stability of registered signals (measured as RSD is 1.3%, n = 7, estradiol concentration 0.5·10-6 mol L-1). Cyclic voltammetry study revealed that in phosphate media estradiol suffers irreversible one-proton and one-electron oxidation process...
December 16, 2017: Analytical Biochemistry
Xiaohan Zhang, Huisheng Zhuang
Polycyclic musks (PCMs) in the aquatic environment have become an emerging environmental issue because of their potential risk. The most commonly used method for analysis of PCMs is gas chromatography-mass spectrometer (GC-MS) with different sample extractions, which are somewhat expensive to operate, laborious and complex. In this paper, a carbon nanotube-enhanced real time immuno-PCR was developed for ultrasensitive detection of AHTN in water for the first time. The SWCNTs were used to immobilize numerous amino-DNA and polyclonal antibody to form polyclonal antibody-CNTs-DNA conjugates, which were used as a signal-amplifier in the proposed immunoassay system...
December 16, 2017: Analytical Biochemistry
Louisa V Forbes, Anthony J Kettle
Myeloperoxidase, an abundant neutrophil enzyme, promotes oxidative damage during inflammation by generating hypohalous acids and free radicals. Currently, there are no selective drugs to inhibit its adverse activity. This short-coming is partly due to the lack of screening assays that mimic the complex enzymatic activities of myeloperoxidase in vivo. We have developed an assay for myeloperoxidase activity that includes its major physiological substrates - chloride, thiocyanate, tyrosine, and urate. The multi-substrate assay monitors bleaching of 5-thio-2-nitrobenzoic acid and measures total oxidant production when hydrogen peroxide activates the enzyme...
December 16, 2017: Analytical Biochemistry
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