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Analytical Biochemistry

Lucia Marri, Anita M Jansson, Caspar E Christensen, Ole Hindsgaul
Diacetyl (2,3-butanedione) is an important metabolic marker of several cancers, as well as an important off-flavour component produced during fermentation. As a small molecule in a complex mixture with many other analytes, existing methods for identification and quantitation of diacetyl invariably involves a chromatographic separation step followed by signal integration with an appropriate stoichiometric detector. Here we demonstrate that the chemical reaction of diacetyl with a 1,2-phenylenediamine derivative yields a chemical adduct, 1,4-quinoxaline which can be conjugated on BSA...
July 21, 2017: Analytical Biochemistry
Liyang Liu, Xia Wang, Juan Yang, Yan Bai
We present a highly selective and sensitive colorimetric method for the detection of selenocystine (SeCys) coexisting with other amino acids, especially cysteine (Cys) using the AuNPs. Firstly, Cys was oxidized to cystine (Cys-Cys) by dissolved oxygen under Cu(2+) catalysis in the pre-reaction, which eliminated the interference of Cys in the SeCys sensing process. Then SeCys induced the rapid aggregation of AuNPs through Au-Se bond and complex formation of Cu(2+)-SeCys in the colorimetric reaction, in which the color change of AuNPs from red to blue or purple with the naked eye detection or with a UV-vis spectrophotometric determination...
July 21, 2017: Analytical Biochemistry
Shiv K Sharma, Roger M Leblanc
Many industries are striving for the development of more reliable and robust β-galactosidase biosensors that exhibit high response rate, increased detection limit and enriched useful lifetime. In a newfangled technological atmosphere, a trivial advantage or disadvantage of the developed biosensor may escort to the survival and extinction of the industry. Several alternative strategies to immobilize β-galactosidase enzyme for their utilization in biosensors have been developed in recent years in the quest of maximum utility by controlling the defects seen in the previous biosensors...
July 20, 2017: Analytical Biochemistry
Shivcharan Prasad, Ipsita Roy
The use of enzymes in organic solvents has considerably widened their repertoire of applications. Such low water containing media also offer the possibility of carrying out enzymatic reactions at higher temperatures and enhancing reaction yields. The utility of such preparations is limited by the damage caused to the protein structure during freeze-drying. This work investigates the result of exposing the proteolytic enzyme subtilisin to high temperature in low water containing n-octane on its activity in aqueous and non-aqueous media...
July 18, 2017: Analytical Biochemistry
Mahmoud Roushani, Faezeh Shahdost-Fard, Azadeh Azadbakht
Based on the unique characteristics of the combination of fullerene and gold nanoparticles, we successfully designed a new and facile nanocomposite (Au@nano-C60) to fabricate an aptasensor for the ultra-sensitive and selective detection of TNT. The gold nanoparticles decorated fullerene onto a glassy carbon electrode was prepared using an electrochemical method by the in situ generation of Au nanoparticles onto the surface of the glassy carbon electrode modified with activated fullerene. Successively, the NH2-Apt as a receptor molecule of 2,4,6-Trinitrotoluen was covalently attached onto the modified electrode surface with the resultant nanocomposite...
July 17, 2017: Analytical Biochemistry
Kazhal Ghanbari, Mahmoud Roushani, Azadeh Azadbakht
In the present study, by using the aptamer proximity binding assay strategy, a novel electrochemical aptasensor is described for ultrasensitive detection of hepatitis C virus (HCV) core antigen. The immobilization surface is prepared by the modification of a glassy carbon electrode (GCE) with a graphene quantum dots (GQD). GQD were introduced as a novel and suitable substrate for aptamers through π-π stacking interactions, the richness of hydrophilic edges as well as hydrophobic plane in GQD which enhances the aptamer absorption on the electrode surface...
July 17, 2017: Analytical Biochemistry
Ben Thompson, Jerry Clifford, Mike Jenns, Andrew Smith, Ray Field, Kalpana Nayyar, David C James
Measurement of recombinant protein product titer critically underpins all biopharmaceutical manufacturing process development, as well as diverse research and discovery activity. Here, we describe a simple rapid (<2 min per 96 samples) 96-well microplate-based assay that enables high-throughput quantitation of recombinant immunoglobulin G and Fc-containing IgG derivatives in mammalian cell culture supernatant over a wide dynamic range of 2.5-80 mg/L, using microplate fluorescence polarization (FP) spectroscopy...
July 14, 2017: Analytical Biochemistry
Kezia Gomes de Oliveira, Juliane Cristina Borba, Alexandre Melo Bailão, Célia Maria de Almeida Soares, Emanuel Carrilho, Gabriela Rodrigues Mendes Duarte
In recent years, isothermal DNA amplification methods have emerged as an alternative in molecular diagnostics due to its ease of operation. The purpose of using isothermal amplification is to simplify the diagnostics work by i) eliminating heating cycles, ii) reducing equipment costs, and iii) providing rapid and accurate results in laboratories with limited resources. Here we show a simple and fast method for E. coli detection in disposable polyester-toner (PeT) microdevice. The amplification by LAMP of the malB gene from E...
July 14, 2017: Analytical Biochemistry
Yulin Yuan, Shanshan Li, Yewei Xue, Jintao Liang, Lijie Cui, Qingbo Li, Sufang Zhou, Yong Huang, Guiyin Li, Yongxiang Zhao
In this study, a Fe3O4@Au-based pseudo-homogeneous electrochemical immunosensor was prepared for detection of alpha fetoprotein (AFP), a well-known hepatocellular carcinoma biomarker. The primary antibody (Ab1) was immobilized on Fe3O4@Au NPs as the capture probe. Horseradish peroxidase (HRP) and secondary antibody (Ab2) were conjugated on gold nanoparticles (GNPs) through electrostatic adsorption to form signal-amplifying labels. In the presence of AFP, a sandwich immunocomplex was formed via specific recognition of antigen-antibody in a Fe3O4@Au-basedpseudo-homogeneousreaction system...
July 13, 2017: Analytical Biochemistry
Ljubava D Zorova, Vasily A Popkov, Egor Y Plotnikov, Denis N Silachev, Irina B Pevzner, Stanislovas S Jankauskas, Valentina A Babenko, Savva D Zorov, Anastasia V Balakireva, Magdalena Juhaszova, Steven J Sollott, Dmitry B Zorov
The mitochondrial membrane potential (ΔΨm) generated by proton pumps (Complexes I, III and IV) is an essential component in the process of energy storage during oxidative phosphorylation. Together with the proton gradient (ΔpH), ΔΨm forms the transmembrane potential of hydrogen ions which is harnessed to make ATP. The levels of ΔΨm and ATP in the cell are kept relatively stable although there are limited fluctuations of both these factors that can occur reflecting normal physiological activity. However, sustained changes in both factors may be deleterious...
July 12, 2017: Analytical Biochemistry
Kalpana Settu, Jen-Tsai Liu, Ching-Jung Chen, Jang-Zern Tsai
In this study, we developed a screen-printed carbon-graphene-based electrochemical biosensor for EN2 protein detection. The engrailed-2 (EN2) protein, a biomarker for prostate cancer, is known to be a strong binder to a specific DNA sequence (5'-TAATTA-3') to regulate transcription. To take advantage of this intrinsic property, aptamer probes with TAATTA sequence was immobilized onto the screen-printed carbon-graphene electrode surface via EDC-NHS coupling approach. Cyclic voltammetry (CV) of the electrochemical measurement technique was employed for the quantitative detection of EN2 protein...
July 11, 2017: Analytical Biochemistry
Zhe Ju, Jun-Zhe Cao
As one of important protein post-translational modifications, N-formylation has been reported to be involved in various biological processes. The accurate identification of N-formylation sites is crucial for understanding the underlying mechanisms of N-formylation. Since the traditional experimental methods are generally labor-intensive and expensive, it is important to develop computational methods to predict N-formylation sites. In this paper, a predictor named NformPred is proposed to improve the prediction of N-formylation sites by using composition of k-spaced amino acid pairs encoding scheme and support vector machine algorithm...
July 11, 2017: Analytical Biochemistry
C F Markwalter, I K Jang, R A Burton, G J Domingo, D W Wright
Predicting antibody pair performance in a sandwich format streamlines development of antibody-based diagnostics and laboratory research tools, such as enzyme-linked immunosorbent assays (ELISAs) and lateral flow immunoassays (LFAs). We have evaluated panels of monoclonal antibodies against the malarial parasite biomarker Plasmodium falciparum histidine rich protein 2 (HRP2), including 9 new monoclonal antibodies, using biolayer interferometry (BLI) and screened antibody pairs in a checkerboard ELISA. This study showed BLI predicts antibody pair ELISA performance for HRP2...
July 8, 2017: Analytical Biochemistry
Zhonghua Xue, Xiaofen Wang, Honghong Rao, Xiuhui Liu, Xiaoquan Lu
Many strategies have been explored for selectively and sensitively detecting cysteine in different samples. Here, a novel colorimetric sensor based on self-assembly nanostructures of Fe(3+)-H2O2/Tetramethylbenzidine system with dual-level logic gate function and colorimetric determination of cysteine were firstly explored. The proposed Fe(3+)-H2O2-TMB system provides a sensitive optical signal due to the selectively reductive ability of cysteine to the oxidized TMB and thus could be successfully applied to the construction of instant on-site visual detection method with a paper based test strip for cysteine determination in a sample solution as well as for a dual-level logic gate fabrication...
July 8, 2017: Analytical Biochemistry
Andrew Osahor, Karthik Deekonda, Choon-Weng Lee, Edmund Ui-Hang Sim, Aurelian Radu, Kumaran Narayanan
Sample preparation for scanning electron microscope analysis involves reagents and equipment that are expensive and often hazardous. Here we demonstrate a circumvention of Osmium tetroxide and critical point drying, greatly reducing the duration, complexity and cost of the process. We captured early stage interactions of invasive-bacteria and HeLa cells during the process of bacteria-mediated gene delivery and illustrate sufficient clarity can be obtained using this procedure to preserve and clearly visualize relevant cellular structures...
July 8, 2017: Analytical Biochemistry
Artyom Y Baev, Pia Elustondo, Alexander Negoda, Evgeny V Pavlov
Mitochondrial permeability transition (PT) is a phenomenon of an increase of the inner membrane permeability in response to an excessive matrix calcium accumulation. PTP is caused by the opening of the large weakly selective channel. Molecular composition and regulation of permeability transition pore (PTP) are not well understood. Here we used isolated mitochondria to investigate dependence of PTP activation on the osmotic pressure. We found that in low osmotic strength solution calcium-induced PTP is significantly inhibited...
July 7, 2017: Analytical Biochemistry
Renu Pandey, Daman Saluja
Efficient electrophoretic separation of isolated total RNA utilizes chemicals and agents to aid in nuclease free environment. However cost, extensive pre-run processing protocols as well as toxic byproducts limit the usage of such protocols. Moreover, these treatments affect the overall electrophoretic results by altering the conductivity of the running buffer and weaken the gel strength. We here provide a protocol for RNA visualization that obviates these shortcomings by preparation of agarose gel with hydrogen peroxide using the regular TAE buffer...
July 6, 2017: Analytical Biochemistry
Jun Wang, Jun Wang, Zhenzhong Zhang, Xiaona Zhang, Shaoguo Ru, YiFei Dong
Vitellogenin (Vtg) is a sensitive biomarker for environmental estrogens. In this study, an immunosensor for quantifying zebrafish Vtg was developed using the Octet system. First, Protein A sensors were immobilized with purified anti-lipovitellin (Lv) antibody that demonstrated specificity to Vtg. Then, antibody-coated biosensors were immersed into zebrafish Lv standards and diluted samples. The Octet system measured and recorded kinetic parameters between antigens and captured antibody within 5 min. Sample Vtg concentrations were automatically calculated by interpolating relative binding rates observed with each sample and the immobilized anti-Lv antibody into the developed standard curve...
July 6, 2017: Analytical Biochemistry
Zitao Zhong, Xin Yao, Xiaomei Gao, Li Jia
A rapid, convenient and efficient DNA extraction method with no need for toxic agents and centrifugation was reported. A polypropylene microfuge (MF) tube was used as the substrate to immobilize polydopamine (PDA). The prepared PDA-coated MF (PDA@MF) tube was used as a pH-responsive platform for rapid extraction of DNA based on pH-induced charge switch of amino and phenolic hydroxyl groups in PDA coating. The extraction procedure is simple and can be finished in 25 min. The PDA@MF tube was applied for extraction of genomic DNA from foodborne pathogens in milk...
July 6, 2017: Analytical Biochemistry
Arthur J L Cooper
No abstract text is available yet for this article.
July 4, 2017: Analytical Biochemistry
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