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Showing 1 - 200 of 3176 Journals sorted alphabetically
A Practical Logic of Cognitive Systems     Full-text available via subscription   (Followers: 8)
AASRI Procedia     Open Access   (Followers: 14)
Academic Pediatrics     Hybrid Journal   (Followers: 28, SJR: 1.402, h-index: 51)
Academic Radiology     Hybrid Journal   (Followers: 22, SJR: 1.008, h-index: 75)
Accident Analysis & Prevention     Partially Free   (Followers: 90, SJR: 1.109, h-index: 94)
Accounting Forum     Hybrid Journal   (Followers: 25, SJR: 0.612, h-index: 27)
Accounting, Organizations and Society     Hybrid Journal   (Followers: 33, SJR: 2.515, h-index: 90)
Achievements in the Life Sciences     Open Access   (Followers: 4)
Acta Anaesthesiologica Taiwanica     Open Access   (Followers: 6, SJR: 0.338, h-index: 19)
Acta Astronautica     Hybrid Journal   (Followers: 377, SJR: 0.726, h-index: 43)
Acta Automatica Sinica     Full-text available via subscription   (Followers: 2)
Acta Biomaterialia     Hybrid Journal   (Followers: 26, SJR: 2.02, h-index: 104)
Acta Colombiana de Cuidado Intensivo     Full-text available via subscription   (Followers: 1)
Acta de Investigación Psicológica     Open Access   (Followers: 3)
Acta Ecologica Sinica     Open Access   (Followers: 8, SJR: 0.172, h-index: 29)
Acta Haematologica Polonica     Free   (Followers: 1, SJR: 0.123, h-index: 8)
Acta Histochemica     Hybrid Journal   (Followers: 3, SJR: 0.604, h-index: 38)
Acta Materialia     Hybrid Journal   (Followers: 236, SJR: 3.683, h-index: 202)
Acta Mathematica Scientia     Full-text available via subscription   (Followers: 5, SJR: 0.615, h-index: 21)
Acta Mechanica Solida Sinica     Full-text available via subscription   (Followers: 9, SJR: 0.442, h-index: 21)
Acta Oecologica     Hybrid Journal   (Followers: 10, SJR: 0.915, h-index: 53)
Acta Otorrinolaringologica (English Edition)     Full-text available via subscription  
Acta Otorrinolaringológica Española     Full-text available via subscription   (Followers: 2, SJR: 0.311, h-index: 16)
Acta Pharmaceutica Sinica B     Open Access   (Followers: 1)
Acta Poética     Open Access   (Followers: 4)
Acta Psychologica     Hybrid Journal   (Followers: 25, SJR: 1.365, h-index: 73)
Acta Sociológica     Open Access  
Acta Tropica     Hybrid Journal   (Followers: 6, SJR: 1.059, h-index: 77)
Acta Urológica Portuguesa     Open Access  
Actas Dermo-Sifiliograficas     Full-text available via subscription   (Followers: 3)
Actas Dermo-Sifiliográficas (English Edition)     Full-text available via subscription   (Followers: 2)
Actas Urológicas Españolas     Full-text available via subscription   (Followers: 3, SJR: 0.383, h-index: 19)
Actas Urológicas Españolas (English Edition)     Full-text available via subscription   (Followers: 1)
Actualites Pharmaceutiques     Full-text available via subscription   (Followers: 5, SJR: 0.141, h-index: 3)
Actualites Pharmaceutiques Hospitalieres     Full-text available via subscription   (Followers: 3, SJR: 0.112, h-index: 2)
Acupuncture and Related Therapies     Hybrid Journal   (Followers: 6)
Acute Pain     Full-text available via subscription   (Followers: 14)
Ad Hoc Networks     Hybrid Journal   (Followers: 11, SJR: 0.967, h-index: 57)
Addictive Behaviors     Hybrid Journal   (Followers: 15, SJR: 1.514, h-index: 92)
Addictive Behaviors Reports     Open Access   (Followers: 7)
Additive Manufacturing     Hybrid Journal   (Followers: 9, SJR: 1.039, h-index: 5)
Additives for Polymers     Full-text available via subscription   (Followers: 22)
Advanced Cement Based Materials     Full-text available via subscription   (Followers: 3)
Advanced Drug Delivery Reviews     Hybrid Journal   (Followers: 129, SJR: 5.2, h-index: 222)
Advanced Engineering Informatics     Hybrid Journal   (Followers: 11, SJR: 1.265, h-index: 53)
Advanced Powder Technology     Hybrid Journal   (Followers: 16, SJR: 0.739, h-index: 33)
Advances in Accounting     Hybrid Journal   (Followers: 8, SJR: 0.299, h-index: 15)
Advances in Agronomy     Full-text available via subscription   (Followers: 12, SJR: 2.071, h-index: 82)
Advances in Anesthesia     Full-text available via subscription   (Followers: 27, SJR: 0.169, h-index: 4)
Advances in Antiviral Drug Design     Full-text available via subscription   (Followers: 2)
Advances in Applied Mathematics     Full-text available via subscription   (Followers: 10, SJR: 1.054, h-index: 35)
Advances in Applied Mechanics     Full-text available via subscription   (Followers: 10, SJR: 0.801, h-index: 26)
Advances in Applied Microbiology     Full-text available via subscription   (Followers: 22, SJR: 1.286, h-index: 49)
Advances In Atomic, Molecular, and Optical Physics     Full-text available via subscription   (Followers: 14, SJR: 3.31, h-index: 42)
Advances in Biological Regulation     Hybrid Journal   (Followers: 4, SJR: 2.277, h-index: 43)
Advances in Botanical Research     Full-text available via subscription   (Followers: 2, SJR: 0.619, h-index: 48)
Advances in Cancer Research     Full-text available via subscription   (Followers: 28, SJR: 2.215, h-index: 78)
Advances in Carbohydrate Chemistry and Biochemistry     Full-text available via subscription   (Followers: 7, SJR: 0.9, h-index: 30)
Advances in Catalysis     Full-text available via subscription   (Followers: 5, SJR: 2.139, h-index: 42)
Advances in Cell Aging and Gerontology     Full-text available via subscription   (Followers: 3)
Advances in Cellular and Molecular Biology of Membranes and Organelles     Full-text available via subscription   (Followers: 12)
Advances in Chemical Engineering     Full-text available via subscription   (Followers: 27, SJR: 0.183, h-index: 23)
Advances in Child Development and Behavior     Full-text available via subscription   (Followers: 10, SJR: 0.665, h-index: 29)
Advances in Chronic Kidney Disease     Full-text available via subscription   (Followers: 10, SJR: 1.268, h-index: 45)
Advances in Clinical Chemistry     Full-text available via subscription   (Followers: 28, SJR: 0.938, h-index: 33)
Advances in Colloid and Interface Science     Full-text available via subscription   (Followers: 19, SJR: 2.314, h-index: 130)
Advances in Computers     Full-text available via subscription   (Followers: 14, SJR: 0.223, h-index: 22)
Advances in Dermatology     Full-text available via subscription   (Followers: 14)
Advances in Developmental Biology     Full-text available via subscription   (Followers: 10)
Advances in Digestive Medicine     Open Access   (Followers: 8)
Advances in DNA Sequence-Specific Agents     Full-text available via subscription   (Followers: 5)
Advances in Drug Research     Full-text available via subscription   (Followers: 21)
Advances in Ecological Research     Full-text available via subscription   (Followers: 42, SJR: 3.25, h-index: 43)
Advances in Engineering Software     Hybrid Journal   (Followers: 27, SJR: 0.486, h-index: 10)
Advances in Experimental Biology     Full-text available via subscription   (Followers: 6)
Advances in Experimental Social Psychology     Full-text available via subscription   (Followers: 42, SJR: 5.465, h-index: 64)
Advances in Exploration Geophysics     Full-text available via subscription   (Followers: 1)
Advances in Fluorine Science     Full-text available via subscription   (Followers: 7)
Advances in Food and Nutrition Research     Full-text available via subscription   (Followers: 54, SJR: 0.674, h-index: 38)
Advances in Fuel Cells     Full-text available via subscription   (Followers: 15)
Advances in Genetics     Full-text available via subscription   (Followers: 14, SJR: 2.558, h-index: 54)
Advances in Genome Biology     Full-text available via subscription   (Followers: 7)
Advances in Geophysics     Full-text available via subscription   (Followers: 6, SJR: 2.325, h-index: 20)
Advances in Heat Transfer     Full-text available via subscription   (Followers: 21, SJR: 0.906, h-index: 24)
Advances in Heterocyclic Chemistry     Full-text available via subscription   (Followers: 9, SJR: 0.497, h-index: 31)
Advances in Human Factors/Ergonomics     Full-text available via subscription   (Followers: 23)
Advances in Imaging and Electron Physics     Full-text available via subscription   (Followers: 1, SJR: 0.396, h-index: 27)
Advances in Immunology     Full-text available via subscription   (Followers: 36, SJR: 4.152, h-index: 85)
Advances in Inorganic Chemistry     Full-text available via subscription   (Followers: 8, SJR: 1.132, h-index: 42)
Advances in Insect Physiology     Full-text available via subscription   (Followers: 2, SJR: 1.274, h-index: 27)
Advances in Integrative Medicine     Hybrid Journal   (Followers: 6)
Advances in Intl. Accounting     Full-text available via subscription   (Followers: 3)
Advances in Life Course Research     Hybrid Journal   (Followers: 8, SJR: 0.764, h-index: 15)
Advances in Lipobiology     Full-text available via subscription   (Followers: 1)
Advances in Magnetic and Optical Resonance     Full-text available via subscription   (Followers: 9)
Advances in Marine Biology     Full-text available via subscription   (Followers: 14, SJR: 1.645, h-index: 45)
Advances in Mathematics     Full-text available via subscription   (Followers: 10, SJR: 3.261, h-index: 65)
Advances in Medical Sciences     Hybrid Journal   (Followers: 6, SJR: 0.489, h-index: 25)
Advances in Medicinal Chemistry     Full-text available via subscription   (Followers: 5)
Advances in Microbial Physiology     Full-text available via subscription   (Followers: 4, SJR: 1.44, h-index: 51)
Advances in Molecular and Cell Biology     Full-text available via subscription   (Followers: 21)
Advances in Molecular and Cellular Endocrinology     Full-text available via subscription   (Followers: 8)
Advances in Molecular Toxicology     Full-text available via subscription   (Followers: 7, SJR: 0.324, h-index: 8)
Advances in Nanoporous Materials     Full-text available via subscription   (Followers: 3)
Advances in Oncobiology     Full-text available via subscription   (Followers: 1)
Advances in Organ Biology     Full-text available via subscription   (Followers: 1)
Advances in Organometallic Chemistry     Full-text available via subscription   (Followers: 15, SJR: 2.885, h-index: 45)
Advances in Parallel Computing     Full-text available via subscription   (Followers: 6, SJR: 0.148, h-index: 11)
Advances in Parasitology     Full-text available via subscription   (Followers: 5, SJR: 2.37, h-index: 73)
Advances in Pediatrics     Full-text available via subscription   (Followers: 24, SJR: 0.4, h-index: 28)
Advances in Pharmaceutical Sciences     Full-text available via subscription   (Followers: 10)
Advances in Pharmacology     Full-text available via subscription   (Followers: 15, SJR: 1.718, h-index: 58)
Advances in Physical Organic Chemistry     Full-text available via subscription   (Followers: 8, SJR: 0.384, h-index: 26)
Advances in Phytomedicine     Full-text available via subscription  
Advances in Planar Lipid Bilayers and Liposomes     Full-text available via subscription   (Followers: 3, SJR: 0.248, h-index: 11)
Advances in Plant Biochemistry and Molecular Biology     Full-text available via subscription   (Followers: 7)
Advances in Plant Pathology     Full-text available via subscription   (Followers: 5)
Advances in Porous Media     Full-text available via subscription   (Followers: 5)
Advances in Protein Chemistry     Full-text available via subscription   (Followers: 17)
Advances in Protein Chemistry and Structural Biology     Full-text available via subscription   (Followers: 18, SJR: 1.5, h-index: 62)
Advances in Psychology     Full-text available via subscription   (Followers: 59)
Advances in Quantum Chemistry     Full-text available via subscription   (Followers: 6, SJR: 0.478, h-index: 32)
Advances in Radiation Oncology     Open Access  
Advances in Small Animal Medicine and Surgery     Hybrid Journal   (Followers: 3, SJR: 0.1, h-index: 2)
Advances in Space Biology and Medicine     Full-text available via subscription   (Followers: 5)
Advances in Space Research     Full-text available via subscription   (Followers: 377, SJR: 0.606, h-index: 65)
Advances in Structural Biology     Full-text available via subscription   (Followers: 5)
Advances in Surgery     Full-text available via subscription   (Followers: 9, SJR: 0.823, h-index: 27)
Advances in the Study of Behavior     Full-text available via subscription   (Followers: 29, SJR: 1.321, h-index: 56)
Advances in Veterinary Medicine     Full-text available via subscription   (Followers: 17)
Advances in Veterinary Science and Comparative Medicine     Full-text available via subscription   (Followers: 13)
Advances in Virus Research     Full-text available via subscription   (Followers: 5, SJR: 1.878, h-index: 68)
Advances in Water Resources     Hybrid Journal   (Followers: 46, SJR: 2.408, h-index: 94)
Aeolian Research     Hybrid Journal   (Followers: 6, SJR: 0.973, h-index: 22)
Aerospace Science and Technology     Hybrid Journal   (Followers: 334, SJR: 0.816, h-index: 49)
AEU - Intl. J. of Electronics and Communications     Hybrid Journal   (Followers: 8, SJR: 0.318, h-index: 36)
African J. of Emergency Medicine     Open Access   (Followers: 6, SJR: 0.344, h-index: 6)
Ageing Research Reviews     Hybrid Journal   (Followers: 9, SJR: 3.289, h-index: 78)
Aggression and Violent Behavior     Hybrid Journal   (Followers: 429, SJR: 1.385, h-index: 72)
Agri Gene     Hybrid Journal  
Agricultural and Forest Meteorology     Hybrid Journal   (Followers: 15, SJR: 2.18, h-index: 116)
Agricultural Systems     Hybrid Journal   (Followers: 31, SJR: 1.275, h-index: 74)
Agricultural Water Management     Hybrid Journal   (Followers: 43, SJR: 1.546, h-index: 79)
Agriculture and Agricultural Science Procedia     Open Access   (Followers: 1)
Agriculture and Natural Resources     Open Access   (Followers: 2)
Agriculture, Ecosystems & Environment     Hybrid Journal   (Followers: 56, SJR: 1.879, h-index: 120)
Ain Shams Engineering J.     Open Access   (Followers: 5, SJR: 0.434, h-index: 14)
Air Medical J.     Hybrid Journal   (Followers: 5, SJR: 0.234, h-index: 18)
AKCE Intl. J. of Graphs and Combinatorics     Open Access   (SJR: 0.285, h-index: 3)
Alcohol     Hybrid Journal   (Followers: 11, SJR: 0.922, h-index: 66)
Alcoholism and Drug Addiction     Open Access   (Followers: 9)
Alergologia Polska : Polish J. of Allergology     Full-text available via subscription   (Followers: 1)
Alexandria Engineering J.     Open Access   (Followers: 1, SJR: 0.436, h-index: 12)
Alexandria J. of Medicine     Open Access   (Followers: 1)
Algal Research     Partially Free   (Followers: 9, SJR: 2.05, h-index: 20)
Alkaloids: Chemical and Biological Perspectives     Full-text available via subscription   (Followers: 2)
Allergologia et Immunopathologia     Full-text available via subscription   (Followers: 1, SJR: 0.46, h-index: 29)
Allergology Intl.     Open Access   (Followers: 5, SJR: 0.776, h-index: 35)
Alpha Omegan     Full-text available via subscription   (SJR: 0.121, h-index: 9)
ALTER - European J. of Disability Research / Revue Européenne de Recherche sur le Handicap     Full-text available via subscription   (Followers: 9, SJR: 0.158, h-index: 9)
Alzheimer's & Dementia     Hybrid Journal   (Followers: 48, SJR: 4.289, h-index: 64)
Alzheimer's & Dementia: Diagnosis, Assessment & Disease Monitoring     Open Access   (Followers: 4)
Alzheimer's & Dementia: Translational Research & Clinical Interventions     Open Access   (Followers: 4)
Ambulatory Pediatrics     Hybrid Journal   (Followers: 6)
American Heart J.     Hybrid Journal   (Followers: 50, SJR: 3.157, h-index: 153)
American J. of Cardiology     Hybrid Journal   (Followers: 50, SJR: 2.063, h-index: 186)
American J. of Emergency Medicine     Hybrid Journal   (Followers: 42, SJR: 0.574, h-index: 65)
American J. of Geriatric Pharmacotherapy     Full-text available via subscription   (Followers: 10, SJR: 1.091, h-index: 45)
American J. of Geriatric Psychiatry     Hybrid Journal   (Followers: 14, SJR: 1.653, h-index: 93)
American J. of Human Genetics     Hybrid Journal   (Followers: 31, SJR: 8.769, h-index: 256)
American J. of Infection Control     Hybrid Journal   (Followers: 26, SJR: 1.259, h-index: 81)
American J. of Kidney Diseases     Hybrid Journal   (Followers: 32, SJR: 2.313, h-index: 172)
American J. of Medicine     Hybrid Journal   (Followers: 42, SJR: 2.023, h-index: 189)
American J. of Medicine Supplements     Full-text available via subscription   (Followers: 3)
American J. of Obstetrics and Gynecology     Hybrid Journal   (Followers: 189, SJR: 2.255, h-index: 171)
American J. of Ophthalmology     Hybrid Journal   (Followers: 62, SJR: 2.803, h-index: 148)
American J. of Ophthalmology Case Reports     Open Access   (Followers: 6)
American J. of Orthodontics and Dentofacial Orthopedics     Full-text available via subscription   (Followers: 6, SJR: 1.249, h-index: 88)
American J. of Otolaryngology     Hybrid Journal   (Followers: 25, SJR: 0.59, h-index: 45)
American J. of Pathology     Hybrid Journal   (Followers: 27, SJR: 2.653, h-index: 228)
American J. of Preventive Medicine     Hybrid Journal   (Followers: 27, SJR: 2.764, h-index: 154)
American J. of Surgery     Hybrid Journal   (Followers: 36, SJR: 1.286, h-index: 125)
American J. of the Medical Sciences     Hybrid Journal   (Followers: 12, SJR: 0.653, h-index: 70)
Ampersand : An Intl. J. of General and Applied Linguistics     Open Access   (Followers: 6)
Anaerobe     Hybrid Journal   (Followers: 4, SJR: 1.066, h-index: 51)
Anaesthesia & Intensive Care Medicine     Full-text available via subscription   (Followers: 61, SJR: 0.124, h-index: 9)
Anaesthesia Critical Care & Pain Medicine     Full-text available via subscription   (Followers: 14)
Anales de Cirugia Vascular     Full-text available via subscription  
Anales de Pediatría     Full-text available via subscription   (Followers: 2, SJR: 0.209, h-index: 27)
Anales de Pediatría (English Edition)     Full-text available via subscription  
Anales de Pediatría Continuada     Full-text available via subscription   (SJR: 0.104, h-index: 3)
Analytic Methods in Accident Research     Hybrid Journal   (Followers: 4, SJR: 2.577, h-index: 7)
Analytica Chimica Acta     Hybrid Journal   (Followers: 39, SJR: 1.548, h-index: 152)
Analytical Biochemistry     Hybrid Journal   (Followers: 166, SJR: 0.725, h-index: 154)
Analytical Chemistry Research     Open Access   (Followers: 10, SJR: 0.18, h-index: 2)
Analytical Spectroscopy Library     Full-text available via subscription   (Followers: 11)
Anesthésie & Réanimation     Full-text available via subscription   (Followers: 1)
Anesthesiology Clinics     Full-text available via subscription   (Followers: 22, SJR: 0.421, h-index: 40)
Angiología     Full-text available via subscription   (SJR: 0.124, h-index: 9)
Angiologia e Cirurgia Vascular     Open Access   (Followers: 1)

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Journal Cover Analytical Biochemistry
  [SJR: 0.725]   [H-I: 154]   [166 followers]  Follow
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
   ISSN (Print) 0003-2697 - ISSN (Online) 1096-0309
   Published by Elsevier Homepage  [3176 journals]
  • Prediction of lysine glutarylation sites by maximum relevance minimum
           redundancy feature selection
    • Authors: Zhe Ju; Jian-Jun He
      Pages: 1 - 7
      Abstract: Publication date: 1 June 2018
      Source:Analytical Biochemistry, Volume 550
      Author(s): Zhe Ju, Jian-Jun He
      Lysine glutarylation is new type of protein acylation modification in both prokaryotes and eukaryotes. To better understand the molecular mechanism of glutarylation, it is important to identify glutarylated substrates and their corresponding glutarylation sites accurately. In this study, a novel bioinformatics tool named GlutPred is developed to predict glutarylation sites by using multiple feature extraction and maximum relevance minimum redundancy feature selection. On the one hand, amino acid factors, binary encoding, and the composition of k-spaced amino acid pairs features are incorporated to encode glutarylation sites. And the maximum relevance minimum redundancy method and the incremental feature selection algorithm are adopted to remove the redundant features. On the other hand, a biased support vector machine algorithm is used to handle the imbalanced problem in glutarylation sites training dataset. As illustrated by 10-fold cross-validation, the performance of GlutPred achieves a satisfactory performance with a Sensitivity of 64.80%, a Specificity of 76.60%, an Accuracy of 74.90% and a Matthew's correlation coefficient of 0.3194. Feature analysis shows that some k-spaced amino acid pair features play the most important roles in the prediction of glutarylation sites. The conclusions derived from this study might provide some clues for understanding the molecular mechanisms of glutarylation.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.04.005
      Issue No: Vol. 550 (2018)
  • Principal coordinate analysis assisted chromatographic analysis of
           bacterial cell wall collection: A robust classification approach
    • Authors: Keshav Kumar; Felipe Cava
      Pages: 8 - 14
      Abstract: Publication date: 1 June 2018
      Source:Analytical Biochemistry, Volume 550
      Author(s): Keshav Kumar, Felipe Cava
      In the present work, Principal coordinate analysis (PCoA) is introduced to develop a robust model to classify the chromatographic data sets of peptidoglycan sample. PcoA captures the heterogeneity present in the data sets by using the dissimilarity matrix as input. Thus, in principle, it can even capture the subtle differences in the bacterial peptidoglycan composition and can provide a more robust and fast approach for classifying the bacterial collection and identifying the novel cell wall targets for further biological and clinical studies. The utility of the proposed approach is successfully demonstrated by analysing the two different kind of bacterial collections. The first set comprised of peptidoglycan sample belonging to different subclasses of Alphaproteobacteria. Whereas, the second set that is relatively more intricate for the chemometric analysis consist of different wild type Vibrio Cholerae and its mutants having subtle differences in their peptidoglycan composition. The present work clearly proposes a useful approach that can classify the chromatographic data sets of chromatographic peptidoglycan samples having subtle differences. Furthermore, present work clearly suggest that PCoA can be a method of choice in any data analysis workflow.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.04.008
      Issue No: Vol. 550 (2018)
  • Determination of enantiomeric excess of some amino acids by second-order
           calibration of kinetic-fluorescence data
    • Authors: Azadeh Naghashian-Haghighi; Bahram Hemmateenejad; Mojtaba Shamsipur
      Pages: 15 - 26
      Abstract: Publication date: 1 June 2018
      Source:Analytical Biochemistry, Volume 550
      Author(s): Azadeh Naghashian-Haghighi, Bahram Hemmateenejad, Mojtaba Shamsipur
      In this investigation a new non-separative kinetic-spectroflourimetric method is proposed for the determination of lysine (lys), leucine (leu) and phenylalanine (phe) enantiomers as their o-phthaldialdehyde (OPA) derivatives in the presence of an optically active chiral thiol compound, 1-mercapto-2-propanol (MP). At ambient temperature and in the borate buffer media of pH 9.6, MP, OPA, as highly selective fluorogenic reagents, and amino acid (AA) enantiomers reacts with each other to yield two fluorescent diasteriomers of D and L-AA with maximum difference in fluorescence intensity at about 450 nm. To achieve information from the small spectral changes, the data are analyzed by Multivariate Curve Resolution Alternating Least Squares (MCR-ALS) method. Linear calibration curves are achieved to distinct D and L-lys, leu and phe in different mole ratios by applying appropriate constraints in MCR-ALS procedures. This is the first application of MCR-ALS in determination of enantiomeric excess (ee) using OPA/MP adduct as chiral reagent, which benefits from direct time dependent-fluorescence spectral analysis and does not require prior separation of chiral analytes. Both the cross-validated correlation coefficient (Q 2) and root mean squares error of prediction (RMSEP) indicated satisfactory prediction ability of this method.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.04.004
      Issue No: Vol. 550 (2018)
  • Development of Quenching-qPCR (Q-Q) assay for measuring absolute
           intracellular cleavage efficiency of ribozyme
    • Authors: Min Woo Kim; Gwanggyu Sun; Jung Hyuk Lee; Byung-gee Kim
      Pages: 27 - 33
      Abstract: Publication date: 1 June 2018
      Source:Analytical Biochemistry, Volume 550
      Author(s): Min Woo Kim, Gwanggyu Sun, Jung Hyuk Lee, Byung-gee Kim
      Ribozyme (Rz) is a very attractive RNA molecule in metabolic engineering and synthetic biology fields where RNA processing is required as a control unit or ON/OFF signal for its cleavage reaction. In order to use Rz for such RNA processing, Rz must have highly active and specific catalytic activity. However, current methods for assessing the intracellular activity of Rz have limitations such as difficulty in handling and inaccuracies in the evaluation of correct cleavage activity. In this paper, we proposed a simple method to accurately measure the “intracellular cleavage efficiency” of Rz. This method deactivates unwanted activity of Rz which may consistently occur after cell lysis using DNA quenching method, and calculates the cleavage efficiency by analyzing the cleaved fraction of mRNA by Rz from the total amount of mRNA containing Rz via quantitative real-time PCR (qPCR). The proposed method was applied to measure “intracellular cleavage efficiency” of sTRSV, a representative Rz, and its mutant, and their intracellular cleavage efficiencies were calculated as 89% and 93%, respectively.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.04.007
      Issue No: Vol. 550 (2018)
  • Mucin and carbon nanotube-based biosensor for detection of glucose in
           human plasma
    • Authors: Fausto N. Comba; Marcelo R. Romero; Fernando S. Garay; Ana M. Baruzzi
      Pages: 34 - 40
      Abstract: Publication date: 1 June 2018
      Source:Analytical Biochemistry, Volume 550
      Author(s): Fausto N. Comba, Marcelo R. Romero, Fernando S. Garay, Ana M. Baruzzi
      This work reports an amperometric enzyme-electrode prepared with glucose oxidase, which have been immobilized by a cross-linking step with glutaraldehyde in a mixture containing albumin and a novel carbon nanotubes-mucin composite (CNT-muc). The obtained hydrogel matrix was trapped between two polycarbonate membranes and then fixed at the surface of a Pt working electrode. The developed biosensor was optimized by evaluating different compositions and the analytical properties of an enzymatic matrix with CNT-muc. Then, the performance of the resulting enzymatic matrix was evaluated for direct glucose quantification in human blood plasma. The novel CNT-muc composite provided a sensitivity of 0.44 ± 0.01 mA M−1 and a response time of 28 ± 2 s. These values were respectively 20% higher and 40% shorter than those obtained with a sandwich-type biosensor prepared without CNT. Additionally, CNT-muc based biosensor exhibited more than 3 orders of magnitude of linear dynamic calibration range and a detection limit of 3 μM. The short-term and long-term stabilities of the biosensors were also examined and excellent results were obtained through successive experiments performed within the first 60 days from their preparation. Finally, the storage stability was remarkable during the first 300 days.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.04.006
      Issue No: Vol. 550 (2018)
  • A label free Ag+ sensing method via in situ formation of metal
           coordination polymer
    • Authors: Yan Lu; Lianjie Meng; Yan Gao; Dongli Liao; Yongxin Li; Yongxing Ai; Yuqin Ma; Cong Yu
      Pages: 21 - 25
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Yan Lu, Lianjie Meng, Yan Gao, Dongli Liao, Yongxin Li, Yongxing Ai, Yuqin Ma, Cong Yu
      Metal ions sensing play critical roles in environmental monitoring and in biology. In this assay, we report the development of a facile fluorometric method for the sensing of Ag+ ions via the in situ formation of metal coordination polymer, based on the selective interactions of GSH with Ag+. The formation of coordination polymer with net multiple negative charges in an aqueous buffer solution (Tris-HAc, pH 9.0) resulted in aggregation and fluorescence quenching of a cationic perylene probe. The difference in emission intensity spurred us to develop a new strategy for sensing Ag+ ions. The proposed Ag+ detection method is simple, convenient, selective and sensitive, and can be used for Ag+ detection in lake water samples.

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.005
      Issue No: Vol. 549 (2018)
  • Fluorescent microplate assay method for high-throughput detection of
           lipase transesterification activity
    • Authors: Jianyong Zheng; Wei Wei; Xing Lan; Yinjun Zhang; Zhao Wang
      Pages: 26 - 28
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Jianyong Zheng, Wei Wei, Xing Lan, Yinjun Zhang, Zhao Wang
      This study describes a sensitive and fluorescent microplate assay method to detect lipase transesterification activity. Lipase-catalyzed transesterification between butyryl 4-methyl umbelliferone (Bu-4-Mu) and methanol in tert-butanol was selected as the model reaction. The release of 4-methylumbelliferone (4-Mu) in the reaction was determined by detecting the fluorescence intensity at λ ex 330 nm and λ em 390 nm. Several lipases were used to investigate the accuracy and efficiency of the proposed method. Apparent Michaelis constant (Km) was calculated for transesterification between Bu-4-Mu and methanol by the lipases. The main advantages of the assay method include high sensitivity, inexpensive reagents, and simple detection process.

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.010
      Issue No: Vol. 549 (2018)
  • HPLC-UV assays for evaluation of inhibitors of mono and diamine oxidases
           using novel phenyltetrazolylalkanamine substrates
    • Authors: Kira Mergemeier; Matthias Lehr
      Pages: 29 - 38
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Kira Mergemeier, Matthias Lehr
      Recently, we have described an HPLC-UV assay for the evaluation of inhibitors of plasma amine oxidase (PAO) using 6-(5-phenyl-2H-tetrazol-2-yl)hexan-1-amine (4) as a new type of substrate. Now we studied, whether this compound or homologues of it can also function as substrate for related amine oxidases, namely diamine oxidase (DAO), monoamine oxidase A (MAO A) and monoamine oxidase B (MAO B). Among these substances, 4 was converted by DAO with the highest rate. The best substrate for MAO A and B was 4-(5-phenyl-2H-tetrazol-2-yl)butan-1-amine (2). To validate the new assays, the inhibition values of known enzyme inhibitors were determined and the data were compared with those obtained with the substrate benzylamine, which is often used in amine oxidase assays. For the DAO inhibitor 2-(4-phenylphenyl)acetohydrazide an about 10fold lower IC50-value against DAO was obtained when benzylamine was applied instead of 4, indicating that 4 binds to the enzyme with higher affinity than benzylamine. The IC50-values of clorgiline and selegiline against MAO A and B, respectively, also decreased (two- and 30fold) replacing 2 by benzylamine. The discrepancies largely disappeared, when the enzymes were pre-incubated with the inhibitors for 15 min. This can be explained with the covalent inhibition mechanism of the inhibitors.

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.007
      Issue No: Vol. 549 (2018)
  • Single step, direct fluorescence immunoassays based on metal enhanced
           fluorescence (MEF-FIA) applicable as micro plate-, array-, multiplexing-
           or point of care-format
    • Authors: G. Hawa; Linda Sonnleitner; A. Missbichler; A. Prinz; G. Bauer; C. Mauracher
      Pages: 39 - 44
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): G. Hawa, Linda Sonnleitner, A. Missbichler, A. Prinz, G. Bauer, C. Mauracher
      Although Enzyme Linked Immuno Sorbent Assay (ELISA) technology is approaching it's 45th year of existence since first described in 1971, it is still the main diagnostic tool in clinical research and routine diagnostics. However, despite its broad usage it suffers from some drawbacks, limiting its use especially in more advanced assay formats like multiplexing platforms, point of care devices or protein arrays. Those limitations result from the need for an enzyme label, a soluble enzyme substrate, washing steps (multiplexing, point care, arrays) and in some cases also insufficient sensitivity, because the majority of circulating proteins and thus potential biomarkers may be found in lower sub-picomolar concentrations. We hereby present a new assay platform based on metal enhanced fluorescence (MEF), that remedies these problems since it eliminates the need for washing steps, for using enzyme labels and allows detection of analytes down to sub-picomolar concentrations. In addition this technology is fully compatible to standard fluorescence reader equipment as it is found in many laboratories nowadays. Since our present work is focused on single biomarker evaluation, we chose a 96 well plate format for convenience, but any other formate like antibody arrays, strip-like point of care devices etc. is feasible too.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.002
      Issue No: Vol. 549 (2018)
  • An improved immobilized enzyme reactor-mass spectrometry-based label free
           assay for butyrylcholinesterase ligand screening
    • Authors: Adriana Ferreira Lopes Vilela; Cláudia Seidl; Juliana Maria Lima; Carmen Lúcia Cardoso
      Pages: 53 - 57
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Adriana Ferreira Lopes Vilela, Cláudia Seidl, Juliana Maria Lima, Carmen Lúcia Cardoso
      Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are key cholinesterase enzymes responsible for the hydrolysis of acetylcholine into choline and acetic acid, an essential process for the restoration of the cholinergic neuron. The loss of cholinergic function in the central nervous system contributes to the cognitive decline associated with advanced age and Alzheimer's disease (AD). Inhibitions assays represent a significant role in the drug discovery process. Herein, we describe an improved label free method to screen and characterize new BChE ligands. The liquid chromatography system uses an immobilized capillary enzyme reactor (ICER) as a low affinity and high selectivity column coupled to a mass spectrometer (MS). The enzyme activity was evaluated by monitoring the choline's precursor ion [M + H]+ m/z 104 for a brief period. The method was validated using two known cholinesterase inhibitors tacrine and galanthamine. The IC50 values were 0.03 ± 0.006 μM and 0.88 ± 0.2 for tacrine and galanthamine respectively, and Ki was 0.11 ± 0.2 for galanthamine. The efficient combination of the huBChE-ICER with sensitive enzymatic assay detection such as MS, improved the reliable, fast identification of new ligands. Moreover, specific direct quantitation of the product contributes to the reduction of false positive and negative results.
      Graphical abstract image

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.012
      Issue No: Vol. 549 (2018)
  • Spherical-supported membranes as platforms for screening against membrane
           protein targets
    • Authors: V. Vasilca; A. Sadeghpour; S. Rawson; L.E. Hawke; S.A. Baldwin; T. Wilkinson; D. Bannister; V.L.G. Postis; M. Rappolt; S.P. Muench; L.J.C. Jeuken
      Pages: 58 - 65
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): V. Vasilca, A. Sadeghpour, S. Rawson, L.E. Hawke, S.A. Baldwin, T. Wilkinson, D. Bannister, V.L.G. Postis, M. Rappolt, S.P. Muench, L.J.C. Jeuken
      Screening assays performed against membrane protein targets (e.g. phage display) are hampered by issues arising from protein expression and purification, protein stability in detergent solutions and epitope concealment by detergent micelles. Here, we have studied a fast and simple method to improve screening against membrane proteins: spherical-supported bilayer lipid membranes (“SSBLM”). SSBLMs can be quickly isolated via low-speed centrifugation and redispersed in liquid solutions while presenting the target protein in a native-like lipid environment. To provide proof-of-concept, SSBLMs embedding the polytopic bacterial nucleoside transporter NupC were assembled on 100- and 200 nm silica particles. To test specific binding of antibodies, NupC was tagged with a poly-histidine epitope in one of its central loops between two transmembrane helices. Fluorescent labelling, small angle X-ray scattering (SAXS) and cryo-electron microscopy (cryo-EM) were used to monitor formation of the SSBLMs. Specific binding of an anti-his antibody and a gold-nitrilotriacetic acid (NTA) conjugate probe was confirmed with ELISAs and cryo-EM. SSBLMs for screening could be made with purified and lipid reconstituted NupC, as well as crude bacterial membrane extracts. We conclude that SSBLMs are a promising new means of presenting membrane protein targets for (biomimetic) antibody screening in a native-like lipid environment.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.006
      Issue No: Vol. 549 (2018)
  • In vitro selection and characterization of single stranded DNA aptamers
           for luteolin: A possible recognition tool
    • Authors: Jinan Tuma Sabah; Razauden Mohamed Zulkifli; Shafinaz Shahir; Farediah Ahmed; Mohammed Rafiq Abdul Kadir; Zarita Zakaria
      Pages: 72 - 79
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Jinan Tuma Sabah, Razauden Mohamed Zulkifli, Shafinaz Shahir, Farediah Ahmed, Mohammed Rafiq Abdul Kadir, Zarita Zakaria
      Distinctive bioactivities possessed by luteolin (3′, 4′, 5, 7-tetrahydroxy-flavone) are advantageous for sundry practical applications. This paper reports the in vitro selection and characterization of single stranded-DNA (ssDNA) aptamers, specific for luteolin (LUT). 76-mer library containing 1015 randomized ssDNA were screened via systematic evolution of ligands by exponential enrichment (SELEX). The recovered ssDNA pool from the 8th round was amplified with unlabeled primers and cloned into PSTBlue-1 vector prior to sequencing. 22 of LUT-binding aptamer variants were further classified into one of the seven groups based on their N40 random sequence regions, wherein one representative from each group was characterized. The dissociation constant of aptamers designated as LUT#28, LUT#20 and LUT#3 was discerned to be 107, 214 and 109 nM, respectively with high binding affinity towards LUT. Prediction analysis of the secondary structure suggested discrete features with typical loop and stem motifs. Furthermore, LUT#3 displayed higher specificity with insignificant binding toward kaempferol and quercetin despite its structural and functional similarity compared to LUT#28 and LUT#20. Further LUT#3 can detect free luteolin within 0.2–1 mM in solution. It was suggested that LUT#3 aptamer were the most suitable for LUT recognition tool at laboratory scale based on the condition tested.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.004
      Issue No: Vol. 549 (2018)
  • A high-throughput pH-based colorimetric assay: application focus on
           alpha/beta hydrolases
    • Authors: Mariétou F. Paye; Harrison B. Rose; John M. Robbins; Diana A. Yunda; Seonggeon Cho; Andreas S. Bommarius
      Pages: 80 - 90
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Mariétou F. Paye, Harrison B. Rose, John M. Robbins, Diana A. Yunda, Seonggeon Cho, Andreas S. Bommarius
      Research involving α/β hydrolases, including α-amino acid ester hydrolase and cocaine esterase, has been limited by the lack of an online high throughput screening assay. The development of a high throughput screening assay capable of detecting α/β hydrolase activity toward specific substrates and/or chemical reactions (e.g., hydrolysis in lieu of amidase activity and/or synthesis instead of thioesterase activity) is of interest in a broad set of scientific questions and applications. Here we present a general framework for pH-based colorimetric assays, as well as the mathematical considerations necessary to estimate de novo the experimental response required to assign a ‘hit’ or a ‘miss,’ in the absence of experimental standard curves. This combination is valuable for screening the hydrolysis and synthesis activity of α/β hydrolases on a variety of substrates, and produces data comparable to the current standard technique involving High Performance Liquid Chromatography (HPLC). In contrast to HPLC, this assay enables screening experiments to be performed with greater efficiency.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.03.009
      Issue No: Vol. 549 (2018)
  • The potential of aptamers for cancer research
    • Authors: Zhizhi Zhou; Mingying Liu; Jiahuan Jiang
      Pages: 91 - 95
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Zhizhi Zhou, Mingying Liu, Jiahuan Jiang
      Aptamers are promising alternatives to antibodies and can be used as high affinity agents for the cancer detection and the targeted drug transportation. In this manuscript, we highlight the advantages of aptamers, such as high affinities, specificity and excellent chemical stabilities, which are likely to benefit for the diagnosis of cancer in its early stages and then achieve molecular-level treatment. Also, we discuss the challenges and problems in the current application of aptamers.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.008
      Issue No: Vol. 549 (2018)
  • Rapid quantification of tyrosine sulfation in therapeutic proteins
    • Authors: Iva Turyan; Ruth Frenkel; Zoran Sosic
      Pages: 96 - 98
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Iva Turyan, Ruth Frenkel, Zoran Sosic
      Protein tyrosine sulfation (Tyr-O-SO3) is a common post-translational modification (PTM), which is important for protein function. Absolute quantitation of Tyr-O-SO3 in recombinant therapeutic proteins has been challenging. We report here an MRM method used for absolute quantitation of Tyr-O-SO3 in the hydrolysate of a recombinant Fc-fusion protein. Quantitation is achieved by monitoring the sum of two transitions: the loss of carboxylic acid from tyrosine sulfate (major transition) and sulfate group from tyrosine sulfate sodium salt. The method exhibits a good sensitivity with a limit of quantitation of 1.4 ng/mL, linearity over three orders of magnitude, good repeatability, precision and accuracy.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.001
      Issue No: Vol. 549 (2018)
  • Producing standard damaged DNA samples by heating: pitfalls and
    • Authors: Paolo Fattorini; Giorgio Marrubini; Serena Bonin; Barbara Bertoglio; Pierangela Grignani; Elisa Recchia; Paola Pitacco; Francesca Procopio; Carolina Cantoni; Irena Zupanič Pajnič; Solange Sorçaburu-Cigliero; Carlo Previderè
      Pages: 107 - 112
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Paolo Fattorini, Giorgio Marrubini, Serena Bonin, Barbara Bertoglio, Pierangela Grignani, Elisa Recchia, Paola Pitacco, Francesca Procopio, Carolina Cantoni, Irena Zupanič Pajnič, Solange Sorçaburu-Cigliero, Carlo Previderè
      Heat-mediated hydrolysis of DNA is a simple and inexpensive method for producing damaged samples in vitro. Despite heat-mediated DNA hydrolysis is being widely used in forensic and clinical validation procedures, the lack of standardized procedures makes it impossible to compare the intra and inter-laboratory outcomes of the damaging treatments. In this work, a systematic approach to heat induced DNA hydrolysis was performed at 70 °C for 0–18 h to test the role both of the hydrolysis buffer and of the experimental conditions. Specifically, a trial DNA sample, resuspended in three different media (ultrapure water, 0.1% DEPC-water and, respectively, TE) was treated both in Eppendorf tubes (“Protocol P”) and in Eppendorf tubes provided with screwcaps (“Protocol S”). The results of these comparative tests were assessed by normalization of the qPCR results. DEPC-water increased the degradation of the samples up to about 100 times when compared to the ultrapure water. Conversely, the TE protected the DNA from degradation whose level was about 1700 times lower than in samples treated in ultrapure water. Even the employment of the “Protocol S” affected the level of degradation, by consistently increasing it (up to about 180 times in DEPC-water). Thus, this comparative approach showed that even seemingly apparently trivial and often underestimated parameters modify the degradation level up to 2–3 orders of magnitude. The chemical-physical reasons of these findings are discussed together with the role of potential factors such as enhanced reactivity of CO2, ROS, NOx and pressure, which are likely to be involved. Since the intra and inter-laboratory comparison of the outcomes of the hydrolytic procedure is the first step toward its standardization, the normalization of the qPCR data by the UV/qPCR ratio seems to be the simplest and most reliable way to allow this. Finally, the supplying (provided with the commercial qPCR kits) of a DNA sample whose degree of degradation is well documented could be helpful in ISO/IEC 17025 validation procedures and in proficiency testing.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.011
      Issue No: Vol. 549 (2018)
  • FIA-MS/MS determination of creatinine in urine samples undergoing
    • Authors: Helena Jurdáková; Renáta Górová; Gabriela Addová; Anna Šalingová; Ivan Ostrovský
      Pages: 113 - 118
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Helena Jurdáková, Renáta Górová, Gabriela Addová, Anna Šalingová, Ivan Ostrovský
      Flow injection analysis-tandem mass spectrometry has become widely used for analysis of many biomarkers in various biological matrices. To improve the sensitivity, the compounds are often determined as their butylesters. Since the concentration of urinary excreted compounds are generally reported after normalization to creatinine, the aim of this study was to investigate the possibility of creatinine determination in urine samples which underwent butylation. The impact of derivatization on urinary creatinine determination was investigated by measuring of underivatized and derivatized samples. The 10% creatine to creatinine conversion was observed during butylation, what above 700 μmol creatine/mmol creatinine caused significant creatinine overestimation. In that case, correction for creatine conversion rate was done. QC samples at six concentration levels were examined and precision and accuracy values fulfill the European Medicine Agency validation requirements. The elaborated method was applied for determination of creatinine in 41 real human urine samples. Determined creatinine concentrations were in the range of 0.27–22.3 mmol/L, linearity was confirmed within the concentration range of 0.27–31.7 mmol/L. Obtained results highly correlated with routinely used enzymatic assay for all tested samples and proposed method provide reliable determination of creatinine in butylated urine in a single run with butylesters of other analytes of interest.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.018
      Issue No: Vol. 549 (2018)
  • A simple and reproducible protocol of glass surface silanization for TIRF
           microscopy imaging
    • Authors: Michał Szkop; Beata Kliszcz; Andrzej A. Kasprzak
      Pages: 119 - 123
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Michał Szkop, Beata Kliszcz, Andrzej A. Kasprzak
      We describe a simple and reproducible protocol for the preparation of microscope glass slides for in vitro motility assays that use total internal reflection fluorescence microscopy. The developed method utilizes trimethylchlorosilane (TMCS) as a silanizing reagent, which in the presence of imidazole as a catalyst and under optimized conditions enables reproducible preparation of high-quality hydrophobic glass surfaces. This method presents a simplification and improvement in reproducibility over the commonly applied protocol utilizing dichlorodimethylsilane (DDS) as a silanizing agent. We demonstrate the applicability of the new method by performing the analysis of the interactions of a molecular motor, kinesin-1 with microtubules.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.020
      Issue No: Vol. 549 (2018)
  • Selection of specific aptamer against enrofloxacin and fabrication of
           graphene oxide based label-free fluorescent assay
    • Authors: Somayeh Dolati; Mohammad Ramezani; Maryam Sadat Nabavinia; Vahid Soheili; Khalil Abnous; Seyed Mohammad Taghdisi
      Pages: 124 - 129
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Somayeh Dolati, Mohammad Ramezani, Maryam Sadat Nabavinia, Vahid Soheili, Khalil Abnous, Seyed Mohammad Taghdisi
      Specific ssDNA aptamers for the antibiotic enrofloxacin (ENR) were isolated from an enriched nucleotide library by SELEX (Systematic Evolution of Ligands by EXponential enrichment) method with high binding affinity. After seven rounds, five aptamers were selected and identified. Apt58 with highest affinity and sensitivity (Kd = 14.19 nM) was employed to develop a label-free fluorescent biosensing approach based on aptamer, graphene oxide (GO) and native fluorescence of ENR for determination of ENR residue in raw milk samples. Under optimized experimental conditions, the linear range was from 5 nM to 250 nM and LOD was calculated to be 3.7 nM, and the recovery rate was between 94.1% and 108.5%. The integration of aptamer and GO in this bioassay provides a promising way for rapid, sensitive and cost-effective detection of ENR in real samples like raw milk.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.021
      Issue No: Vol. 549 (2018)
  • An aptasensor for staphylococcus aureus based on nicking enzyme
           amplification reaction and rolling circle amplification
    • Authors: Jingguo Xu; Jia Guo; Sarah Wanjiku Maina; Yumeng Yang; Yimin Hu; Xuanxuan Li; Jiarong Qiu; Zhihong Xin
      Pages: 136 - 142
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Jingguo Xu, Jia Guo, Sarah Wanjiku Maina, Yumeng Yang, Yimin Hu, Xuanxuan Li, Jiarong Qiu, Zhihong Xin
      An ultra-sensitive aptamer-based biosensor for the detection of staphylococcus aureus was established by adopting the nicking enzyme amplification reaction (NEAR) and the rolling circle amplification (RCA) technologies. Aptamer-probe (AP), containing an aptamer and a probe sequence, was developed to act as the recognition unit of the biosensor, which was specifically bound to S. aureus. The probe was released from AP and initiated into the subsequent DNA amplification reactions where S. aureus was present, converting the detection of S. aureus to the investigation of probe oligonucleotide. The RCA amplification products contained a G-quadruplex motif and formed a three dimensional structure in presence of hemin. The G4/hemin complex showed horseradish peroxidase (HRP)-mimic activity and catalyzed the chemiluminescence reaction of luminol mediated by H2O2. The results showed that the established biosensor could detect S. aureus specifically with a good linear correlation at 5–104 CFU/mL. The signal values based on NEAR-RCA two-step cycle were boosted acutely, much higher than that relied on one-cycle magnification. The limit of detection (LoD) was determined to be as low as 5 CFU/mL. The established aptasensor exhibited a good discrimination of living against dead S. aureus, and can be applied to detect S. aureus in the food industry.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.013
      Issue No: Vol. 549 (2018)
  • Characterization of antibody-C1q interactions by Biolayer Interferometry
    • Authors: Wei Zhou; Shanshan Lin; Rongying Chen; Jun Liu; Yali Li
      Pages: 143 - 148
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Wei Zhou, Shanshan Lin, Rongying Chen, Jun Liu, Yali Li
      IgG molecules exert important effector functions including complement-dependent cytotoxicity (CDC). Different IgG isotypes induce CDC effect with variation, largely due to their differential binding to C1q, the initiating molecule of the classical CDC pathway. Here we report a method to characterize the binding of IgG to C1q using label-free technique. With this method, we determined the binding affinities of multiple IgG1, IgG2 and IgG4 antibodies to C1q. To explore whether antigen binding to antibodies affects C1q binding, we assessed the binding of Trastuzumab and Adalimumab with bound antigen proteins to C1q. The results showed that although the two tested IgG1 mAbs alone bind C1q similarly, their FC binding to C1q was significantly impacted by antigen binding to the Fab. The data suggested that the first step of complement pathway, whether C1q binds target cell bound antibody molecules, may significantly affect the CDC activities of antibody drugs.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.022
      Issue No: Vol. 549 (2018)
  • Prediction of DNase I hypersensitive sites in plant genome using multiple
           modes of pseudo components
    • Authors: Shanxin Zhang; Weichao Zhuang; Zhenghong Xu
      Pages: 149 - 156
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Shanxin Zhang, Weichao Zhuang, Zhenghong Xu
      DNase I hypersensitive sites (DHSs) are accessible chromatin zones hypersensitive to DNase I endonucleases in plant genome. DHSs have been used as markers for the presence of transcriptional regulatory elements. It is an important complement to develop computational methods to identify DHSs for discovering potential regulatory elements. To the best of our knowledge, several machine learning approaches have been proposed for the DHSs prediction, but there is still room for improvements. In this work, a new predictor called pDHS-WE was proposed for prediction of DHSs in plant genome by using weighted ensemble learning framework. Here, five classes of heterogeneous features were used to represent the sequences. Five random forest (RF) operators were constructed based on these five classes of features. The proposed pDHS-WE was formed by fusing the five individual RF classifiers into an ensemble predictor. Genetic algorithm was employed to obtain the weights of different classes of features. In the experiments, pDHS-WE obtained accuracy of 88.5%, sensitivity of 89.1%, specificity of 88.0%, and AUC of 0.958, which was more than 2.7%, 2%, 3.5% and 2.6% higher than state-of-the-art methods, respectively. The results suggested that pDHS-WE may become a useful tool for transcriptional regulatory elements analysis in plant genome.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.025
      Issue No: Vol. 549 (2018)
  • Application of nanomaterials for the electrical and optical detection of
           the hepatitis B virus
    • Authors: Danielle A. Oliveira; Jussara V. Silva; José M.R. Flauzino; Ana C.H. Castro; Anna C.R. Moço; Márcia M.C.N. Soares; João M. Madurro; Ana G. Brito-Madurro
      Pages: 157 - 163
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Danielle A. Oliveira, Jussara V. Silva, José M.R. Flauzino, Ana C.H. Castro, Anna C.R. Moço, Márcia M.C.N. Soares, João M. Madurro, Ana G. Brito-Madurro
      This work describes different approaches for the detection of hepatitis B virus (HBV) genomic DNA, using electrochemical and optical techniques. The platforms consisted of a single-stranded DNA probe (HEPB1S), specific to HBV, grafted on a gold electrode modified with reduced graphene oxide or gold nanoparticles. Differential pulse voltammetry analysis indicates that the addition of HBV genomic DNA caused an increase of about 1.4 times in the current peak value, when compared to the negative control. It was observed a linear dependence with the log HBV-genomic DNA concentration and the electrochemical biosensor detected until 7.65 pg μL−1 of the target. Electrochemical impedance spectroscopy measurements showed an increase of about 2 times in the charge transfer resistance, after the addition of HBV genomic DNA. Assays using colloidal suspension of gold nanoparticles showed a shift of the peak wavelength, linearly proportional to the HBV-genomic DNA concentration, with a detection limit of 0.15 ng μL−1. The applicability of the gold nanoparticles for clinical samples was tested with success in the blood plasma. All the approaches used in this work were effective in detecting genomic DNA or blood plasma in positive samples for HBV.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.023
      Issue No: Vol. 549 (2018)
  • A fluorometric assay for lysosomal phospholipase A2 activity using
           fluorescence-labeled truncated oxidized phospholipid
    • Authors: Akira Abe; Miki Hiraoka; James A. Shayman; Hiroshi Ohguro
      Pages: 164 - 170
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Akira Abe, Miki Hiraoka, James A. Shayman, Hiroshi Ohguro
      Lysosomal phospholipase A2 (LPLA2) is a key enzyme involved in the homeostasis of cellular phospholipids. Recently, LPLA2 was reported to preferentially degrade some truncated oxidized phospholipids at the sn-1 position. A commercially available, truncated oxidized phospholipid conjugated with a fluorescent dye, 1-palmitoyl-2-glutaroyl-sn-glycero-3-phosphoethanolamine-N-[4-(dipyrrometheneboron difluoride) butanoyl] (PGPE-BODIPY), was used to develop a specific assay for this enzyme. When recombinant mouse LPLA2 was incubated with liposomes consisting of 1,2-O-octadecyl-sn-glycero-3-phosphocholine/PGPE-BODIPY under acidic conditions, PGPE-BODIPY was converted to palmitic acid and a polar BODIPY-product. After phase partitioning by chloroform/methanol, the polar BODIPY-product was recovered in the aqueous phase and identified as 1-lyso-PGPE-BODIPY. The formation of 1-lyso-PGPE-BODIPY was quantitatively determined by fluorescent measurements. The Km and Vmax values of the recombinant LPLA2 for PGPE-BODIPY were 5.64 μM and 20.7 μmol/min/mg protein, respectively. Detectable activity against PGPE-BODIPY was present in LPLA2 deficient mouse sera, but the deacylase activity was completely suppressed by treatment with 4-(2-aminoethyl)benzenesulfonyl fluoride (AEBSF). AEBSF had no effect on LPLA2 activity. The LPLA2 activity of mouse serum pre-treated with AEBSF was specifically and quantitatively determined by this assay method. The PGPE-BODIPY and AEBSF based LPLA2 assay is convenient and can be used to measure LPLA2 activity in a variety of biological specimens.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.024
      Issue No: Vol. 549 (2018)
  • The detection of a mismatched DNA by using hairpin DNA-templated silver
    • Authors: Seyeon Kim; Jongback Gang
      Pages: 171 - 173
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Seyeon Kim, Jongback Gang
      Fluorescence assays have been developed to detect biomolecules using DNA-templated silver nanoclusters (DNA-AgNCs) utilizing their unique physical and optical properties. This study was designed to detect single-mismatched DNA by the hybridization of target DNA to template DNA either before or after DNA-AgNCs synthesis. The results showed that the detection specificity of a single-mismatched DNA was clearly enhanced when the target DNA (cDNA) was hybridized to template DNA prior to DNA-AgNCs synthesis compared with cDNA hybridization subsequent to DNA-AgNCs synthesis.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.03.026
      Issue No: Vol. 549 (2018)
  • Quantification of proteins by data independent acquisition: Performance
           assessment of the Hi3 methodology
    • Authors: Guillaume Chevreux; Nolwenn Tilly; Nicolas Bihoreau
      Pages: 184 - 187
      Abstract: Publication date: 15 May 2018
      Source:Analytical Biochemistry, Volume 549
      Author(s): Guillaume Chevreux, Nolwenn Tilly, Nicolas Bihoreau
      Proteomics greatly benefited from the development of mass spectrometry. Over the last years, data-independent acquisitions increased in popularity in an effort to provide routine label free quantitative information. In this report, the performance of the Hi3 label free method was assessed based on the analysis of a plasma-derived protein mixture. The following parameters of the method (CVs) were determined: repeatability 13.8%, intermediate precision 27.6%, bias 32.3% and linearity observed over 3 orders of magnitude. Finally an accuracy of 42.5% corresponding to a confidence interval within 2 fold the expected protein abundance should be a good approximation of the method performance.

      PubDate: 2018-04-15T10:34:19Z
      DOI: 10.1016/j.ab.2018.03.019
      Issue No: Vol. 549 (2018)
  • Simultaneous detection of Mycoplasma pneumoniae IgG and IgM using
           dual-label time resolved fluoroimmunoassay
    • Authors: Yi Zhang; Xue Yang; Jun Qian; Xiaohong Gu; Jue Zhang; Jie Liu; Zhigang Hu
      Pages: 1 - 6
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Yi Zhang, Xue Yang, Jun Qian, Xiaohong Gu, Jue Zhang, Jie Liu, Zhigang Hu
      Anti-Mycoplasma pneumoniae (MP) IgM and IgG are useful serological markers for detection of MP infection. In this study, a simultaneous quantification of MP IgM and IgG was performed by time-resolved fluoroimmunoassay (TRFIA). The europium-labeled anti-human IgM and samarium-labeled anti-human IgG were used as tracers, and MP IgM and IgG were recognized in serum samples. After dissociating europium and samarium ions from the immune complex, their fluorescence intensity was recorded and used to calculate the concentrations. The linear range and sensitivity of detection were 2–5500 BU/mL and 0.5 BU/mL for IgM, and 1.5–1500 BU/mL and 0.2 BU/mL for IgG, respectively. The intra- and inter-assay coefficients of variation were 5.14% and 8.41% for IgM, and 5.44% and 8.76% for IgG, respectively. The recovery rate was 94.9–106.8% for IgM and 96.1–109.4% for IgG. The correlation rates of serum detection for 38 respiratory infected patients between dual-label TRFIA and ELISA were 0.9294 and 0.9366 for IgM and IgG, respectively. The coincidence rate between passive particle agglutination and TRFIA is 93.3%. Dual-label TRFIA is a sensitive and reliable technique for measuring MP IgM and IgG levels and could be useful for the early diagnosis of MP infection.

      PubDate: 2018-02-26T00:26:16Z
      DOI: 10.1016/j.ab.2018.02.015
      Issue No: Vol. 548 (2018)
  • An efficient screening method for purifying and crystallizing membrane
           proteins using modified clear-native PAGE
    • Authors: Nanao Suzuki; Yuuki Takamuku; Tomohiro Asakawa; Makoto Inai; Tomoya Hino; So Iwata; Toshiyuki Kan; Takeshi Murata
      Pages: 7 - 14
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Nanao Suzuki, Yuuki Takamuku, Tomohiro Asakawa, Makoto Inai, Tomoya Hino, So Iwata, Toshiyuki Kan, Takeshi Murata
      Membrane proteins, such as G-protein coupled receptors, control communication between cells and their environments and are indispensable for many cellular functions. Nevertheless, structural studies on membrane proteins lag behind those on water-soluble proteins, due to their low structural stability, making it difficult to obtain crystals for X-ray crystallography. Optimizing conditions to improve the stability of membrane proteins is essential for successful crystallization. However, the optimization usually requires large amounts of purified samples, and it is a time-consuming and trial-and-error process. Here, we report a rapid method for precrystallization screening of membrane proteins using Clear Native polyacrylamide gel electrophoresis (CN-PAGE) with the modified Coomassie Brilliant Blue G-250 (mCBB) stain that was reduced in sodium formate. A2A adenosine receptor (A2AAR) was selected as a target membrane protein, for which we previously obtained the crystal structure using an antibody, and was expressed as a red fluorescent protein fusion for in-gel fluorescence detection. The mCBB CN-PAGE method enabled the optimization of the solubilization, purification, and crystallization conditions of A2AAR using the solubilized membrane fraction expressing the protein without purification procedures. These data suggest the applicability of mCBB CN-PAGE technique to a wide variety of integral membrane proteins.

      PubDate: 2018-02-26T00:26:16Z
      DOI: 10.1016/j.ab.2018.02.007
      Issue No: Vol. 548 (2018)
  • Functional electrospun nanofibers-based electrochemiluminescence
           immunosensor for detection of the TSP53 using RuAg/SiO2NPs as signal
    • Authors: Xiaoying Wang; Yijie Wang; Meng Jiang; Yanqun Shan; Xin Jin; Miao Gong; Xiaoning Wang
      Pages: 15 - 22
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Xiaoying Wang, Yijie Wang, Meng Jiang, Yanqun Shan, Xin Jin, Miao Gong, Xiaoning Wang
      A functional electrospun nanofibers-based electrochemiluminescence (ECL) immunosensor for detection of the tumor suppressor protein p53 (TSP53) at trace level using core-shell nanocomposite as signal enhancers is fabricated. The gold nanoparticles (AuNPs) were decorated on the surface of the electrospun carbon-nanotubes (MWCNTs) doped chitosan (CTS) nanofibers (MWCNTs-CTS) by in-situ electrodeposition. The functional electrospun nanofibers (MWCNTs-CTS-AuNPs) was utilized as supporting scaffolds for TSP53 capture antibody (CAb) immobilization firstly. They can dramatically increase the amount and stability of CAb attachment, and efficiently enhance the sensitivity of detection. After a sandwich immunoreaction, TSP53 and Ru(bpy)3 2+/silver nanoparticles doped silica core-shell nanocomposite (RuAg/SiO2NPs)-labeled detection antibody (RuAg/SiO2NPs@DAb) captured onto the electrode surface. It was observed that the increase of response ECL signal was proportional to the TSP53 concentration in the range of 1 pg mL−1 to 1 ng mL−1. The detection limit was 0.5 pg mL−1, which is comparable or better than that in reported TSP53 assays. The immunosensor was successfully applied to determination of TSP53 in normal human cubital vein blood samples with good recovery, and the results are basically consistent with the TSP53 Kit (ELISA). Excellent sensitivity and selectivity make the developed ECL immunosensor a potential and simple tool for the detection of tumor biomarkers.
      Graphical abstract image

      PubDate: 2018-02-26T00:26:16Z
      DOI: 10.1016/j.ab.2018.02.006
      Issue No: Vol. 548 (2018)
  • A methodological approach for the thermal stability and stress exposure
           studies of a model antibody
    • Authors: Gaëlle Coussot; Clément Faye; Aurélie Le Postollec; Michel Dobrijevic
      Pages: 23 - 31
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Gaëlle Coussot, Clément Faye, Aurélie Le Postollec, Michel Dobrijevic
      The anti-horseradish peroxidase (HRP) antibody is conventionally used in immunohistochemistry. More recently, it has been used as the key element in a gold standard method to evaluate the functionality of antibody-based materials. However, few information are available about its melting temperature and its stability after exposition to laboratory stress conditions including freeze-drying and freeze-thawing cycles. The aim of this study was to evaluate the effects of these environmental constraints on the anti-HRP antibody in order to further use it as a reference in quality control and in the development of new antibody-based materials. In the developed method, the anti-HRP antibody is covalently immobilized onto a solid surface. After the direct recognition of its antigen HRP, the signal is proportional to the number of antibody active binding sites. The method was successfully utilized to accurately evaluate the anti-HRP antibody melting temperature (Tm was 73.5 ± 0.2 °C). The method is a rapid and reliable tool with minimal cost for studying the anti-HRP antibody stability to solvent stress, freeze-thawing cycles, and freeze-drying process. The obtained information may be useful for routine analysis or in the development of antibody-based materials. This can be also proposed as an easy way to control antibody freeze-drying process.
      Graphical abstract image

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.02.019
      Issue No: Vol. 548 (2018)
  • A highly sensitive electrochemical sensor for the determination of
           methanol based on PdNPs@SBA-15-PrEn modified electrode
    • Authors: Ziba Karimi; Mojtaba Shamsipur; Mahmoud Amouzadeh Tabrizi; Sadegh Rostamnia
      Pages: 32 - 37
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Ziba Karimi, Mojtaba Shamsipur, Mahmoud Amouzadeh Tabrizi, Sadegh Rostamnia
      In this study, a novel electrochemical sensor for the determination of methanol based on palladium nanoparticles supported on Santa barbara amorphous-15- PrNHEtNH2 (PdNPs@SBA-15-PrEn) as nanocatalysis platform is presented. Scanning electron microscopy (SEM), transmission electron microscopy (TEM), X-ray diffraction (XRD) and electrochemical methods are employed to characterize the PdNPs@SBA-15-PrEn nanocomposite. The Nafion-Pd@SBA-15-PrEn modified glassy carbon electrode (Nafion-PdNPs@SBA-15-PrEn/GCE) displayed the high electrochemical activity and excellent catalytic characteristic for electro-oxidation of methanol in an alkaline solution. The electro-oxidation performance of the proposed sensor was investigated using cyclic voltammetry (CV) and amperometry. The sensor exhibits a good sensitivity of 0.0905 Amol−1 Lcm−2, linear range of 20–1000 μM and the corresponding detection limit of 12 μM (3σ). The results demonstrate that the Nafion-PdNPs@SBA-15-PrEn/GCE has potential as an efficient and integrated sensor for methanol detection.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.01.033
      Issue No: Vol. 548 (2018)
  • Quack: A quality assurance tool for high throughput sequence data
    • Authors: Adam Thrash; Mark Arick; Daniel G. Peterson
      Pages: 38 - 43
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Adam Thrash, Mark Arick, Daniel G. Peterson
      The quality of data generated by high-throughput DNA sequencing tools must be rapidly assessed in order to determine how useful the data may be in making biological discoveries; higher quality data leads to more confident results and conclusions. Due to the ever-increasing size of data sets and the importance of rapid quality assessment, tools that analyze sequencing data should quickly produce easily interpretable graphics. Quack addresses these issues by generating information-dense visualizations from FASTQ files at a speed far surpassing other publicly available quality assurance tools in a manner independent of sequencing technology.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.01.028
      Issue No: Vol. 548 (2018)
  • An electrochemical immunosensor based on poly p-phenylenediamine and
           graphene nanocomposite for detection of neuron-specific enolase via
           electrochemically amplified detection
    • Authors: Jafar Amani; Mehdi Maleki; Alireza Khoshroo; Ali Sobhani-Nasab; Mehdi Rahimi-Nasrabadi
      Pages: 53 - 59
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Jafar Amani, Mehdi Maleki, Alireza Khoshroo, Ali Sobhani-Nasab, Mehdi Rahimi-Nasrabadi
      In this work, a label-free electrochemical immunosensor was constructed on the base of poly p-phenylenediamine (PPD) and GR nanocomposite (PPD-GR). Screen-printed electrodes modified with PPD-GR nanocomposite and applied to advance enzyme-free and label free electrochemical immunosensor for detection of protein biomarker neuron-specific enolase (NSE). It was found that the PPD-GR nanocomposite exhibits excellent electrocatalytic activity towards ascorbic acid (AA) oxidation as analytical signal based on EC' mechanism. Due to the excellent electrocatalytic activity of PPD-GR nanocomposite, determination of NSE antigen was based on its obstruction to the electrocatalytic oxidation of AA after binding to the surface of electrode through interaction with the anti-NSE. The proposed immunosensor exhibited a wide linear range of 1.0–1000 ng mL−1, with a low detection limit of 0.3 ng mL−1. Furthermore, the proposed immunosensor were successfully used for the determination of NSE antigen in human serum samples.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.02.024
      Issue No: Vol. 548 (2018)
  • Multi-laboratory analysis of the variability of shipped samples for
           proteomics following non-cooled international transport
    • Authors: Pascal Steffen; Christoph Krisp; Wang Yi; Pengyuan Yang; Mark P. Molloy; Hartmut Schlüter
      Pages: 60 - 65
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Pascal Steffen, Christoph Krisp, Wang Yi, Pengyuan Yang, Mark P. Molloy, Hartmut Schlüter
      Transporting biological samples such as cells or tissues is complicated by the need to maintain integrity and minimise modification and degradation, but this is economically costly as the samples must be shipped in a frozen state. This multi-laboratory study investigated sample variability introduced by non-cooled transport of dried peptide samples for proteomic analysis using mass spectrometry. Human cancer cell tryptic lysates were proteolysed and dried in Australia and shipped by air to Europe and China. Samples were measured using label free mass spectrometry on similar LC-MS systems at all three sites. Preparation and analysis of the specimens in this manner resulted in only minor differences in protein identification and showed high quantitative reproducibility amongst the participating laboratories. We examined any impact on peptide chemical modification and report no discrepancies compared to the starting, non-shipped sample. We conclude that transport of non-cooled, dried peptides has negligible effect on sample integrity for downstream LC-MS analysis and therefore represents a cost-effective option to facilitate international proteomic collaborations. Data is available via ProteomeXchange with identifier PXD008160.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.02.026
      Issue No: Vol. 548 (2018)
  • Microparticle−based platform for point-of-care immunoassays
    • Authors: Teppo Salminen; Etvi Juntunen; Merja Lahdenranta; Iida Martiskainen; Sheikh M. Talha; Kim Pettersson
      Pages: 66 - 68
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Teppo Salminen, Etvi Juntunen, Merja Lahdenranta, Iida Martiskainen, Sheikh M. Talha, Kim Pettersson
      There is a need for quantitative and sensitive, yet simple point-of-care immunoassays. We have developed a point-of-care microparticle-based immunoassay platform which combines the performance of a microtiter well-based assay with the usability of a rapid assay. The platform contained a separate reaction and detection chambers and microparticles for the solid-phase. Photoluminescent up-converting nanoparticles (UCNPs) were used as labels. The platform was tested with a cardiac troponin I assay, and a limit of detection of 19.7 ng/L was obtained. This study demonstrates the feasibility of developing point-of-care assays on the new platform for various analytes of interests.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.021
      Issue No: Vol. 548 (2018)
  • Manufacturing of a novel double-function ssDNA aptamer for sensitive
           diagnosis and efficient neutralization of SEA
    • Authors: Hamid Sedighian; Raheleh Halabian; Jafar Amani; Mohammad Heiat; Ramezan Ali Taheri; Abbas Ali Imani Fooladi
      Pages: 69 - 77
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Hamid Sedighian, Raheleh Halabian, Jafar Amani, Mohammad Heiat, Ramezan Ali Taheri, Abbas Ali Imani Fooladi
      Staphylococcal enterotoxin A (SEA) is an enterotoxin produced mainly by Staphylococcus aureus. In recent years, it has become the most prevalent compound for staphylococcal food poisoning (SFP) around the world. In this study, we isolate new dual-function single-stranded DNA (ssDNA) aptamers by using some new methods, such as the Taguchi method, by focusing on the detection and neutralization of SEA enterotoxin in food and clinical samples. For the asymmetric polymerase chain reaction (PCR) optimization of each round of systematic evolution of ligands by exponential enrichment (SELEX), we use Taguchi L9 orthogonal arrays, and the aptamer mobility shift assay (AMSA) is used for initial evaluation of the protein-DNA interactions on the last SELEX round. In our investigation the dissociation constant (K D ) value and the limit of detection (LOD) of the candidate aptamer were found to be 8.5 ± 0.91 of nM and 5 ng/ml using surface plasmon resonance (SPR). In the current study, the Taguchi and mobility shift assay methods were innovatively harnessed to improve the selection process and evaluate the protein–aptamer interactions. To the best of our knowledge, this is the first report on employing these two methods in aptamer technology especially against bacterial toxin.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.017
      Issue No: Vol. 548 (2018)
  • Enzymatic quantification and length determination of polyphosphate down to
           a chain length of two
    • Authors: Jonas Johannes Christ; Lars Mathias Blank
      Pages: 82 - 90
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Jonas Johannes Christ, Lars Mathias Blank
      Polyacrylamide gel electrophoresis, being the current method of choice for length determination of inorganic polyphosphate (polyP), requires a sequencing apparatus, relies on commercially not available polyP length standards and yields only a chain length distribution. State of the art polyP quantification involves enzymatic hydrolysis of polyP to orthophosphate with the Saccharomyces cerevisiae exopolyphosphatase 1 (scPpx1p) and subsequent colorimetric orthophosphate detection. Because scPpx1p leaves one pyrophosphate per polyP, short chain polyPs are only partially detected. To overcome this analytical limitation, a method involving both the scPpx1p and the S. cerevisiae inorganic pyrophosphatase (scIpp1p) is proposed. Differential enzymatic hydrolysis of polyP with scPpx1p, and a combination of scIpp1p and scPpx1p allows not only for comprehensive quantification of polyP (excluding cyclic polyP) down to a chain length of two, but also absolute average chain length determination in the range of two to approximately 80. An optimized one-reagent method for rapid (2 min) orthophosphate quantification is part of the assay. Biological phosphorous containing molecules at equimolar phosphorous concentrations regarding polyP do not interfere. The method requires 1.5 μg polyP and calls only for a plate reader. This is the first enzymatic method for simultaneous average polyP chain length determination as well as comprehensive quantification.
      Graphical abstract image

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.018
      Issue No: Vol. 548 (2018)
  • Lectin-carbohydrate complex evaluation by chemiluminescence
    • Authors: Luiza Rayanna Amorim de Lima; Lúcia Patrícia Bezerra Gomes da Silva; Sinara Mônica Vitalino de Almeida; Thiago Barbosa Cahú; Eduardo Isidoro Carneiro Beltrão; Luiz Bezerra de Carvalho Júnior
      Pages: 91 - 95
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Luiza Rayanna Amorim de Lima, Lúcia Patrícia Bezerra Gomes da Silva, Sinara Mônica Vitalino de Almeida, Thiago Barbosa Cahú, Eduardo Isidoro Carneiro Beltrão, Luiz Bezerra de Carvalho Júnior
      In order to characterize the affinity between specific carbohydrate-binding proteins such as lectins, a model is proposed to study these interactions using a polysaccharide membrane to simulate such adsorption. Here, lectin-carbohydrate interactions were chemiluminescently investigated using lectins conjugated to acridinium ester (AE) and polysaccharides composed of their respective specific carbohydrates. The lectin-AE conjugates were incubated with discs (0.0314–0.6358 cm2) of phytagel, chitosan and carrageenan. The complex formation chemiluminescently detected followed the Langmuir isotherm from which constants were estimated. The association constant (Ka) and maximum binding sites on the membranes were 2.4 × 10−7 M−1 ± 0.8 × 10−7 M−1 and 1.3 × 10−3 mol. mg−1 ± 0.3 × 10−3 mol. mg−1 (Con A); 0.9 × 10−6 M−1 ± 0.4 × 10−6 M−1 and 0.021 × 10−3 mol. mg−1 ± 0.003 × 10−3 mol. mg−1 (WGA) and 2.0 × 10−6 M−1 ± 0.9 × 10−6 M−1 and 0.069 × 10−3 mol. mg−1 ± 0.010 × 10−3 mol. mg−1 (PNA). The proposed model might be useful to study binding affinity and estimate the amount of binding not limited by the sugar content in the membrane.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.01.024
      Issue No: Vol. 548 (2018)
  • Label-free and simple detection of endotoxins using a sensitive LSPR
           biosensor based on silver nanocolumns
    • Authors: Mohamad Zandieh; Seyed Nezamedin Hosseini; Manouchehr Vossoughi; Maryam Khatami; Sara Abbasian; Ahmad Moshaii
      Pages: 96 - 101
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Mohamad Zandieh, Seyed Nezamedin Hosseini, Manouchehr Vossoughi, Maryam Khatami, Sara Abbasian, Ahmad Moshaii
      This paper describes the construction of a silver-based LSPR biosensor for endotoxin detection. We used GLAD method to procure reproducible silver nanocolumns. In this work, the silver nanostructures were considerably stabilized by a SAM of MPA, and the limit of detection of biosensor was measured to be 340 pg/ml for endotoxin E. coli. Considering endotoxin B. abortus as the second type of endotoxin contamination in our target samples (HBs-ag produced in Institute Pasteur, Iran), we investigated selectivity of the biosensor in various experiments. We showed that this biosensor can selectively detect both types of endotoxins compared to other biological species. Overall, this study proposes that LSPR biosensing can be considered as a sensitive, simple, and label-free method for endotoxin detection in the quality control laboratories.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.023
      Issue No: Vol. 548 (2018)
  • Ultrasensitive environmental assessment of xeno-estrogens in water samples
           using label-free graphene immunosensors
    • Authors: Huw Barton; Waldir M. Berbel-Filho; Sofia Consuegra; Lewis Francis; Chedly Tizaoui; R. Steven Conlan; Sofia Rodrigues Teixeira
      Pages: 102 - 108
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Huw Barton, Waldir M. Berbel-Filho, Sofia Consuegra, Lewis Francis, Chedly Tizaoui, R. Steven Conlan, Sofia Rodrigues Teixeira
      There is a growing interest in the possible environmental health impact posed by endocrine-disrupting chemicals (EDCs). A challenge to the field of endocrine disruption is that these substances are diverse and may not appear to share any structural similarity other than usually being low molecular mass (<1000 Da) compounds. Here we demonstrate the effectiveness of sensor device for the detection of low molecular weight, poorly water soluble, estrogenic compounds E1, E2 and EE2, fabricated by electropolymerization over graphene screen printed electrode (SPE). The PANI/Gr-SPE-devices displayed linear responses to estrogenic substances, in EIS assays, from 0.0975 ng/L to 200 ng/L in water samples, with a detection limit of 0.043 pg/L for E1, 0.19 ng/L for E2 and 0.070 pg/L for EE2 which is lower than other current biosensing techniques. This portable, disposable immunosensor offers a solution for immediate measurement at sample collection sites, due to its excellent sensitivity and selectivity when testing water samples obtained directly from rivers and waste water treatment facilities. The simple screen printing production method will enable the low cost, high volume production required for this type of environmental analysis.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.027
      Issue No: Vol. 548 (2018)
  • Developing a colorimetric assay for Fe(II)/2-oxoglutarate-dependent
    • Authors: Cuixia Guo; Yiling Hu; Chunyu Yang; Ankanahalli N. Nanjaraj Urs; Yan Zhang
      Pages: 109 - 114
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Cuixia Guo, Yiling Hu, Chunyu Yang, Ankanahalli N. Nanjaraj Urs, Yan Zhang
      The Fe(II)/2-oxoglutarate-dependent dioxygenases (2-OGDs) catalyze the oxidation of substrates ranging from small molecules to large biomolecules with concomitant oxidation of co-substrate (2-oxoglutarate) into succinate. In the present study, we reported a coupled colorimetric assay that can be generally applied to measure the activities of all members of 2-OGDs family. Succinyl-CoA synthetase is employed as the coupling enzyme to transform succinate produced from 2-OGDs catalysis to form succinyl-CoA with concomitant hydrolysis of ATP to form ADP and orthophosphate. Orthophosphate can be quantitated by reacting it with molybdic acid forming a blue pigment. As a proof of concept, kinetic parameters of ectoine hydroxylase obtained using this method are compared to a traditional time- and labor-consuming HPLC based method. As 2-OGDs family enzymes are important drug targets due to their impressive versatility in catalyzing numerous oxidative reactions that are still very challenging using synthetic chemistry, colorimetric method detailed in the manuscript has the potential to enable the practice of high throughput drug screening for 2-OGDs.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.013
      Issue No: Vol. 548 (2018)
  • Complete solubilization of cartilage using the heat-stable protease
           thermolysin for comprehensive GAG analysis
    • Authors: Harumi Osago; Mikiko Kobayashi-Miura; Yoshifumi Hamasaki; Nobumasa Hara; Mineyoshi Hiyoshi; Mikako Tsuchiya
      Pages: 115 - 118
      Abstract: Publication date: 1 May 2018
      Source:Analytical Biochemistry, Volume 548
      Author(s): Harumi Osago, Mikiko Kobayashi-Miura, Yoshifumi Hamasaki, Nobumasa Hara, Mineyoshi Hiyoshi, Mikako Tsuchiya
      Articular cartilage comprises collagens, proteoglycans, and glycosaminoglycans (GAGs) together with water, in hyaline matrixes. Articular cartilage is resistant to proteolytic solubilization for comprehensive GAG analyses partly because of assemblies of collagen fibers with thermolabile hydrogen bonds. In this study, we used the heat-stable protease thermolysin to digest collagen in solid articular cartilage at 70 °C and compared the efficiencies of collagen digestion and GAG extraction to those with collagenase digestion at 50 °C. Overnight digestion with thermolysin completely solubilized cartilage, whereas collagenase with >10-times higher proteolytic activity digested <20% of collagen. Following thermolysin treatments, almost all GAGs were extracted from the cartilage, whereas only 56% of GAGs were extracted after collagenase digestion. Disaccharide analyses of extracted GAG chains revealed >98% extraction efficiencies of several GAG classes from thermolysin-treated cartilage, compared with <60% extraction efficiencies using collagenase, depending on GAG classes. These results indicate that thermolysin allows complete GAG extraction from solid articular cartilage and that complete solubilization is required for accurate and reproducible analyses of cartilage GAGs. Hence, thermolysin offers a tool for complete solubilization of cartilage prior to comprehensive GAGomic analysis, and is likely applicable to other collagen-rich tissues such as ligaments, skin, and blood vessels.

      PubDate: 2018-04-11T10:25:23Z
      DOI: 10.1016/j.ab.2018.02.028
      Issue No: Vol. 548 (2018)
  • A mass spectrometry based transport assay for studying EmrE transport of
           unlabeled substrates
    • Authors: Anne E. Robinson; Jeffrey P. Henderson; Katherine A. Henzler-Wildman
      Abstract: Publication date: Available online 17 March 2018
      Source:Analytical Biochemistry
      Author(s): Anne E. Robinson, Jeffrey P. Henderson, Katherine A. Henzler-Wildman
      Membrane transporters are an important class of proteins which remain challenging to study. Transport assays are crucial to developing our understanding of such proteins as they allow direct measurement of their transport activity. However, currently available methods for monitoring liposomal loading of organic substrates primarily rely on detection of radioactively or fluorescently labeled substrates. The requirement of a labeled substrate significantly restricts the systems and substrates that can be studied. Here we present a mass spectrometry based detection method for liposomal uptake assays that eliminates the need for labeled substrates. We demonstrate the efficacy of the assay with EmrE, a small multidrug resistance transporter found in E. coli that has become a model transport system for the study of secondary active transport. Furthermore, we develop a method for differentiation between bound and transported substrate, enhancing the information gained from the liposomal uptake assay. The transport assay presented here is readily applicable to other transport systems and substrates.
      Graphical abstract image

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.017
  • Facile preparation of highly active casein kinase 1 using Escherichia coli
           constitutively expressing lambda phosphatase
    • Authors: Kazutoshi Akizuki; Taku Toyama; Masashi Yamashita; Yasunori Sugiyama; Atsuhiko Ishida; Isamu Kameshita; Noriyuki Sueyoshi
      Abstract: Publication date: Available online 17 March 2018
      Source:Analytical Biochemistry
      Author(s): Kazutoshi Akizuki, Taku Toyama, Masashi Yamashita, Yasunori Sugiyama, Atsuhiko Ishida, Isamu Kameshita, Noriyuki Sueyoshi
      Casein kinase 1 (CK1) is a widely expressed Ser/Thr kinase in eukaryotic organisms that is involved in various cellular processes (e.g., circadian rhythm and apoptosis). Therefore, preparing highly active CK1 and investigating its properties in vitro have important implications for understanding the biological roles of the kinase. However, recombinant CK1 undergoes autoinactivation via autophosphorylation in Escherichia coli cells and thus is undesirably prepared as a phosphorylated and inactivated kinase. To circumvent this problem, we established a protein expression system using E. coli strain BL21(DE3)pλPP in which λ protein phosphatase (λPPase) is constitutively expressed. Using this system, recombinant CK1 isoforms (α, δ and ε) were readily prepared as unphosphorylated forms. Furthermore, we found that CK1s prepared using BL21(DE3)pλPP showed markedly higher activity than those prepared by the conventional BL21(DE3). Finally, we demonstrated that the kinase activity of CK1δ from BL21(DE3)pλPP was higher than that prepared by a conventional method consisting of troublesome steps such as in vitro λPPase treatment. Thus, this simple method using BL21(DE3)pλPP is valuable for preparing highly active CK1s. It may also be applicable to other kinases that are difficult to prepare because of phosphorylation in E. coli cells.
      Graphical abstract image

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.015
  • An ELISA for the study of calcineurin-NFAT unstructured region interaction
    • Authors: Nesly Dotan; Vera Gayder; Itai Bloch; Maayan Gal
      Abstract: Publication date: Available online 16 March 2018
      Source:Analytical Biochemistry
      Author(s): Nesly Dotan, Vera Gayder, Itai Bloch, Maayan Gal
      Calcineurin is a phosphatase that targets the transcription factor, nuclear factor of activated T-cells (NFAT) dephosphorylates multiple sites along NFAT's regulatory domain. The calcineurin-NFAT complex interaction is mediated through two conserved binding motifs known as the PxIxIT and LxVP, which are located at the N- and C- terminus to the phosphorylation sites. The vast range of cellular processes regulated by the calcineurin-NFAT interaction has aroused great interest in the investigation of the structural aspects that govern their complex formation and in the discovery of protein-protein interaction inhibitors; the latter interfere with calcineurin-NFAT complex formation while keeping calcineurin's catalytic site free. To assist additional biophysical study of the calcineurin-NFAT structure-function relation and to screen for new inhibitors, we present a robust and cost-effective Enzyme Linked Immuno Sorbent Assay (ELISA) that is based on the interaction of calcineurin with the NFAT homology region. The latter includes the two calcineurin's binding sites, in addition to the phosphorylation sites. The ELISA experiment shown here can thus be applied towards the study of important structural aspects of the complex and for the discovery of new inhibitors. This will allow for a better understanding of T-cell activation switch.

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.014
  • Metabolic profiling of moulds with laser desorption/ionization mass
           spectrometry on gold nanoparticle enhanced target
    • Authors: Adrian Arendowski; Justyna Szulc; Joanna Nizioł; Beata Gutarowska; Tomasz Ruman
      Abstract: Publication date: Available online 16 March 2018
      Source:Analytical Biochemistry
      Author(s): Adrian Arendowski, Justyna Szulc, Joanna Nizioł, Beata Gutarowska, Tomasz Ruman
      Surface-assisted laser desorption/ionization mass spectrometry on gold nanoparticle enhanced target (AuNPET) technique was used for metabolomic analysis and secondary metabolites detection of two mould strains – Aspergillus versicolor and Penicillium chrysogenum in model conditions on microbiological malt extract agar medium. Results obtained with the use of AuNPET-based mass spectrometry technique were compared with traditional matrix-assisted laser desorption/ionization (MALDI) method based on α-cyano-4-hydroxycinnamic acid (CHCA) and 2,5-dihydroxybenzoic acid (DHB) matrices. Gold nanoparticle enhanced target method enabled effective ionization of microbial cellular extract ingredients without interference from the matrix and also improved calibration of spectra resulting in the detection of much higher amount of characteristic metabolites for studied organisms than MALDI.
      Graphical abstract image

      PubDate: 2018-03-19T20:43:07Z
      DOI: 10.1016/j.ab.2018.03.016
  • 3-Amino-1-phenyl-2-pyrazoline-5-ketone as a heterobifunctional chromogenic
           reagent to derivatize reducing glycans for subsequent online LC/MS
    • Authors: Yu Lu; Chengjian Wang; Rendan Liu; Wanjun Jin; Yanan Wen; Linjuan Huang; Zhongfu Wang
      Abstract: Publication date: Available online 7 March 2018
      Source:Analytical Biochemistry
      Author(s): Yu Lu, Chengjian Wang, Rendan Liu, Wanjun Jin, Yanan Wen, Linjuan Huang, Zhongfu Wang
      Sensitive analysis of glycans by liquid chromatography/mass spectrometry is significantly hampered by the lack of chromogenic or fluorescent groups on the glycan structures, as well as, their poor ionization properties. In the present, a heterobifunctional chromogenic reagent 3-amino-1-phenyl-2-pyrazoline-5-ketone (PAP) bearing amino and active methylene groups, which readily reacts with reducing glycans, was used for detection of the pre-column-labeled glycans via high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS). The PAP derivatives with active methylene and amino groups were obtained via reductive amination in acidic medium and condensation of an active PAP methylene group with the reducing end of glycans in alkaline medium, respectively, and the PAP derivatives could be further functionalized, e.g., via glycan microarray preparation. The conditions for the two reaction modes were optimized, the HPLC separation method of PAP derivatives was investigated, and the PAP derivatives of some glycans derived from biological samples were obtained and analyzed by ESI-MS and LC-MS. Using this new reagent, reducing glycans can be selectively derivatized by different reaction mechanisms, having great importance for functional glycomics studies.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.02.029
  • Analyzing mitochondrial function in human peripheral blood mononuclear
    • Authors: Chao-Pin Hsiao; Charles Hoppel
      Abstract: Publication date: Available online 2 March 2018
      Source:Analytical Biochemistry
      Author(s): Chao-Pin Hsiao, Charles Hoppel
      Mitochondrial oxidative phosphorylation (OXPHOS) is responsible for producing most of the adenosine triphosphate required by eukaryotic cells. Lymphocytes make up the majority of the peripheral blood mononuclear cells. Peripheral blood mononuclear cells are readily obtainable, providing an ideal sample to monitor systemic changes and understand molecular signaling mechanisms in disease processes. Mitochondrial energy metabolism of lymphocyte has been used to screen for OXPHOS disorders. While there are increasing studies of lymphocyte OXPHOS, few studies examined activity of electron transport chain of lymphocyte mitochondria. We present an optimal protocol to harvest fresh peripheral blood mononuclear cells from human whole blood, determine integrated mitochondrial function, and analyze electron transport chain complex activity. Analyzing integrated mitochondrial function using OXPHOS provides data to uncover defects in the transport of substrates into the mitochondria, generation of reducing equivalents, the electron transport chain, and coupling to the production of adenosine triphosphate. The optimal conditions to harvest peripheral blood mononuclear cells were using blood anticoagulated with ethylenediaminetetraacetic acid, processed utilizing Lymphoprep™, and washed in phosphate buffered saline, all at room temperature. Using isolated peripheral blood mononuclear cells, integrated mitochondrial function and the activities of electron transport chain were determined.

      PubDate: 2018-03-08T03:19:02Z
      DOI: 10.1016/j.ab.2018.03.003
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