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Shenyang Tong - One of the best experts on this subject based on the ideXlab platform.
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A Novel Protein Assay With an Azo Dye Using Rayleigh Light Scattering Technioque
Analytical Letters, 2000Co-Authors: Ya-ting Wang, Feng-lin Zhao, Shenyang TongAbstract:ABSTRACT The interaction of SPADNS [3-(4-sulfophenylazo)-4, 5-dihydroxy-2, 7-naphthalene disulfonic acid] with Proteins in acidic solution was studied by spectrophotometric and Rayleigh light scattering (RLS) methods. SPADNS reacts rapidly with albumin at room temperature, and the intensity of the weak RLS of SPADNS was enhanced remarkably and quantitatively by this reaction. Based on this observation, a novel RLS method for serum albumin determination was developed. The linear range is 0.125-14.9 μg/ml for BSA. The relative standard deviation (n=10) for 5.0 μg/ml BSA was 3.94%. The method was applied to the determination of Proteins in human plasma, and the results were compared with the traditional Proteins Assay (CBB method). There is almost no interference from amino acids, metal ions and other coexistent substances. The reaction mechanism was also discussed.
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A linear regression method for the study of the Coomassie brilliant blue Protein Assay.
Talanta, 1997Co-Authors: Yong-ju Wei, Shenyang TongAbstract:Abstract The interactions of Coomassie brilliant blue G-250 (CBB) with bovine serum albumin (BSA) and γ-globulin at low pH are investigated by a spectrophotometric method. It is considered that the binding of CBB to Protein is because of the weak interactions (ionic, van der Waals, hydrogen bonding, and hydrophobic). The solution equilibria involving the binding of three dye species (blue, green, and red) to Protein are treated in the same way as Ringbom model used in the treatment of complexation in analytical chemistry. Based on this treatment, the formation of an isosbestic point in the absorption spectra of CBB-BSA mixtures is discussed, two mathematical models for the description of the CBB Protein Assay are developed. The first model is a nonlinear equation which is rigorous in theory but unreliable in use because of its optimization procedure. The second model based on an approximation is a linear equation, it allows to estimate apparent binding constant, maximum binding number, and molar absorptivity of bound dye from Assay data by a linear regression method. The results of the linear regression operations are reasonable and in agreement with experimental findings. Factors which influence the sensitivity of the CBB Protein Assay are studied using this method. Ionic strength and acidity are found to have significant effect on the binding of CBB to Protein.
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Enhancement of Rayleigh Light Scattering of Acid Chrome Blue K by Proteins and Protein Assay by the Scattering Technique
Analyst, 1997Co-Authors: Shenyang TongAbstract:Rayleigh light scattering of Acid Chrome Blue K (ACBK) is enhanced greatly by Proteins. Based on this, a method for Protein Assay in aqueous solution was developed. This Assay matches the sensitivity of the colorimetric dye-binding method with a linear range of 0.136–10.88 µg ml - 1 . The measurements can be made easily on a common fluorimeter. The reaction between ACBK and Proteins is completed in 2 min and the scattered light signal is stable for at least 3 h. Protein-to-Protein variability is encountered in this method as in many other Protein Assays. There is little or no interference from amino acids, most metal ions and complexing agents (e.g., EDTA). Interferences from salts, urea and detergents can be minimized by dilution.
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A Novel Protein Assay Method Using Tetraphenylporphin tetrasulfonate (TPPS4)
Analytical Letters, 1995Co-Authors: Shenyang TongAbstract:Abstract A sensitive Protein Assay method which involves the reaction of TPPS4 with Protein is described. When Protein is added to TPPS4 solution, an absorption band with the maximum at 488 nm appears and the absorbance is proportional to the concentration of Protein. Just Like the Soret absorption of the porphyrin, the new band is very narrow and there is no overlap at all between them, which means the free dyes would not give any background for the detection of the Protein-TPPS4 complexes. A new spectrophotometric method for determination of Protein has been constructed and applied to the determination of human plasma Protein and urinary Protein; The Assay using microtiter plates has also been studied.
Thierry Rabilloud - One of the best experts on this subject based on the ideXlab platform.
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Optimization of the cydex blue Assay: A one-step colorimetric Protein Assay using cyclodextrins and compatible with detergents and reducers.
PLOS ONE, 2018Co-Authors: Thierry RabilloudAbstract:Sodium dodecyl sulfate electrophoresis (SDS) is a Protein separation technique widely used, for example, prior to immunoblotting. Samples are usually prepared in a buffer containing both high concentrations of reducers and high concentrations of SDS. This conjunction renders the samples incompatible with common Protein Assays. By chelating the SDS, cyclodextrins make the use of simple, dye-based colorimetric Assays possible. In this paper, we describe the optimization of the Assay, focussing on the cyclodextrin/SDS ratio and the use of commercial Assay reagents. The adaptation of the Assay to a microplate format and using other detergent-containing conventional extraction buffers is also described.
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A single step Protein Assay that is both detergent and reducer compatible: The cydex blue Assay.
Electrophoresis, 2016Co-Authors: Thierry RabilloudAbstract:Determination of Protein concentration in often an absolute pre-requisite in preparing samples for biochemical and proteomic analyses. However, current Protein Assay methods are not compatible with both reducers and detergents, which are however present simultaneously in most denaturing extraction buffers used in proteomics and electrophoresis, and in particular in SDS electrophoresis. We found that inclusion of cyclodextrins in a Coomassie blue-based Assay made it compatible with detergents, as cyclodextrins complex detergents in a 1:1 molecular ratio. As this type of Assay is intrinsically resistant to reducers, we have thus developed a single step Assay that is both detergent and reducer compatible. Depending on the type and concentration of detergents present in the sample buffer, either beta-cyclodextrin or alpha-cyclodextrin can be used, the former being able to complex a wider range of detergents and the latter being able to complex higher amounts of detergents due to its greater solubility in water. Cyclodextrins are used at final concentrations of 2-10 mg/mL in the Assay mix. This typically allows to measure samples containing as little as 0.1 mg/mL Protein, in the presence of up to 2% detergent and reducers such as 5 % mercaptoethanol or 50 mM DTT in a single step with a simple spectrophotometric Assay. This article is protected by copyright. All rights reserved.
Jamila Kalthoum Cherif - One of the best experts on this subject based on the ideXlab platform.
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Study of Tunisian nettle leaves (Urtica dioica L.)
International Journal of Pharmacognosy and Phytochemical Research, 2016Co-Authors: F. Sidaoui, Souad Igueld Belghith, Malika Trabelsi-ayadi, Danielle Barth, Jamila Kalthoum CherifAbstract:This work was undertaken to explore the potential of extract of leaves Nettle (Urtica dioica L.) of Tunisia as sources of minerals and natural antioxidants. Two extraction methods were used and compared, the supercritical fluid extraction (SFE) with carbon dioxide (CO2) and traditional maceration extraction using ethanol (EtOH) as solvent. The SFE was explored at various operating conditions of pression (15 and 18 MPa) and temperature (40 and 60°C) (with or without glass beads). The phenolic and flavonoid contents were determined respectively by Folin-Ciocalteu and Aluminium chloride colorimetric method. The ABTS Assay was used for determining the antioxidant activity. The Protein Assay was performed by the Kjeldahl method and the multielemental analysis by atomic emission spectrometry with inductively coupled plasma. The results show some variations between both extracts in terms of extraction yield and antioxidant activity. The SFE, with carbon dioxide (SC-CO2) is an interesting alternative to the conventional extraction using organic solvents. However, higher values of phenolic content (11.62 mg GAE.g-1 DW), flavonoid content (7.10 mgCE.g-1DW) and antioxidant activity TEAC (8.11 μM) correspond to extracts obtained by maceration. In all SFE extract Assays, the highest yield was reached at 60°C – 15MPa with glass beads (2.5%). The ICP analysis shows the abundance of Urtica dioica L. in calcium (1116 mg.100g-1DW) and magnesium (544). The Protein Assay showed high content (15.75 %.). The present study indicates that the leaves of Urtica dioica L. of Tunisia are a potential source of minerals, Proteins and antioxidants
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Study of Tunisian nettle leaves (Urtica dioica L.): mineral composition and antioxidant capacity of their extracts obtained by maceration and supercritical fluid extraction
International Journal of Pharmacognosy and Phytochemical Research, 2015Co-Authors: F. Sidaoui, Souad Igueld Belghith, Malika Trabelsi-ayadi, Danielle Barth, Jamila Kalthoum CherifAbstract:This work was undertaken to explore the potential of extract of leaves Nettle (Urtica dioica L.) of Tunisia as sources of minerals and natural antioxidants. Two extraction methods were used and compared, the supercritical fluid extraction (SFE) with carbon dioxide (CO2) and traditional maceration extraction using ethanol (EtOH) as solvent. The SFE was explored at various operating conditions of pression (15 and 18 MPa) and temperature (40 and 60°C) (with or without glass beads). The phenolic and flavonoid contents were determined respectively by Folin-Ciocalteu and Aluminium chloride colorimetric method. The ABTS Assay was used for determining the antioxidant activity. The Protein Assay was performed by the Kjeldahl method and the multielemental analysis by atomic emission spectrometry with inductively coupled plasma. The results show some variations between both extracts in terms of extraction yield and antioxidant activity. The SFE, with carbon dioxide (SC-CO2) is an interesting alternative to the conventional extraction using organic solvents. However, higher values of phenolic content (11.62 mg GAE.g-1 DW), flavonoid content (7.10 mgCE.g-1DW) and antioxidant activity TEAC (8.11 μM) correspond to extracts obtained by maceration. In all SFE extract Assays, the highest yield was reached at 60°C – 15MPa with glass beads (2.5%). The ICP analysis shows the abundance of Urtica dioica L. in calcium (1116 mg.100g-1DW) and magnesium (544). The Protein Assay showed high content (15.75 %.). The present study indicates that the leaves of Urtica dioica L. of Tunisia are a potential source of minerals, Proteins and antioxidants
Katherine M Williams - One of the best experts on this subject based on the ideXlab platform.
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extent of aminoglycoside interference in the pyrogallol red molybdate Protein Assay depends on the concentration of sodium oxalate in the dye reagent
Clinical Chemistry, 2004Co-Authors: Thomas Marshall, Katherine M WilliamsAbstract:The Pyrogallol Red-molybdate (PRM) reagent of Fujita et al. (1) has been modified by Watanabe et al.(2) for urinary Protein determination. The Assay has been widely adopted for this purpose, but interference from aminoglycoside antibiotics can produce falsely increased urinary Protein values in acute-care patients, and the extent of interference varies with different PRM reagents (3)(4). Thus, the Dade Behring and Sigma reagents are susceptible to interference, whereas the Cobas Fara and Roche Integra 700 reagents are resistant (3)(4). The present study compares the PRM reagents of Fujita et al. (1) and Watanabe et al. (2) and demonstrates that the aminoglycoside interference varies with the concentration of sodium oxalate in the dye reagent. Amikacin (cat. no. A1774), dihydrostreptomycin (cat. no. D7253), geneticin (cat. no. G5013), gentamicin (cat. no. G1914), kanamycin (cat. no. K4000), neomycin (cat. no. N5285), paromomycin (cat. no. P9297), streptomycin (cat. no. S6501), and tobramycin (cat. no. T1783) were purchased from Sigma-Aldrich. Aqueous aminoglycoside solutions (10 g/L) were prepared gravimetrically and diluted to 1 g/L before Assay. Pierce Prediluted Protein Assay Standard (1 g/L bovine serum albumin 23208) was purchased from Perbio Science UK Ltd. Urine control (AU2353) was purchased from Randox Laboratories Ltd. and reconstituted in deionized water or aqueous aminoglycoside (final concentration, 0.2 or 1.0 …
Justin J Cooperwhite - One of the best experts on this subject based on the ideXlab platform.
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one step homogeneous c reactive Protein Assay for saliva
Faculty of Health; Institute of Health and Biomedical Innovation, 2011Co-Authors: Chamindie Punyadeera, Goce Dimeski, Karam Kostner, Peter Beyerlein, Justin J CooperwhiteAbstract:Background: Cardiovascular disease is the leading cause of death in the world. Human C-reactive Protein (CRP) has been used in the risk assessment of coronary events. Human saliva mirrors the body's health and well-being and is non-invasive, easy to collect and ideal for third world countries as well as for large patient screening. The aim was to establish a saliva CRP reference range and to demonstrate the clinical utility of salivary CRP levels in assessing the coronary events in a primary health care setting. Methods: We have used a homogeneous bead based Assay to detect CRP levels in human saliva. We have developed a rapid 15 min (vs 90 min), sequential, one-step Assay to detect CRP in saliva. Saliva was collected from healthy volunteers (n = 55, ages 20-70 years) as well as from cardiac patients (n = 28, ages 43-86 years). Results: The Assay incubation time was optimised from 90 min to 15 mm and generated a positive correlation (n = 29, range 10-2189 pg/mL, r2 = 0.94; Passing Bablok slope 0.885. Intercept 0, p>0.10), meaning we could decrease the incubation time and produce equivalent results with confidence. The mean CRP level in the saliva of healthy human volunteers was 285 pg/mL and in cardiac patients was 1680 pg/mL (p<0.01). Analysis of CRP concentrations in paired serum and saliva samples from cardiac patients gave a positive correlation (r2 = 0.84, p<0.001) and the salivary CRP concentration capable of distinguishing healthy from diseased patients. Conclusions: The results suggest that this minimally invasive, rapid and sensitive Assay will be useful in large patient screening studies for risk assessment of coronary events. (C) 2011 Elsevier B.V. All rights reserved.
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one step homogeneous c reactive Protein Assay for saliva
Journal of Immunological Methods, 2011Co-Authors: Chamindie Punyadeera, Goce Dimeski, Karam Kostner, Peter Beyerlein, Justin J CooperwhiteAbstract:Background: Cardiovascular disease is the leading cause of death in the world. Human C-reactive Protein (CRP) has been used in the risk assessment of coronary events. Human saliva mirrors the body's health and well-being and is non-invasive, easy to collect and ideal for third world countries as well as for large patient screening. The aim was to establish a saliva CRP reference range and to demonstrate the clinical utility of salivary CRP levels in assessing the coronary events in a primary health care setting. Methods: We have used a homogeneous bead based Assay to detect CRP levels in human saliva. We have developed a rapid 15. min (vs 90. min), sequential, one-step Assay to detect CRP in saliva. Saliva was collected from healthy volunteers (n = 55, ages 20-70. years) as well as from cardiac patients (n = 28, ages 43-86. years). Results: The Assay incubation time was optimised from 90. min to 15. min and generated a positive correlation (n = 29, range 10-2189. pg/mL, r2 = 0.94; Passing Bablok slope 0.885, Intercept 0, p > 0.10), meaning we could decrease the incubation time and produce equivalent results with confidence. The mean CRP level in the saliva of healthy human volunteers was 285. pg/mL and in cardiac patients was 1680. pg/mL (p < 0.01). Analysis of CRP concentrations in paired serum and saliva samples from cardiac patients gave a positive correlation (r2 = 0.84, p < 0.001) and the salivary CRP concentration capable of distinguishing healthy from diseased patients. Conclusions: The results suggest that this minimally invasive, rapid and sensitive Assay will be useful in large patient screening studies for risk assessment of coronary events.
