Shen Yang Tong

Microdetermination of proteins by Rayleigh light scattering technique with acid green 25

Abstract Based on the enhancement of Rayleigh light scattering (RLS) of acid green 25 (Ag25) by proteins, a new quantitative determination method for proteins in aqueous solutions has been developed. This assay is very sensitive (0.136-10.20 μg/ml), rapid (

Determination of proteins by fluorescence quenching of erythrosin B

Abstract Erythrosin B (EB) binding to proteins causes a decrease in the fluorescence maximum of EB at 550 nm. Based on this, a new, fast and simple fluorescence quenching method for the determination of proteins is developed. The linear range of this assay is 1.36–20.4 μg ml −1 . The method has very few interferences, most of which can be minimized by dilution. In the detection of proteins in human serum, this method gave values close to that of the Coomassie brilliant blue method.

Enhancement of Rayleigh light scattering of acid chrome blue K by proteins and protein assay by the scattering technique.

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 micrograms 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 lest 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.

Spectrophotometry of nucleic acids by their effect on the complex of cobalt(II) with 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene

Abstract A new ternary azo-dye complex reaction involving nucleic acids has been studied to provide a sensitive spectrophotometric method for nucleic acids. In alkaline conditions, single-stranded nucleic acids react with the complex of Co(II) with 4-[(5-chloro-2-pyridyl)azo]-1,3-diaminobenzene (5-Cl-PADAB), resulting in purple ternary compounds with maximum absorbance at 545.0 nm against the binary complex blank. The mole ratio of Co(II) to 5-Cl-PADAB in the ternary compounds is 1:2, and each nucleotide residue can bind two binary complex molecules. There are linear relationships between the absorbance and the concentrations of the tested nucleic acids, such as calf thymus DNA, fish sperm DNA and yeast RNA in the range 0–4.0 μg ml −1 . The 3σ limit of detection for calf thymus DNA is 40 ng ml −1 , the relative standard deviation at 3.0 μg ml −1 is 0.79% ( n = 10) and the Sandells sensitivity is 5.3 ng cm −2 . The same parameters for fish sperm DNA are 41 ng ml −1 , 0.84% and 5.4 ng cm −2 , and for yeast RNA are 49 ng ml −1 , 0.58% and 6.2 ng cm −2 . Synthetic samples were analyzed satisfactorily.

A study on the interaction between concanavalin A and glycogen by light scattering technique and its analytical application.

The mixture of concanavalin A (Con A) and glycogen shows strong light scattering character. Based on it, the interaction between Con A and glycogen was studied on a common spectrofluorimeter by light scattering technique. Many factors affecting the light scattering intensity (LSI), such as pH, temperature, reaction time, ion strength and the denaturing agent of protein were studied in detail. Experimental results showed that the LSI reached its maximum after mixing Con A with glycogen for about 20 min in pH 7.4 Tris-HCl buffer at 37 degrees C. The results also suggested that the conformation of Con A was critical for its unique binding affinity to glycogen. Electrostatic forces should not be the primary interaction between glycogen and Con A. Under proper experimental conditions, the determination method for glycogen by light scattering technique was developed. The glycogen determination can be performed in the range of 0.48-32.0, 0.50-32.0 and 0.32-24.0 mug/ml for Rabbit liver glycogen (RL Gly), Oyster glycogen (O Gly) and Clam Glycogen (C Gly), respectively. The influence of co-existing substances such as proteins, mono- and di-saccharides and metal ions was evaluated, and little interference came from the foreign substances. The determinations of glycogen in synthetic samples demonstrated that the recovery rate was in the range of 98.1-103% and the relative standard deviations (RSD) were lower than 5.0%.

Molecular spectroscopy study of the reaction of nucleic acids with brilliant cresol blue.

The interaction of brilliant cresol blue (BCB) with nucleic acids in aqueous solution has been studied by spectrophotometry and Rayleigh light scattering (RLS) spectroscopy. Under suitable conditions, the RLS spectra of BCB changed significantly due to the presence of nucleic acids. RLS intensity of BCB at 364 nm is greatly enhanced with the addition of nucleic acids, and a new RLS peak is observed at 552 nm. This peak is about half the intensity of that at 364 nm. The results of this study show that BCB interacts with DNA possibly due to the cooperative effect of electrostatic attraction, intercalation, coordination and hydrophobic effect. Under optimum conditions, the increase of RLS at 364 nm of a BCB solution is proportional to the concentration of nucleic acids added. This result is the basis for a new RLS method for determination of nucleic acids. The linear range of ctDNA, fsDNA and yRNA is 0.12-4.70, 0.11-4.64 and 0.43-7.07 microg ml(-1), respectively.

Spectrophotometric micromethod for protein determination with tetrachloro tetraiodo fluorescein

Abstract Reaction of tetrachloro tetraiodo fluorescein (Na2TCTIF) with protein results in an increase in its absorption at 568nm. This assay gives a linear response to BSA at 0.68–34.0 μg /ml, and is almost as sensitive as the Bradford method1. The reaction is completed in approximately 2 min and has good color stability for at least 1 h. Addition of triton X-100 increases the sensitivity and stability of Na2TCTIF binary system and the reason was studied. The binary and ternary assays were compared in protein-to-protein variability, interferences and the type of binding effort. In detection of human serum albumin, these two ways give results with good reproducibility and are comparable with the bromocresol green (BCG) method.

Spectrofluorimetric determination of nucleic acids as 8-hydroxyquinoline/ yttrium ternary complexes

The formation of nucleic acids/8-hydroxyquinoline/yttrium(III) ternary complexes and their fluorescent properties have been studied. The nucleic acids studied include native and thermally denatured calf thymus DNA, fish sperm DNA and yeast RNA. In the range of pH 7.6–8.5, controlled by NH3-NH4C1 buffer, ternary complexes are formed that fluoresce at different wavelengths with different nucleic acids. Based on the fluorescence reactions, sensitive spectrofluorimetric methods for nucleic acids are proposed. In optimal conditions, the calibration curves were linear in the range 0.5–4.0 μgml−1 for calf thymus DNA, 0.5–2.5 μg ml–1 for fish sperm DNA and 0.5–4.0 μg ml–1 for yeast RNA. The limits of determination (3 σ) were 0.030 μg ml−1 for calf thymus DNA, 0.020 μg ml−1 for fish sperm DNA and 0.090 μg ml−1 for yeast RNA. Corresponding to the interferences of coexisting substances, six synthetic samples were constructed and the results of determination were satisfactory.