Novel approach for the effective determination of DNA scission site using the Sanger method

This paper describes the effective determination of DNA scission site using a novel approach that is based on the Sanger method of nucleotide sequencing. The DNA scission site is determined by contrast with the nucleotide sequence of the DNA. Here, instead of the traditional Maxam-Gilbert method for the determination of the DNA sequence, we utilized the Sanger method and studied its effectiveness in the determination of DNA scission sites. Using this method, the determination of DNA scission site becomes more facile and exact. And the total time for the determination is reduced nearly by half in comparison to the Maxam-Gilbert method. Further advantages of this novel approach include the reduced risks of radiation exposure for researchers and contamination of the apparatus.     

呼吸气体一次进样气相色谱分析方法

本文报道在 SP-2305E 型气相色谱仪上,采用由聚合物固定相 GDX-104柱(80/100目,φ4×600ram)、钠石灰柱(40/60目,φ4×350mm)及5A 分子筛柱(60/80目,φ4×2000mm)所组成的单气路双柱跨鉴定器(热导池)色谱流程,以氢为载气(40ml/min),桥流160mA,柱箱温度50°С,一次进样测定呼吸气体氧及二氧化碳含量的方法。进样量1ml,完成一次分析的周期为4 1/2min。氧测定的精确度与 Scholander 微量呼吸气体分析方法相当,二氧化碳测定的精确度则优于后者。还比较了分别以氢或氩为载气时的定量结果。并就定量方法及“氩校正”问题进行了讨论。     

Rapid method of determining the dry mass of whooping cough microbes

A colorimetric method for the rapid determination of the quantitative content of microbial mass in B. pertussis suspensions has been developed. The method is based on the indirect determination of carbon in microbial suspensions by its oxidation with the mixture consisting of potassium bichromate in concentrated sulfuric acid and the subsequent colorimetric analysis of the products of this reaction. The method ensures sufficient accuracy, the determination procedure is simple, takes not more than 2 hours and requires no complex reagents. The results thus obtained are well comparable with those obtained by the classical gravimetric method. The new method permits the determination of microbial mass in B. pertussis suspensions with a minimum concentration of 0.5 mg/ml. The method is recommended for the determination of dry microbial mass in B. pertussis suspensions.     

A spectrophotometric method for simultaneous determination of protein-bound hexoses and fucose with a mixture of L-cysteine and phenol

A very suitable spectrophotometric method for simultaneous determination of protein-bound hexoses and fucose is presented. A mixture of L-cysteine and phenol in sulfuric acid was used as reagent, whereas absorption measurements were carried out at two wavelengths, namely at 398 nm for fucose and 490 nm for hexoses determination. Optimum conditions for the application of the method were established, special attention being paid to the possible interference of fucose determination with that of hexoses, and vice versa. The proposed method was applied to the determination of hexoses and fucose in serum glycoproteins and seromucoids. The method was found to be very practical enabling a simultaneous determination of both kinds of carbohydrates; moreover, it was proved to be more sensitive and specific in comparison with methods commonly used for individual determination of fucose with L-cysteine and hexoses with phenol.     

Method of determining proteolytic activity by using a conjugate of bovine serum albumin with peroxidase

The authors propose a method for determination of proteolytic activity, based on the hydrolysis of peroxidase-labeled molecules of bovine serum albumin immobilized on the surface of polystyrene microassay plates with the subsequent determination of peroxidase activity on the carrier or in the solution. The optimum conditions for the sorption of the labeled substrate have been established. The method permits the determination of bacillary alkaline protease at a concentration of 01. microgram/ml within 45 minutes. The determination of four proteases has demonstrated that this method shows good correlation with the routine one (r = 0.98), but is more sensitive and less time- and labor-consuming.     

A semi-integrated method for the determination of enzyme kinetic parameters and graphical representation of the Michaelis-Menten equation

A semi-integrated method for the determination of the enzyme kinetics parameters (Km and V) and graphical representation of the Michaelis-Menten equation is proposed as a variation of determination of initial reaction rate (v) as a function of initial substrate concentration ([S]0). The method is based on the determination of the time required to exhaust half of the initial substrate concentration as a function of the initial substrate concentration. The advantages and limitations of this method are discussed.     

Use of the respiration activity ofThiobacillus ferrooxidans for the specific determination of iron(II,III)

A method is described for the determination of Fe2+ and Fe3+ after its reduction to Fe2+ on the basis of oxygen uptake rate as a function of Fe2+ concentration. By using substrate-specific Thiobacillus ferrooxidans in combination with the standard addition method a specific determination of iron(II, III) is possible with the determination limit of 3 mumol/L.     

Determination of the enzymatic traits of anaerobic gram-negative nonsporulating bacteria

The method requiring the use of a semi-liquid medium with the reduced content of protein hydrolysates and with bromo-thymol blue added as indicator and permitting the determination of sugar fermentation by bacteroids and fusobacteria is presented. The determination is made by a change in the color of the indicator and the intensity of microbial growth (opacity) after the incubation of the inoculated medium under free access of air. A study has been made for comparing the above method with the electrometric method for the determination of fermentative features in anaerobic microorganisms.     

Deoxycorticosterone, 18-OH-deoxycorticosterone and corticosterone determination by high performance liquid chromatography in monolayer adrenal cell culture.

Reversed-phase HPLC offers a rapid, qualitative as well as quantitative method for separation and determination of deoxycorticosterone, 18-OH-deoxycorticosterone and corticosterone in primary culture of adrenocortical cells. The resolution is sufficient for the purpose of quantitative determination. The limitation of this method lies in its sensitivity for serum steroid determination, but is perfectly applicable at cell cultures where the concentration of these steroids is highest.     

An electrochemical approach to quantitation and characterization of metallothioneins

An analysis was made of the potential of the electrochemical approach to quantitation and characterization of metallothioneins. The Brdi ka procedure for determination of sulfhydryl groups was shown to be of utility as a standard determination method while anodic stripping voltametry was found to offer advantage in determination of protein-bound zinc and cadmium. Cyclic voltametry was observed to differentiate metallothioneins from other sulfhydryl- or disulfide-containing compounds, suggesting considerable potential as a characterizing method for metallothioneins.     

Identification of both termini of an oligonucleotide in a nanomole quantity

The use of the Varian Aerograph LCS-1000 is extended to the determination of mononucleotides and mononucleoside 3′,5′-diphosphates on the nanomole level. In combination with the previously described method for the determination of nucleosides the new method allows identification of both termini in an oligonucleotide of any length. Only one enzyme is used to degrade the oligonucleotide, venom exonuclease for a 3′-oligonucleotide or micrococcal nuclease for a 5′-oligonucleotide. The same method also allows the elucidation of the sequence of a trinucleotide, and the determination of chain length of oligonucleotide.     

A method for the determination of nitrate reductase

A procedure for the assay of nitrate reductase based on Szekely's diaminodiphenylsulphone method of nitrate determination (Szekely, E. (1967) Talanta 14, 941–950) is described. The method is simple and sensitive, allowing determination of 1 μg of nitrate in a volume of 1 ml or less. It is particularly suited to the determination of nitrate reductase.     

Limb locomotion--speed distribution analysis as a new method for stance phase detection.

The stance phase is used for the determination of many parameters in motion analysis. In this technical note the authors present a new kinematical method for determination of stance phase. From the high-speed video data, the speed distribution of the horizontal motion of the distal limb is calculated. The speed with the maximum occurrence within the motion cycle defines the stance phase, and this speed is used as threshold for beginning and end of the stance phase. In seven horses the results obtained with the presented method were compared to synchronous stance phase determination using a force plate integrated in a hard track. The mean difference between the results was 10.8 ms, equalling 1.44% of mean stance phase duration. As a test, the presented method was applied to a horse trotting on the treadmill, and to a human walking on concrete. This article describes an easy and safe method for stance phase determination in continuous kinematic data and proves the reliability of the method by comparing it to kinetic stance phase detection. This method may be applied in several species and all gaits, on the treadmill and on firm ground.     

Spectrofluorimetric determination of certain biologically active phenothiazines in commercial dosage forms and human plasma

A validated simple and sensitive spectrofluorimetric method was developed for the determination of chlorpromazine hydrochloride, promethazine hydrochloride, trifluperazine hydrochloride, thioridazine hydrochloride, perazine maleate and oxomemazine. The method was based on condensation of malonic acid/acetic anhydride (MAA) under the catalytic effect of the tertiary amine moiety of the studied phenothiazines to provide a deep yellow to brown colour with green florescence. Relative fluorescence intensity of the products was measured at λ_exc 398 nm and λ_em 432 nm. Different variables affecting the reaction were studied and optimized. The method was successfully applied for the determination of the studied drugs in commercial dosage forms. The lower detection limits allowed the application of this method for the determination of the compounds in plasma as an example of a biological fluid. In addition, the method was considered specific for the determination of tertiary amines in the presence of primary and secondary amines; as a result, it was deemed suitable for the determination of the cited drugs in the presence of their degradation products resulting from N‐dealkylation or oxidation of the corresponding sulphoxides or sulphones. Copyright © 2012 John Wiley & Sons, Ltd.     

Filtration-fluorescence determination of DNA sedimentation profiles

A method for determination of DNA sedimentation profiles in density gradients of sucrose or salts is proposed. The method consists in isolation and purification of DNA from the fractions by molecular filtration and a subsequent determination of DNA content by the fluorescence of the DNA-ethidium bromide complex.     

A method for parallel determination of choline acetyltransferase and muscarinic cholinergic receptors: Application in aged-rat brain

We have devised a method for the parallel determination of choline acetyltransferase (CAT) and muscarinic cholinergic receptor (mCh-R) in the same brain tissue. The method for CAT activity determination is more rapid, simplified, stable, and economical than the usual Fonnum's method. With our method, age-associated changes in CAT activity and mCh-R levels were examined. Although CAT activity hardly changed with age except in a few areas, mCh-R binding of aged-rats was markedly reduced in all areas. These results suggest that the change in mCh-R represents an age-associated biochemical change in the brain and that determination of CAT activity is not sufficient for the study of age-associated changes in the brain cholinergic system.     

Flow injection chemiluminescence determination of captopril based on its enhancing effect on the luminol–ferricyanide/ferrocyanide reaction

A new flow injection chemiluminescence method is described for the determination of captopril. It is based on the enhancing effect of captopril on the chemiluminescence reaction of luminol with potassium ferricyanide in alkaline solution in the presence of potassium ferrocyanide. The method allows the determination of captopril over 0.1–40 µg/mL range, with a relative standard deviation (SD) of 1.0% for the determination of 0.5 µg/mL captopril solution in 11 repeated measurements. The method was satisfactorily applied to the determination of captopril in commercial captopril tablets. The possible reaction mechanism is also discussed briefly. Copyright © 2002 John Wiley & Sons, Ltd.     

需氧菌阴道炎／细菌性阴道病联合测定技术诊断需氧菌性阴道炎的临床价值

目的探讨需氧菌阴道炎／细菌性阴道病联合测定技术诊断需氧菌性阴道炎（AV）的临床价值。方法分别采用镜检法和需氧菌阴道炎／细菌性阴道病联合测定技术对200例疑似AV患者进行检测。结果在200例疑似AV患者中,镜检法检出阳性患者174例,需氧菌阴道炎／细菌性阴道病联合测定法检出阳性患者169例。以镜检法为诊断标准,需氧菌阴道炎／细菌性阴道病联合测定技术的敏感性为94．83％、特异性为84．62％。两种方法的符合率为93．50％[（165＋22）／200],两者呈中度一致性（Kappa=0．735,P〉0．05）。结论需氧菌阴道炎／细菌性阴道病联合测定技术诊断AV具有很高的敏感度和特异度,与目前临床使用的常规镜检法符合率高,且该方法操作简便、快速,值得临床推广使用。     

生物传感器法测定麦芽汁中赖氨酸含量的研究

本研究利用固定化赖氨酸氧化酶和生物传感器,建立了麦芽汁中赖氨酸含量的测定方法。研究了该方法的线性范围、检出限、最适pH值范围、稳定性及专一性。以大生产麦芽汁为样品,与其它检测方法进行了比较。结果表明,生物传感器法测定赖氨酸含量操作简单而准确,加标回收率为97．41％～103．23％,RSD为2．23％;线性范围为2～100 mg/100 mL,检出限为2 mg/100 mL;稳定性及专一性强,不受其他氨基酸的干扰,能够用于氨基酸种类复杂的麦汁赖氨酸的检测。本方法与氨基酸分析仪、DBL、HPLC等方法相比较,测定结果无显著差异（P＞0．05）。     

Determination of cadmium and lead levels in human blood of a general Czech population by GFAAS

The development of a method for the direct determination of cadmium (Cd) and lead (Pb) in blood samples by GFAAS, is described. Samples were properly diluted by a matrix modifier to enable measuring both analytes in one solution. For the determination of Cd, a matrix-matched, and for the determination of Pb, an aqueous calibration was used. The precision, accuracy, and detection limits of this method are presented. A method is applied to the investigation of Cd and Pb levels in a general Czech population, selected according to the WHO-MONICA project criteria. To avoid possible contaminations, samples were treated in a clean room class 100.