化学发光单孵多层免疫技术检测粪样中轮状病毒

以酶联免疫吸附分析（ＥＬＩＳＡ）夹心测定粪样中轮状病毒（ＲＶ）实验为模型，将传统又孵式免疫操作变成单孵式，即在包被ＲＶ抗体Ａｂ的微孔中同时加入含ＲＶ的粪样上清液和根过氧化物酶（ＨＲＰ）标记抗ＲＶ抗原共同孵育，其结果在载体表面形成多层酶免疫复合物，以ＨＲＰ催化鲁米诺－对碘苯酚－过氧化氢化学体系作为最终信号检测系统，从而建立起化学发光单孵多层免疫技术（ＣＬ－ＳＩＭＩＴ）。其灵敏度是ＥＬＩＳＡ的１６倍，     

Chemical composition,antioxidant and cytotoxic activities of extracts from the leaves of Smilax brasiliensis Sprengel (Smilacaceae)

The antioxidant and cytotoxic activities of petroleum ether and methanol extracts, fatty acids and methyl esters from leaves of Smilax brasiliensis were evaluated, and the composition of the extracts was determined. Palmitic, linoleic and linolenic acids were major components of the extracts. For antioxidant activity, all samples exhibited IC₅₀ values lower than BHT (2,6-di-tert-butyl-4-methylphenol). The extracts, fatty acids and methyl esters from S. brasiliensis presented no toxicity to larvae of the brine shrimp, Artemia salina. Among the purified substances, only methyl linolenate showed toxicity (LD₅₀ = 21.47 μg/mL). This study showed, for the first time, the composition of petroleum ether and methanol extracts from S. brasiliensis leaves, as well as the antioxidant and cytotoxic activities of extracts, fatty acids and methyl esters.     

Spectrophotometric Label-Free Determination of Lead Using Thiol-Functionalized Gold Nanoparticles

A method for the determination of lead is described using thiol-functionalized gold nanoparticle. The detection method is based on the prevention of thiol-induced aggregation of gold nanoparticles by lead. Among six thiols, e.g., 4-mercapto-1-butanol, meso-2, 3-dimercaptosuccinic acid, mercaptosuccinic acid, 6-mercapto-1-hexanol, 4-(methylthio)-1-butanol, 1-propanethiol, four (4-mercapto-1-butanol, 6-mercapto-1-hexanol, 4-(methylthio)-1-butanol and 1-propanethiol) induced the aggregation of the gold nanoparticles which was measured by the change in absorbance at 520 and 650?nm. Prior incubation of the gold nanoparticles with lead decreased the 4-(methylthio)-1-butanol-induced aggregation of gold nanoparticles in a dose-dependent manner. A linear inverse relationship between the logarithmic concentration of lead and the ratio of absorbance at 650 to 520 was noted. The method has a dynamic range from 10?nM to 100?µM. However, metals such as mercury and chromium were more effective in comparison with lead in preventing the 4-methylthio-1-butanol-induced aggregation of gold nanoparticles. The method can be used for assessing the heavy metal load in water samples.     

Applying green analytical chemistry for development and validation of RP-HPLC stability indicating assay method for estimation of fenoverine in bulk and dosage form using quality by design approach

A green and robust reverse-phase liquid chromatographic method has been developed for the determination of fenoverine (FEN), by applying combined principles of green analytical chemistry and quality by design approaches on a Spherisorb C18 column (150?×?4.6?mm, 3?µm) with UV detection at 262?nm. A two level fractional factorial design (2^^7-3) Res IV was used for screening of influential chromatographic factors. The critical method parameters actively affecting critical quality attributes (CQAs) were identified and further optimized using Box–Behnken design. The predicted optimum assay conditions comprised of methanol and ammonium acetate buffer 20?mM, in an extent of 81:19% v/v individually having a flow rate of 1.0?mL/min with a column oven temperature of 33°C. The drug was stressed in hydrolytic, oxidative, reductive, thermal, and photolytic conditions. The developed method was validated successfully. The detector response was linear in the concentration of 0.5–160?µg/mL with a limit of detection (LOD) and limit of quantitation (LOQ) as 0.1 and 0.3?µg/mL, respectively. The % recovery was found to be 99.7%. The analytical method volume intensity value for developed method was 45?mL and the environment assessment tool (EAT) score was 41.07. The method is simple, environmentally benign, rapid, and robust for the determination of FEN in bulk and in its dosage form.     

A novel efficient medium for chromogenic catalysis of tetramethylbenzidine with horseradish peroxidase

The traditional surfactant sodium dodecyl sulfate (SDS) and ionic liquid 1-ethyl-3-methylimidazolium tetrafluoroborate ([Emim][BF₄]) have been combined to create a novel efficient medium for chromogenic catalysis of 3,3',5,5'-tetramethylbenzidine with horseradish peroxidase in presence of H₂O₂. The results have shown the [Emim][BF₄] in the mediums can promote the rate of formation of the blue chromogen, the SDS is responsible for the stabilization of the blue chromogen due to the electrostatic attraction between positively charged blue chromogen and the negatively charged surfactant. The SDS/[Emim][BF₄] combination not only enhance catalytic activity of HRP remarkably but also stabilize the blue chromogen formed in the HRP oxidation of the substrate TMB compared to the conventional medium. Based on the superior combination of SDS and [Emim][BF₄], the colorimetric assay for detecting HRP activity and H₂O₂ concentration was established. This work demonstrates a novel efficient medium for chromogenic catalysis with potential applications in biosensors and clinical diagnosis.     

Determination of Immunoglobulin G by a Hemin–Manganese(IV) Oxide-Labeled Enzyme-linked Immunosorbent Assay

An enzyme-linked immunosorbent assay (ELISA) is reported for human immunoglobulin G based on synthesized hemin–MnO₂ nanocomposite as the label. Enhanced sensitivity was obtained due to the increased catalytic activity of the hemin–MnO₂ nanocomposite toward 3,3′,5,5′-tetramethylbenzidine compared to hemin and MnO₂ alone. The synthesized hemin–MnO₂ nanocomposite was characterized by transmission electron microscopy and its catalytic activity to 3,3′,5,5′-tetramethylbenzidine was investigated by ultraviolet–visible absorption spectroscopy. After assembly of the sandwich-type immunoassay in the 96 wells of the plate the hemin–MnO₂-based label catalyzed 3,3′,5,5′-tetramethylbenzidine into blue compounds that were monitored by a plate reader. The absorbance increased with the concentration of human immunoglobulin G. The immunoassay displayed high sensitivity, a long linear dynamic range, and good selectivity for human immunoglobulin G. The immunoassay was also used for the determination of human immunoglobulin G in serum with favorable results. The developed assay combines the high throughput and low cost of ELISA with the simplicity of nanocomposite labeling and is suitable for application in clinical diagnosis.     

A spectrophotometric assay for solid phase primary amino groups

A rapid, sensitive, and convenient spectrophotometric assay was developed for the measurement of amino groups on solid supports. This method is based on the reaction of amino groups of solids with an excess ofo-phthaldialdehyde (OPA) and subsequent quantitative determination of unreacted OPA by reaction with glycine. Four solids possessing variable quantities of amino groups were examined. Results indicate that about 70% of the total surface amino concentration (determined by the microKjeldahl method) are available for ligand attachment. Unlike the spectrophotometric 2,4,6-trinitrobenzenesulfonic acid method, the OPA spectrophotometric assay is more rapid, sensitive, and convenient, and unlike the spectrofluorimetric OPA, it does not require sophisticated instrumentation.     

初发系统性红斑狼疮患者外周血清IL-17的表达

目的探讨IL-17在初发系统性红斑狼疮（SLE）患者外周血清中的表达水平及其临床意义。方法选择42例初次诊断、未经治疗的SLE患者（SLE组）及26例同期健康志愿者（对照组）,采用双抗体夹心酶联免疫吸附试验（ELISA）方法测定外周血清IL-17水平,同时检测外周血抗ds- DNA等抗体及免疫球蛋白、补体等水平并进行相关分析。结果活动期和缓解期SLE患者血清IL-17水平均较对照组明显升高,差异均有统计学意义（均P＜0.05）。活动期和缓解期SLE患者间血清IL-17水平的差异无统计学意义（P＞0.05）。具有皮肤损害的SLE患者血清IL-17水平较无皮肤损害者高,使用环磷酰胺的SLE患者血清IL-17水平较未使用环磷酰胺者高,差异有统计学意义（P＜0.05）。SLE患者血清IL-17水平与SLEDAI、血沉、CRP、IgG、C3、C4均无相关性（均P＞0.05）。19例SLE活动期患者治疗1个月后血清IL-17水平较治疗前明显下降,差异有统计学意义（P＜0.05）。结论 IL-17与狼疮患者的皮肤损害密切相关;IL-17在SLE的发病中起到重要作用并与疾病严重性有关。     

Dipyrrylmetheneboron Difluorides as Labels in Two-Photon Excited Fluorometry. Part II–Nucleic Acid Hybridization Assays

Five two-photon excitable dipyrrylmetheneboron difluoride labels (dipyrrylmethene-BF₂ labels) with fluorescence emission maximum between 530 and 590 nm, and a frequently used rhodamine label, TAMRA, were conjugated to aminomodified oligonucleotides. The performance of the labeled oligonucleotides was studied in a separation-free nucleic acid hybridization assay using ArcDia^™ TPX bioaffinity assay technology. The results show that oligonucleotide conjugates of dipyrrylmethene-BF₂ labels provide higher two-photon excited fluorescence yield and better assay sensitivity than corresponding TAMRA conjugate. The effect of conjugation on photophysical properties of the labels and performance of the labeled oligonucleotides in separation-free hybridization assay is discussed.