Elution Method (elution + method)

Distribution by Scientific Domains

Selected Abstracts

Microaffinity purification of proteins based on photolytic elution: Toward an efficient microbead affinity chromatography on a chip

Woo-Jae Chung
Abstract A bead affinity chromatography system, which was based on the photolytic elution method, was integrated into a glass-silicon microchip to purify specific target proteins. CutiCore® beads, which were coupled with a photo-cleavable ligand, such as biotin and an RNA aptamer, were introduced into a filter chamber in the microchip. The protein mixture containing target protein labeled with fluorescein isothiocyanate (FITC) was then passed through the packed affinity beads in the microchamber by pressure-driven flow. During the process, the adsorbed protein on the bead was monitored by fluorescence. The concentrated target protein on the affinity bead was released by simple irradiation with UV light at a wavelength of 360 nm, and subsequently eluted with the phosphate buffer flow. The eluted target protein was quantitatively detected via the fluorescence intensity measurements at the downstream of the capillary connected to the outlet of the microchip. The microaffinity purification allowed for a successful method for the identification of specific target proteins from a protein mixture. In addition, the feasibility of this system for use as a diagnosis chip was demonstrated. [source]

Assessment of norovirus contamination in environmental samples from Florianópolis City, Southern Brazil

M. Victoria
Abstract Aims:, To assess norovirus (NoV) contamination in aquatic ecosystems in the city of Florianópolis, in Southern Brazil, to provide epidemiological data that can support actions for environmental contamination control. Methods and Results:, An adsorption,elution method, followed by ultrafiltration, was performed to concentrate the viruses. NoV were detected using semi-nested PCR and quantified by real-time PCR. From June 2007 to May 2008, NoV were detected in 23% (22/94) of the samples analysed, including seawater, drinking water, superficial water (creek and brackish lagoon) and treated sewage. The mean viral loads for genogroups (G)I and GII in treated sewage samples were 297 and 440 genomic copies (gc) l,1, respectively, whereas creek water samples contained 2603 and 1361 gc l,1, respectively. Six samples were sequenced: two samples were GII.4, two were GII.2 and two were GI.3. Conclusions:, NoV were detected in all water types analysed, demonstrating the widespread contamination of this geographical area with several cocirculating strains belonging to GI and GII. Significance and Impact of the Study:, This study demonstrates the environmental spread of NoV in environmental waters and highlights the potential hazard for human health following the consumption of or contact with these waters, which could result in waterborne or foodborne acute gastroenteritis. [source]

Protein expression pattern of P,glycoprotein along the gastrointestinal tract of the yucatan micropig

Huadong Tang
Abstract The purpose of this study is to characterize the distribution pattern of P,gp protein levels along the entire GI tract in the Yucatan micropig, which is being developed as a model for human drug bioavailability. Small and large intestines were freshly obtained and divided into about 37 segments and 10 segments, respectively (ca., 1 foot/segment). Epithelial cells from the small intestine were obtained by an elution method; whereas, a scraping method was applied to the large intestine. Total cellular protein was isolated from the epithelial cells. Western blot analysis using P,gp antibody showed that the amount of P,gp protein increased distally from the duodenum to the ileum over approximately a 10,fold range. P,gp protein in the large intestine was present at a higher level in the central portion, but the absolute amount was much less than what was found in the small intestine. © 2004 Wiley Periodicals, Inc. J Biochem Mol Toxicol 18:18,22, 2004; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20001 [source]

Technical Approach to Simplify the Purification Method and Characterization of Microbial Transglutaminase Produced from Streptoverticillium ladakanum

M.-L. Ho
ABSTRACT: In order to fast and economically purify MTGase from Streptoverticillium ladakanum, a stepwise elution method was developed and compared with linear gradient elution method. MTGase was purified to electrophoretical homogeneity by using CM Sepharose CL-6B and Blue Sepharose Fast Flow chromatographies by linear gradient or stepwise methods. The recovery of MTGase by linear gradient and stepwise methods were 68.4% and 81.0%, respectively. The optimal temperature and pH were 40 °C and 5.5, respectively. It was stable at pH 5.0 to 7.0 and had a rate constant (KD) of 6.21 °o 10 -5 min -1 for thermal inactivation at 45 °C. The purified MTGase was activated by K+ Na+, Ca2+, Mn2+, and Mg2+, not affected by Fe3+, EDTA, but inhibited by Cu2+, Zn2+, Hg2+, Ni2+, Co2+, Cd2+, PCMB, NEM, IAA, and PMSF. A simple stepwise method was developed for the purification of MTGase from S. ladakanum. [source]