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Enrichment Methods (enrichment + methods)

Distribution by Scientific Domains

Chemistry	80%

Selected Abstracts

A Rapid Method for the Pre-Enrichment and Detection of Salmonella Typhimurium by Immunomagnetic Separation and Subsequent Fluorescence Microscopical Techniques

ENGINEERING IN LIFE SCIENCES (ELECTRONIC), Issue 3 2005
J. Steingroewer
Abstract Detection of food-borne pathogens is of great importance in order to minimize the risk of infection for customers. These analyses should be as fast as possible. Any detection method requires enrichment and quantitative analysis of the enriched microbes. Conventional enrichment methods, which take several days, need to be replaced by faster techniques such as immunomagnetic separation (IMS). This technique is based on the use of paramagnetic microspheres coated with antibodies as ligands that have specific affinity to the microbes that have to be detected. In the studies reported here, a rapid method for the detection of Salmonella enterica serovar Typhimurium (Salmonella Typhimurium), combining IMS and Direct Epifluorescence Filter Technique (DEFT), was developed. It was focused on releasing the target cells from the magnetic beads after IMS, because this is a premise for combining IMS, as an alternative pre-enrichment, with DEFT. Otherwise, the high number of beads form a layer on the filter membrane that makes the following microscopic analysis for the detection of the contaminants impossible. The CELLectionTM Dynabeads® used in this study, are coated with recombinant streptavidin (rSA) via a DNA linker. The rSA binds biotinylated antibodies that are able to capture target cells. The DNA linker provides the cleavable site, so that the beads can be removed from the captured cells after isolation. In this study a releasing procedure was developed. This procedure allows for an average 74,% ± 4,% of the bead-bound Salmonella Typhimurium cells to be released from the beads after IMS, so that the detection of the separated cells by DEFT will be possible. [source]

OCCURRENCE OF HEMOLYSIN-PRODUCING AEROMONADS IN MEAT AND OFFAL SOLD IN PORT HARCOURT, NIGERIA

JOURNAL OF FOOD SAFETY, Issue 3 2005
E.N. AMADI
ABSTRACT Fresh, different meat types and offal were examined for the occurrence of Aeromonas species by the direct-plating and enrichment methods. The enrichment method (coefficient of variation = 15.1%) enhanced the recovery of Aeromonas species. The major Aeromonas species identified were A. sobria (67.3%) and A. hydrophila (21.2%). Other species occurred in minor proportions and were A. caviae, A. proteolytica and A. salmonicida. Out of the 52 isolates, 50 were motile species except two which were not motile and identified as the species A. proteolytica. Sixteen motile species occurred in cow, 18 in goat and 16 in lamb. The motile species and the two nonmotile species were hemolytic. They were all sensitive to chloramphenicol (30 mg) and resistant to ampicillin (10 mg). The nonmotile A. proteolytica were all resistant to tetracycline. Aeromonas caviae, A. salmonicida, A. hydrophila and A. sobria were killed after exposure to 50C (decimal reduction time, D10 = 30 s). Aeromonads are unlikely to pose a public health problem in Nigeria where meat undergoes prolonged cooking. Meat is a possible factor in the epidemiology of Aeromonas -associated gastroenteritis in man. [source]

Techniques for phosphopeptide enrichment prior to analysis by mass spectrometry

MASS SPECTROMETRY REVIEWS, Issue 1 2010
Jamie D. Dunn
Abstract Mass spectrometry is the tool of choice to investigate protein phosphorylation, which plays a vital role in cell regulation and diseases such as cancer. However, low abundances of phosphopeptides and low degrees of phosphorylation typically necessitate isolation and concentration of phosphopeptides prior to MS analysis. This review discusses the enrichment of phosphopeptides with immobilized metal affinity chromatography, reversible covalent binding, and metal oxide affinity chromatography. Capture of phosphopeptides on TiO2 seems especially promising in terms of selectivity and recovery, but the success of all methods depends on careful selection of binding, washing, and elution solutions. Enrichment techniques are complementary, such that a combination of methods greatly enhances the number of phosphopeptides isolated from complex samples. Development of a standard series of phosphopeptides in a highly complex mixture of digested proteins would greatly aid the comparison of different enrichment methods. Phosphopeptide binding to magnetic beads and on-plate isolation prior to MALDI-MS are emerging as convenient methods for purification of small (µL) samples. On-plate enrichment can yield >70% recoveries of phosphopeptides in mixtures of a few digested proteins and can avoid sample-handling steps, but this technique is likely limited to relatively simple samples such as immunoprecipitates. With recent advances in enrichment techniques in hand, MS analysis should provide important insights into phosphorylation pathways. © 2009 Wiley Periodicals, Inc., Mass Spec Rev 29:29,54, 2010 [source]

Affinity prefractionation for MS-based plasma proteomics

PROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 6 2009
Maria Pernemalm
Abstract The plasma proteome has proven to be one of the most challenging proteomes to profile using currently available proteomics technologies. A plethora of methodologies have been used to profile human plasma in order to discover potential biomarkers for disease and for therapy optimization. Affinity-based prefractionation coupled to MS has been shown to be one of the most successful ways to dig deeper into the plasma proteome. Depletion of high abundant plasma proteins is becoming an initial method of choice in any plasma profiling project. However, several other affinity-based enrichment methods have been published in recent years. Here we review both protein and peptide affinity prefractionation methods coupled with MS-based proteomics. Analysis of the proportion of cellular and extracellular annotated proteins of publicly available MS plasma proteomics data is performed to estimate the analytical depth of various prefractionation methods. [source]

Large-scale phosphoproteome analysis of human liver tissue by enrichment and fractionation of phosphopeptides with strong anion exchange chromatography

PROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 7 2008
Guanghui Han
Abstract The mixture of phosphopeptides enriched from proteome samples are very complex. To reduce the complexity it is necessary to fractionate the phosphopeptides. However, conventional enrichment methods typically only enrich phosphopeptides but not fractionate phosphopeptides. In this study, the application of strong anion exchange (SAX) chromatography for enrichment and fractionation of phosphopeptides was presented. It was found that phosphopeptides were highly enriched by SAX and majority of unmodified peptides did not bind onto SAX. Compared with Fe3+ immobilized metal affinity chromatography (Fe3+ -IMAC), almost double phosphopeptides were identified from the same sample when only one fraction was generated by SAX. SAX and Fe3+ -IMAC showed the complementarity in enrichment and identification of phosphopeptides. It was also demonstrated that SAX have the ability to fractionate phosphopeptides under gradient elution based on their different interaction with SAX adsorbent. SAX was further applied to enrich and fractionate phosphopeptides in tryptic digest of proteins extracted from human liver tissue adjacent to tumorous region for phosphoproteome profiling. This resulted in the highly confident identification of 274 phosphorylation sites from 305 unique phosphopeptides corresponding to 168 proteins at false discovery rate (FDR) of 0.96%. [source]