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Protein Removal (protein + removal)
Selected AbstractsProtein stabilisation of Chardonnay wine using trisacryl and bentonite: a comparative studyINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 2 2009Johannes De Bruijn Summary The stabilisation of a Chilean Chardonnay wine by SP-Trisacryl-M and bentonite was investigated, evaluating protein, polyphenol and polysaccharide adsorption, turbidity and wine quality. The wine could be stabilised by adding at least 0.3 kg m,3 of bentonite or 12 kg m,3 of trisacryl, removing 95% and 76% of the wine proteins, respectively. The protein adsorption data for bentonite and trisacryl were fitted using the Freundlich isotherm. The wine protein adsorption isotherm on trisacryl was unfavourable. Protein removal from Chardonnay by trisacryl in a packed column at continuous operation was about 50% during the first 70 bed volumes (BV) of treated wine and decreased progressively until the end of the treatment (100 BV). The adsorbents showed a higher selectivity for proteins than for polyphenols and polysaccharides. A sensorial panel could not detect statistically significant differences between the bentonite and trisacryl treatments of wine at P , 0.05. [source] 2-D difference gel electrophoresis of the lung squamous cell carcinoma versus normal sera demonstrates consistent alterations in the levels of ten specific proteinsELECTROPHORESIS, Issue 23 2007Paul Dowling Dr. Abstract Most lung cancers are diagnosed too late for curative treatment to be possible, therefore early detection is crucial. Serum proteins are a rich source of biomarkers and have the potential to be used as diagnostic and prognostic indicators for lung cancer. In order to examine differences in serum levels of specific proteins associated with human lung squamous carcinoma, immunodepletion of albumin and five other high-abundant serum proteins followed by 2-D difference gel electrophoresis (DIGE) analysis and subsequent MS was used to generate a panel of proteins found to be differentially expressed between the cancer and normal samples. Proteins found to have increased abundance levels in squamous cell carcinoma sera compared to normal sera included apolipoprotein A-IV precursor, chain F; human complement component C3c, haptoglobin, serum amyloid A protein precursor and Ras-related protein Rab-7b. Proteins found to have lower abundance levels in squamous cell carcinoma sera compared to normal sera included alpha-2-HS glycoprotein, hemopexin precursor, proapolipoprotein, antithrombin III and SP40; 40. The data presented here demonstrate that high-abundant protein removal combined with 2-D DIGE is a powerful strategy for the discovery of potential biomarkers. The identification of lung cancer-specific biomarkers is crucial to early detection, which in turn could lead to a dramatic increase in survival rates. [source] An effective skeletal muscle prefractionation method to remove abundant structural proteins for optimized two-dimensional gel electrophoresisELECTROPHORESIS, Issue 11 2005Bradley Jarrold Abstract Proteomic analysis of biological samples in disease models or therapeutic intervention studies requires the ability to detect and identify biologically relevant proteins present in relatively low concentrations. The detection and analysis of these low-level proteins is hindered by the presence of a few proteins that are expressed in relatively high concentrations. In the case of muscle tissue, highly abundant structural proteins, such as actin, myosin, and tropomyosin, compromise the detection and analysis of more biologically relevant proteins. We have developed a practical protocol which exploits high-pH extraction to reduce or remove abundant structural proteins from skeletal muscle crude membrane preparations in a manner suitable for two dimensional gel electrophoresis. An initial whole-cell muscle lysate is generated by homogenization of powdered tissue in Tris-base. This lysate is subsequently partitioned into a supernatant and pellet containing the majority of structural proteins. Treatment of the pellet with high-pH conditions effectively releases structural proteins from membrane compartments which are then removed through ultracentrifugation. Mass spectrometric identification shows that the majority of protein spots reduced or removed by high-pH treatment were contractile proteins or contractile-related proteins. Removal of these proteins enabled successful detection and identification of minor proteins. Structural protein removal also results in significant improvement of gel quality and the ability to load higher amounts of total protein for the detection of lower abundant protein classes. [source] Fluoride down-regulates the expression of matrix metalloproteinase-20 in human fetal tooth ameloblast-lineage cells in vitroEUROPEAN JOURNAL OF ORAL SCIENCES, Issue 2006Yan Zhang Fluoride is associated with a decrease in the incidence of dental caries, but excessive fluoride intake during tooth enamel formation can result in enamel fluorosis. Fluorosed enamel has increased porosity, which has been related to a delay in the removal of amelogenin proteins as the enamel matures. This delay in protein removal suggests that fluoride may affect either the amount or the activity of enamel matrix proteinases. In this study, we investigated the role of fluoride in the synthesis and secretion of matrix metalloproteinase-20 (MMP-20), the proteinase primarily responsible for the initial hydrolysis of amelogenin during the secretory stage of enamel formation. Cultured human fetus tooth organ ameloblast-lineage cells were exposed to 10 µM fluoride and analyzed for synthesis of MMP-20. Immunoblotting showed that 10 µM NaF down-regulated the synthesis of MMP-20 by 21% compared with control cells, but did not alter the amount of amelogenin or kalikrein-4 (KLK-4) synthesized by the cells. Real-time polymerase chain reaction (PCR) showed that 10 µM NaF down-regulated MMP-20 mRNA expression to 28% of the levels found in the non-treated cells. These in vitro results suggest that fluoride can alter the expression of MMP-20 by ameloblasts, resulting in a disturbance of the balance between MMP-20 and its substrate that may contribute to the retention of amelogenins in the formation of fluorosed enamel. [source] The use of muscle protein for egg production in the Zebra Finch Taeniopygia guttataIBIS, Issue 2 2002Mat Cottam Pectoral muscle can be an important source of protein for birds. During egg formation Zebra Finches Taeniopygia guttata are able to compensate for nutritional inadequacies in their diet by utilization of the protein in their flight muscles. This analysis of flight muscle sarcoplasm supported earlier observations of protein depletion during egg production. However, SDS gel electrophoresis of the sarcoplasm produced no evidence to support a previous suggestion of the existence of a high molecular weight storage protein, and it is thought that the original observation may have arisen as an artefact of experimental methodology. During laying, protein removal from the sarcoplasm occurred over a range of different proteins and was not confined to any one specific protein band. Additionally, the protein band most reduced over the course of laying did not contain elevated levels of the amino acids most limiting to egg production. These results indicate that during laying, flight muscle sarcoplasm contributes towards the nutrient requirements of egg production from general protein reserves, rather than from a specific storage protein containing elevated levels of limiting amino acids. [source] Optimization of the extraction and purification of oligosaccharides from defatted soybean mealINTERNATIONAL JOURNAL OF FOOD SCIENCE & TECHNOLOGY, Issue 3 2003Suna Kim Summary Optimization of the extraction of oligosaccharides from defatted soybean meal (DSM) was investigated under various conditions. The optimal ratio of water to DSM and optimal temperature for oligosaccharide extraction was 5:1 and 50 °C, respectively. The use of a stirring process, without any further grinding, improved the extractability of oligosaccharides from DSM, a 10% ethanol-water solution was more effective than distilled water alone. To purify the oligosaccharides, ultrafiltration was used. More than 90% of the protein was removed from the extracts at a volume concentration ratio (VCR) of 3,5. The percentages of fructose, sucrose, raffinose and stachyose in permeate for a VCR of 5 were 38.6, 51.4, 54.2 and 52.6%, respectively. A VCR of 5 was the most effective for protein removal and recovery of oligosaccharides. [source] Differences among techniques for high-abundant protein depletionPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 13 2005Nina Zolotarjova Abstract The need to identify protein or peptide biomarkers via readily available biological samples like serum, plasma, or cerebrospinal fluid is often hindered by a few particular proteins present at relatively high concentrations. The ability to remove these proteins specifically, reproducibly, and with high selectivity is increasingly important in proteomic studies, and success in this procedure is leading to an ever-increasing list of lower abundant proteins being identified in these biological fluids. The current work addresses some of the potential problems in depleting proteins in typical biomarker studies, including nonspecific binding during depletion procedures and whether low molecular weight (LMW) species bind to the column in a so-called "sponge" effect caused by the ability of albumin or other high-abundant proteins to bind peptides or protein fragments. LC-MS/MS methods were applied to the comparative analysis of an IgG-based immunodepletion method and a Cibacron blue (CB)-dye-based method, for specificity of removing targeted proteins (binding fraction), as well as for assessing efficiency of target removal. This analysis was extended to examine the effects of repeated use of materials (cycles of binding and elution), in order to assess potential for carryover of one sample to the next. Capacity studies and efficiency of protein removal from the serum samples were followed for the IgG-based system using both immunochemical assays (ELISA) as well as LC-MS/MS methods. Additionally, the IgG-based system was further characterized for the removal of LMW polypeptides by nonspecific binding. We conclude that the IgG-based system provided effective removal of targeted proteins, with minimal carryover, high longevity, and minimal nonspecific binding. Significant differences are noted between the depletion techniques employed, and this should be considered based on the expectations set during experimental design. [source] Immunoaffinity separation of plasma proteins by IgY microbeads: Meeting the needs of proteomic sample preparation and analysisPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 13 2005Lei Huang Abstract Separation of complex protein mixtures that have a wide dynamic range of concentration, such as plasma or serum, is a challenge for proteomic analysis. Sample preparation to remove high-abundant proteins is essential for proteomics analysis. Immunoglobulin yolk (IgY) antibodies have unique and advantageous features that enable specific protein removal to aid in the detection of low-abundant proteins and biomarker discovery. This report describes the efficiency and effectiveness of IgY microbeads in separating 12 abundant proteins from plasma with an immunoaffinity spin column or LC column. The protein separation and sample preparation process was monitored via SDS-PAGE, 2-DE, LC-MS/MS, or clinical protein assays. The data demonstrate the high specificity of the protein separation, with removal of 95,99.5% of the abundant proteins. IgY microbeads against human proteins can also selectively remove orthologous proteins of other mammals such as mouse, rat, etc. Besides the specificity and reproducibility of the IgY microbeads, the report discusses the factors that may cause potential variations in protein separation such as protein,protein interactions (known as "Interactome"), binding and washing conditions of immunoaffinity reagents, etc. A novel concept of Seppromics is introduced to address methodologies and science of protein separation in a context of proteomics. [source] Matching Efficacy of Online Hemodiafiltration in Simple Hemodialysis ModeARTIFICIAL ORGANS, Issue 12 2008Detlef H. Krieter Abstract PUREMA H (referred to as PES) is an innovative dialysis membrane for enhanced low-molecular-weight (LMW) protein removal. The purpose of the study was to prove whether its efficacy in hemodialysis (HD) matches that of online hemodiafiltration (HDF) with conventional high-flux membranes. In a prospective, randomized, cross-over study on eight maintenance dialysis patients, treatment efficacy of HD with PES was compared with online postdilution HDF with the two synthetic high-flux membranes polysulfone (referred to as PSU) and Polyamix (referred to as POX). Apart from the infusion of replacement fluid, which was set at 20% of the blood flow rate of 300 mL/min, operating conditions in HD and HDF were kept identical. Small solute and LMW protein plasma clearances as well as the reduction ratio (RR) of cystatin C and retinol-binding protein were not different between the therapies. HDF with POX resulted in a significantly lower myoglobin RR as compared with HD with PES, and HDF with PSU. A 4% higher beta2 -microglobulin RR was determined in HDF with PSU (73 ± 5%) as compared with PES in HD (69 ± 5%). The albumin loss was below 1 g for all treatments. Despite the fact that simple HD did not fully exploit the characteristics of PES, it achieved essentially similar LMW protein removal and albumin loss as compared with online postdilution HDF with the conventional synthetic high-flux membranes PSU and POX. Therefore, HD with PES may have beneficial effects on the outcome of maintenance dialysis patients similar to high-efficiency HDF. [source] | ||||||||||||||||