Development of a C12Im-Cl-assisted method for rapid sample preparation in proteomic application

2021 ◽  
Author(s):  
Chang Liu ◽  
Xiaoxia Si ◽  
Shumei Yan ◽  
Xinyuan Zhao ◽  
Xiaohong Qian ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Protein Extraction ◽  
Sample Processing ◽  
Processing Methods ◽  
Proteomic Analyses

Chromatography and mass spectrometry (MS) techniques have greatly improved the power of proteomic analyses. However, sample processing methods, including protein extraction and digestion, before MS remain as bottlenecks in the...

scholarly journals Evaluation of sample processing methods for the polar contaminant analysis of sewage sludge using liquid chromatography - mass spectrometry (LC/MS)

Química Nova ◽  
2010 ◽  
Vol 33 (5) ◽  
pp. 1194-1198 ◽  
Author(s):  
Rênnio F. de Sena ◽  
José L. Tambosi ◽  
Regina F. P. M. Moreira ◽  
Humberto J. José ◽  
Wilheim Gebhardt ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Liquid Chromatography ◽  
Sewage Sludge ◽  
Liquid Chromatography Mass Spectrometry ◽  
Sample Processing ◽  
Processing Methods ◽  
Chromatography Mass Spectrometry

scholarly journals Progress in Radiocarbon Target Preparation at the Antares AMS Centre

Radiocarbon ◽  
2001 ◽  
Vol 43 (2A) ◽  
pp. 275-282 ◽  
Author(s):  
Q Hua ◽  
G E Jacobsen ◽  
U Zoppi ◽  
E M Lawson ◽  
A A Williams ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Accelerator Mass Spectrometry ◽  
Sample Pretreatment ◽  
Small Samples ◽  
Sample Processing ◽  
Target Preparation ◽  
Recent Developments ◽  
Technical Details ◽  
Fe Reduction

We present routine methods of target preparation for radiocarbon analysis at the ANTARES Accelerator Mass Spectrometry (AMS) Centre, as well as recent developments which have decreased our procedural blank level and improved our ability to process small samples containing less than 200 μg of carbon. Routine methods of 14C sample preparation include sample pretreatment, CO2 extraction (combustion, hydrolysis and water stripping) and conversion to graphite (graphitization). A new method of cleaning glassware and reagents used in sample processing, by baking them under a stream of oxygen, is described. The results show significant improvements in our procedural blanks. In addition, a new graphitization system dedicated to small samples, using H2/Fe reduction of CO2, has been commissioned. The technical details of this system, the graphite yield and the level of fractionation of the targets are discussed.

Comparison of Sample Preparation Methods for Recovering Salmonella from Raw Fruits, Vegetables, and Herbs

2001 ◽  
Vol 64 (10) ◽  
pp. 1459-1465 ◽  
Author(s):  
ANDREA B. BURNETT ◽  
LARRY R. BEUCHAT
Keyword(s):  
Sample Preparation ◽  
Pathogenic Bacteria ◽  
Preparation Method ◽  
Morphological Characteristics ◽  
Processing Method ◽  
Sample Processing ◽  
Processing Methods ◽  
Preparation Methods ◽  
Wide Range ◽  
The Mean

Methods for preparing raw fruits, vegetables, and herbs for enrichment or direct plating to determine the presence and populations of pathogenic bacteria vary greatly. A study was done to compare three sample processing methods (washing in 0.1% peptone, stomaching, and homogenizing) for their influence on recovery of Salmonella inoculated onto 26 types of raw produce. The mean numbers of Salmonella recovered from 10 fruits, 11 vegetables, and 5 herbs using all three processing methods were 7.17, 7.40, and 7.27 log10 CFU/sample, respectively. Considering all 26 types of produce and all processing methods, the number of Salmonella recovered ranged from 7.24 to 7.29 log10 CFU/sample, with no significant differences attributable to a particular sample processing method. Mean percent recoveries of Salmonella from washed, stomached, and homogenized produce were 39.4, 44.7, and 42.4%, respectively. Mean percent recoveries from fruits, vegetables, and herbs, regardless of sample preparation method, were 41.7, 50.1, and 25.9%, respectively. The number of Salmonella recovered from stomached and homogenized produce, but not washed produce, with pH ≤ 4.53 was significantly less than the number recovered from produce with pH from 5.53 to 5.99, suggesting that the acidic environment in stomachates and homogenates was lethal to a portion of Salmonella. Reduced percent recoveries from herbs (pH 5.94 to 6.34) is attributed, in part, to antimicrobials released from plant cells during sample preparation. Overall, the type of processing method did not substantially affect the number of Salmonella recovered from the 26 types of raw produce representing a wide range of structural and morphological characteristics, composition, and pH. The influence of sample size, diluent composition, and processing time on efficiency of recovery of Salmonella and other pathogens needs to be evaluated before a method(s) for processing samples of raw produce can be recommended.

scholarly journals Gel Electrophoresis/Electroelution Sorting Fractionator combined with Filter Aided Sample preparation (FASP) for deep proteomic analysis

2021 ◽  
Author(s):  
Yassel Ramos ◽  
Alexis Almeida ◽  
Jenis Carpio ◽  
Arielis Rodríguez-Ulloa ◽  
Yasser Perera ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Protein Identification ◽  
Protein Extraction ◽  
Complex Mixture ◽  
Fold Increase ◽  
Mass Spectrometry Analysis ◽  
Protein Fractionation ◽  
Spectrometry Analysis ◽  
Sds Page

AbstractSample preparation and protein fractionation are important issues in proteomic studies in spite of the technological achievements on protein mass spectrometry. Protein extraction procedures strongly affect the performance of fractionation methods by provoking protein dispersion in several fractions. The most notable exception is SDS-PAGE-based protein fractionation due to its extraordinary resolution and the effectiveness of SDS as a solubilizing agent. Its main limitation lies in the poor recovery of the gel-trapped proteins, where protein electro-elution is the most successful approach to overcome this drawback. We created a device to separate complex mixture of proteins and peptides (named “GEES fractionator”) that is based on the continuous Gel Electrophoresis/Electro-elution Sorting of these molecules. In an unsupervised process, complex mixtures of proteins or peptides are fractionated into the gel while separated fractions are simultaneously and sequentially electro-eluted to the solution containing wells. The performance of the device was studied for SDS-PAGE-based protein fractionation in terms of reproducibility, protein recovery and loading capacity. In the SDS-free PAGE setup, complex peptide mixtures can also be fractionated. More than 11 700 proteins were identified in the whole-cell lysate of the CaSki cell line by using the GEES fractionator combined with the Filter Aided Sample Preparation (FASP) method and mass spectrometry analysis. GEES-based proteome characterization shows a 1.7 fold increase in the number of identified proteins compared to the unfractionated sample analysis. Proteins involved in the co-regulated transcription activity, as well as cancer related pathways such as apoptosis signaling, P53 and RAS pathways are more represented in the protein identification output of GEES-based fractionation approaches.

scholarly journals Sample Preparation by Easy Extraction and Digestion (SPEED) - A Universal, Rapid, and Detergent-free Protocol for Proteomics based on Acid Extraction

2018 ◽  
Author(s):  
Joerg Doellinger ◽  
Andy Schneider ◽  
Marcell Hoeller ◽  
Peter Lasch
Keyword(s):  
Sample Preparation ◽  
Protein Extraction ◽  
Spectrometric Analysis ◽  
Acid Extraction ◽  
Sample Processing ◽  
Special Equipment ◽  
One Pot ◽  
Tissue Samples ◽  
Main Challenge ◽  
Speed Performance

SUMMARYMass spectrometry is the method of choice for deep and comprehensive analysis of proteomes and has become a key technology to support the progress in life science and biomedicine. However, sample preparation in proteomics is not standardized and contributes to a lack of reproducibility. The main challenge is to extract all proteins in a manner that enables efficient digestion into peptides and is compatible with subsequent mass spectrometric analysis. Current methods are based on the idea of removing detergents or chaotropic agents during sample processing, which are essential for protein extraction but interfere with digestion and LC-MS. These multi-step preparations are prone to losses, biases and contaminations, while being time-consuming and labor-intensive.We report a universal detergent-free method, named Sample Preparation by Easy Extraction and Digestion (SPEED), which is based on a simple three-step procedure, acidification, neutralization and digestion. SPEED is a one-pot method for peptide generation from various sources and is easily applicable even for lysis-resistant sample types as pure trifluoroacetic acid (TFA) is used for highly efficient protein extraction. SPEED-based sample processing is highly reproducible, provides exceptional peptide yields and enables preparation even of tissue samples with less than 15 min hands-on time and without any special equipment. Evaluation of SPEED performance revealed, that the number of quantified proteins and the quantitative reproducibility are superior compared to the well-established sample processing protocols FASP, ISD-Urea and SP3 for various sample types, including human cells, bacteria and tissue, even at low protein starting amounts.

scholarly journals NEW OPPORTUNITIES TO IDENTIFY AND TYPE STAPHYLOCOCCUS spp. BY USING MALDI-TOF MASS SPECTROMETRY

2019 ◽  
Vol 8 (4) ◽  
pp. 489-496
Author(s):  
A. S. Stepanov ◽  
N. V. Vasilyeva
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Protein Extraction ◽  
Mass Range ◽  
Intensity Level ◽  
Immunocompromised Patients ◽  
Microbial Identification ◽  
Maldi Tof Mass Spectrometry ◽  
Maldi Tof ◽  
Staphylococcus Spp

Abstract.Mass spectrometry profiles of microorganisms obtained by time-of-flight matrix-associated laser desorption/ionization (MALDI-TOF) mass spectrometry are a source of information about peptide profiles can be used for microbial identification and typing. A variety of technical and bioinformational solutions complicate developing of a united mass-spectro-profile database. Staphylococcus spp. strains are good studied objects for identification by MALDI-TOF mass spectrometry, frequently resulting in nosocomial infections, especially in immunocompromised patients. Rapid differentiation of nosocomial, multiresistant and highly virulent isolates of Staphylococcus spp. Allows to reduce the lethality in weakened and immunocompromised patients. The study was aimed at assessing comparability and reproducibility of identification and typing results for Staphylococcus spp by MALDI-TOF mass spectrometry. Comparing 292 Staphylococcus spp. isolates in clinical specimens obtained fron the multidisciplinary hospital at the NWSMU im. I.I. Mechnikov was carried out by using MALDI-TOF mass spectrometer BactoSCREEN ID (Litech, Russia) and Bruker Biotyper 3.1 (Bruker GmbH, Germany). Comparability of Staphylococcus spp. identification showed that 95.9%; 12 isolates (4.1%) by “Bruker Biotyper 3.1” and 3 isolates (1.1%) by using “BactoSCREEN ID” were incorrectly identified. Repeated identification leveled the differences between the systems used. In addition, it was shown that the method of protein extraction did not affect reliability of Staphylococcus spp. species identification by using databases (÷2, p > 0.05) compared to intraspecific typing (÷2, p < 0.0001). Using different extraction protocols showed that Staphylococcus spp. mass-spectra differed by peak intensity level within the mass range up to 4000 m/z, 5300±600 m/z and 6500±500 m/z, as well as higher than 7000 m/z. Peaks of low-molecular weight peptides were detected under full protein extraction compared to sample preparation on plate extraction. To develop a unified protocol for mass-spectrometry profile processing, a reliability of the basic statistical variables (mode, median, maximum, minimum and arithmetic mean) was evaluated. Analysis of the median mass spectrometry profiles is recommended for Staphylococcus spp. intraspecific typing by using MALDI-TOF mass spectrometry as the most reproducible and consistent approach. As a result, two systems for MALDI-TOF mass spectrometry reliably identify Staphylococcus spp., but standardization of sample preparation and bioinformation analysis is required for Staphylococcus spp. typing.

scholarly journals Assessment and refinement of sample preparation methods for deep and quantitative plant proteome profiling

2018 ◽  
Author(s):  
Gaoyuan Song ◽  
Polly Y Yingshan Hsu ◽  
Justin W. Walley
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Protein Extraction ◽  
Leaf Tissue ◽  
Extraction Methods ◽  
Label Free ◽  
Preparation Methods ◽  
Proteome Profiling ◽  
Leaf Tissues ◽  
Sample Preparation Methods

SummaryA major challenge in the field of proteomics is obtaining high quality peptides for comprehensive proteome profiling by liquid chromatography mass spectrometry for many organisms. Here we evaluate and modify a range of sample preparation methods using photosynthetically active Arabidopsis leaf tissues from several developmental timepoints. We find that inclusion of FASP-based on filter digestion improves all protein extraction methods tested. Ultimately, we show that a detergent-free urea-FASP approach enables deep and robust quantification of leaf proteomes. For example, from 4-day-old leaf tissue we profiled up to 11,690 proteins from a single sample replicate. This method should be broadly applicable to researchers working on difficult to process samples from a range of plant and non-plant organisms.AbbreviationsChloroMethanol/Chloroform ExtractionFASPFilter Aided Sample PrepGOGene OntologyIAAIodoacetamideLFQLabel Free QuantificationMS/MSTandem mass spectrometryTFTranscription FactorUAUrea Extraction1D1 Dimensional2D2 Dimensional

scholarly journals Grass Material as a Modern Process Standard for 14C Analysis of n-Alkanes

Radiocarbon ◽  
2016 ◽  
Vol 58 (3) ◽  
pp. 445-458 ◽  
Author(s):  
L M Cisneros-Dozal ◽  
X Xu ◽  
C Bryant ◽  
E J Pearson ◽  
J A J Dungait
Keyword(s):  
Mass Spectrometry ◽  
Gas Chromatography ◽  
Sample Preparation ◽  
Extraction Methods ◽  
Chemical Extraction ◽  
Sample Processing ◽  
Elution Time ◽  
Modern Process ◽  
Chemical Extraction Methods ◽  
Suitable Process

AbstractOne of the difficulties in reporting accurate radiocarbon results from compound-specific radiocarbon analysis (CSRA) is the lack of suitable process standard materials to correct for the amount and 14C content of carbon added during extensive sample processing. We evaluated the use of n-alkanes extracted from modern grass material (1.224±0.006 fraction modern) as process standards for CSRA. The n-alkanes were isolated using preparative capillary gas chromatography (PCGC) from two independent chemical extraction methods applied to the grass. Since this was our first assessment of the 14C content of the grass n-alkanes, we corrected for extraneous carbon derived from PCGC isolation using commercially available single compounds of modern and 14C-free content. Results were consistent across the two extraction methods showing that the C29n-alkane has a fraction modern value that is within 1σ of the bulk value of the grass while C31n-alkane and less abundant n-alkanes have values within 2σ of the bulk value of the grass. C29 and C31n-alkanes were the most abundant n-alkanes in the grass and, as such, the more feasible for collection of sufficient amounts of carbon for accelerator mass spectrometry (AMS) analysis. Our results suggest that choosing a grass n-alkane with an elution time closest to that of the unknowns may be advisable due to possibly greater effect from GC column bleed (14C-free) at later elution times. We conclude that C29 and C31n-alkanes in modern grass of known 14C content can be used as in-house standards to correct for the addition of 14C-free carbon during sample preparation for 14C analysis of n-alkanes.

scholarly journals Radiocarbon Measurement Program at the Centro Nacional de Aceleradores (CNA), Spain

Radiocarbon ◽  
2009 ◽  
Vol 51 (2) ◽  
pp. 883-889 ◽  
Author(s):  
F Javier Santos Arévalo ◽  
Isabel Gómez Martínez ◽  
Manuel García León
Keyword(s):  
Mass Spectrometry ◽  
Sample Preparation ◽  
Accelerator Mass Spectrometry ◽  
Scientific Community ◽  
Sample Processing ◽  
Main Research ◽  
Research Programs ◽  
Set Up ◽  
Reference Samples

In September 2005, an accelerator mass spectrometry (AMS) system based on a 1MV Tandetron accelerator arrived at the Centro Nacional de Aceleradores (CNA). One of the main research programs for this AMS facility is based on radiocarbon. At the same time as the AMS facility was installed and tested, the 14C sample preparation laboratory was designed and set up. A graphitization line that allows the preparation of 5 samples in parallel was designed and built in October 2006. The first months were mainly dedicated to check and optimize all the sample processing. For such a task, several reference samples have been prepared and measured. Since the beginning of 2007, the laboratory has been fully operational and is currently performing as a service for the scientific community. During 2007, nearly 100 unknown samples were prepared and measured in our AMS system. Most of them were for dating purposes, but also other applications were investigated. The performance of the 14C laboratory and dating service will be shown, with some examples as illustration.

scholarly journals Comparison and performance of two cosmogenic nuclide sample preparation procedures of in situ produced 10Be and 26Al

2021 ◽  
Author(s):  
Zsófia Ruszkiczay-Rüdiger ◽  
Stephanie Neuhuber ◽  
Régis Braucher ◽  
Johannes Lachner ◽  
Peter Steier ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Sample Processing ◽  
Processing Methods ◽  
Cosmogenic Nuclide ◽  
Environmental Materials ◽  
And Performance ◽  
Cosmogenic Radionuclide ◽  
Data Reproducibility

AbstractCosmogenic radionuclide 10Be and 26Al targets (BeO and Al2O3) for AMS analysis are produced by a growing number of geochemical laboratories, employing different sample processing methods for the extraction of Be and Al from environmental materials. The reliability of this geochronological tool depends on data reproducibility independent from the preparation steps and the AMS measurements. Our results demonstrate that 10Be and 26Al concentrations of targets processed following different, commonly used protocols and measured at two AMS facilities lead to consistent results. However, insoluble fluoride precipitates, if formed during processing, can cause decreased 26Al results, while 10Be concentrations are unaffected.

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