A Novel UHPLC-MS/MS Method for Measuring 8-iso-Prostaglandin F2α in Bronchoalveolar Lavage Fluid.
Ontology highlight
ABSTRACT:
SUBMITTER: Holder C
PROVIDER: S-EPMC8406638 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:We provide a review of proteomic techniques used to characterize the bronchoalveolar lavage fluid (BALF) proteome of normal healthy subjects. Bronchoalveolar lavage (BAL) is the most common technique for sampling the components of the alveolar space. The proteomic techniques used to study normal BALF include protein separation by 2DE, whereby proteins were identified by comparison to a reference gel as well as high pressure liquid chromatography (HPLC)-MS/MS, also known as shotgun proteomics. We summarize recent progress using shotgun MS technologies to define the normal BALF proteome. Surprisingly, we find that despite advances in shotgun proteomic technologies over the course of the last 10 years, which have resulted in greater numbers of proteins being identified, the functional landscape of normal BALF proteome was similarly described by all methods examined.
| S-EPMC4239657 | biostudies-literature
Project description:bronchoalveolar lavage fluid Metagenome
| PRJNA449183 | ENA
Project description:The 8-iso-prostaglandin F2α (8-iso-PGF2α) biomarker is used as the gold standard for tracing lipid oxidative stress in vivo. The analysis of urinary 8-iso-PGF2α is challenging when dealing with trace amounts of 8-iso-PGF2α and the complexity of urine matrixes. A packed-fiber solid-phase extraction (PFSPE)-coupled with HPLC-MS/MS-method, based on polystyrene (PS)-electrospun nanofibers, was developed for the specific determination of 8-iso-PGF2α in urine and compared with other newly developed LC-MS/MS methods. The method, which simultaneously processed 12 samples within 5 min on a self-made semi-automatic array solid-phase extraction processor, was the first to introduce PS-electrospun nanofibers as an adsorbent for the extraction of 8-iso-PGF2α and was successfully applied to real urine samples. After optimizing the PFSPE conditions, good linearity in the range of 0.05-5 ng/mL with R2 > 0.9996 and a satisfactory limit of detection of 0.015 ng/mL were obtained, with good intraday and interday precision (RSD < 10%) and recoveries of 95.3-103.8%. This feasible method is expected to be used for the batch quantitative analysis of urinary 8-iso-PGF2α.
| S-EPMC9318247 | biostudies-literature
Project description:Acute respiratory distress syndrome (ARDS) remains a significant hazard to human health and is clinically challenging because there are no prognostic biomarkers and no effective pharmacotherapy. The lung compartment metabolome may detail the status of the local environment that could be useful in ARDS biomarker discovery and the identification of drug target opportunities. However, neither the utility of bronchoalveolar lavage fluid (BALF) as a biofluid for metabolomics nor the optimal analytical platform for metabolite identification is established. To address this, we undertook a study to compare metabolites in BALF samples from patients with ARDS and healthy controls using a newly developed liquid chromatography (LC)-mass spectroscopy (MS) platform for untargeted metabolomics. Following initial testing of three different high-performance liquid chromatography (HPLC) columns, we determined that reversed phase (RP)-LC and hydrophilic interaction chromatography (HILIC) were the most informative chromatographic methods because they yielded the most and highest quality data. Following confirmation of metabolite identification, statistical analysis resulted in 37 differentiating metabolites in the BALF of ARDS compared with health across both analytical platforms. Pathway analysis revealed networks associated with amino acid metabolism, glycolysis and gluconeogenesis, fatty acid biosynthesis, phospholipids, and purine metabolism in the ARDS BALF. The complementary analytical platforms of RPLC and HILIC-LC generated informative, insightful metabolomics data of the ARDS lung environment.
| S-EPMC4068805 | biostudies-literature
Project description:Respiratory tract infections are a major health problem and indication for antimicrobial use in cattle and in humans. Currently, most antimicrobial treatments are initiated without microbiological results, holding the risk of inappropriate first intention treatment. The main reason for this empirical treatment is the long turnaround time between sampling and availability of identification and susceptibility results. Therefore the objective of the present study was to develop a rapid identification procedure for pathogenic respiratory bacteria in bronchoalveolar lavage fluid (BALf) samples from cattle by MALDI-TOF MS, omitting the cultivation step on agar plates to reduce the turnaround time between sampling and identification of pathogens. The effects of two different liquid growth media and various concentrations of bacitracin were determined to allow optimal growth of Pasteurellaceae and minimise contamination. The best procedure was validated on 100 clinical BALf samples from cattle with conventional bacterial culture as reference test. A correct identification was obtained in 73% of the samples, with 59.1% sensitivity (Se) (47.2-71.0%) and 100% specificity (Sp) (100-100%) after only 6?hours of incubation. For pure and dominant culture samples, the procedure was able to correctly identify 79.2% of the pathogens, with a sensitivity (Se) of 60.5% (45.0-76.1%) and specificity (Sp) of 100% (100-100%). In mixed culture samples, containing ?2 clinically relevant pathogens, one pathogen could be correctly identified in 57% of the samples with 57.1% Se (38.8-75.5%) and 100% Sp (100-100%). In conclusion, MALDI-TOF MS is a promising tool for rapid pathogen identification in BALf. This new technique drastically reduces turnaround time and may be a valuable decision support tool to rationalize antimicrobial use.
| S-EPMC6895124 | biostudies-literature
Project description:BackgroundClinical bronchoalveolar lavage fluid (BALF) samples are rich in biomolecules, including proteins, and useful for molecular studies of lung health and disease. However, mass spectrometry (MS)-based proteomic analysis of BALF is challenged by the dynamic range of protein abundance, and potential for interfering contaminants. A robust, MS-based proteomics compatible sample preparation workflow for BALF samples, including those of small and large volume, would be useful for many researchers.ResultsWe have developed a workflow that combines high abundance protein depletion, protein trapping, clean-up, and in-situ tryptic digestion, that is compatible with either qualitative or quantitative MS-based proteomic analysis. The workflow includes a value-added collection of endogenous peptides for peptidomic analysis of BALF samples, if desired, as well as amenability to offline semi-preparative or microscale fractionation of complex peptide mixtures prior to LC-MS/MS analysis, for increased depth of analysis. We demonstrate the effectiveness of this workflow on BALF samples collected from COPD patients, including for smaller sample volumes of 1-5 mL that are commonly available from the clinic. We also demonstrate the repeatability of the workflow as an indicator of its utility for quantitative proteomic studies.ConclusionsOverall, our described workflow consistently provided high quality proteins and tryptic peptides for MS analysis. It should enable researchers to apply MS-based proteomics to a wide-variety of studies focused on BALF clinical specimens.
| S-EPMC10068177 | biostudies-literature
Project description:BackgroundIt is now possible to comprehensively characterize the microbiota of the lungs using culture-independent, sequencing-based assays. Several sample types have been used to investigate the lung microbiota, each presenting specific challenges for preparation and analysis of microbial communities. Bronchoalveolar lavage fluid (BALF) enables the identification of microbiota specific to the lower lung but commonly has low bacterial density, increasing the risk of false-positive signal from contaminating DNA. The objectives of this study were to investigate the extent of contamination across a range of sample densities representative of BALF and identify features of contaminants that facilitate their removal from sequence data and aid in the interpretation of BALF sample 16S sequencing data.ResultsUsing three mock communities across a range of densities ranging from 8E+?02 to 8E+?09 16S copies/ml, we assessed taxonomic accuracy and precision by 16S rRNA gene sequencing and the proportion of reads arising from contaminants. Sequencing accuracy, precision, and the relative abundance of mock community members decreased with sample input density, with a significant drop-off below 8E+?05 16S copies/ml. Contaminant OTUs were commonly inversely correlated with sample input density or not reproduced between technical replicates. Removal of taxa with these features or physical concentration of samples prior to sequencing improved both sequencing accuracy and precision for samples between 8E+?04 and 8E+?06 16S copies/ml. For the lowest densities, below 8E+?03 16S copies/ml BALF, accuracy and precision could not be significantly improved using these approaches. Using clinical BALF samples across a large density range, we observed that OTUs with features of contaminants identified in mock communities were also evident in low-density BALF samples.ConclusionRelative abundance data and community composition generated by 16S sequencing of BALF samples across the range of density commonly observed in this sample type should be interpreted in the context of input sample density and may be improved by simple pre- and post-sequencing steps for densities above 8E+?04 16S copies/ml.
| S-EPMC6821041 | biostudies-literature
Project description:Wastewater analysis has been demonstrated to be a complementary approach for assessing the overall patterns of drug use by a population while the full potential of wastewater-based epidemiology has yet to be explored. F2-isoprostanes are a prototype wastewater biomarker to study the cumulative oxidative stress at a community level. In this work, 8-iso-prostaglandin F2α (8-iso-PGF2α) was analysed in raw 24 h-composite wastewater samples collected from 4 Norwegian and 7 other European cities in 2014 and 2015. Using the same samples, biomarkers of alcohol (ethyl sulfate) and tobacco (trans-3'-hydroxycotinine) use were also analysed to investigate any possible correlation between 8-iso-PGF2α and the consumption of the two drugs. The estimated per capita daily loads of 8-iso-PGF2α in the 11 cities ranged between 2.5 and 9.9 mg/day/1000 inhabitants with a population-weighted mean of 4.8 mg/day/1000 inhabitants. There were no temporal trends observed in the levels of 8-iso-PGF2α, however, spatial differences were found at the inter-city level correlating to the degree of urbanisation. The 8-iso-PGF2α mass load was found to be strongly associated with that of trans-3'-hydroxycotinine while it showed no correlation with ethyl sulfate. The present study shows the potential for 8-iso-PGF2α as a wastewater biomarker for the assessment of community public health.
| S-EPMC5157025 | biostudies-literature
Project description:The long-term goal of our study is to identify chronic obstructive pulmonary disease (COPD)-related bronchoalveolar lavage fluid (BALF) nitroproteins to clarify COPD pathological mechanisms and to discover biomarkers of COPD. The goal of the present study was to detect the presence of, and potential roles of, nitroproteins in, human ex-smoker (without COPD) BALF samples. Nitroproteins were immunoprecipitated from two separate BALF samples, and digested with trypsin; and tryptic peptides were analyzed with matrix-assisted laser desorption/ionization (MALDI)-tandem mass spectrometry (MS/MS). Each MS/MS spectrum was composed of accumulated scans (n = 50-100). The MS/MS data were searched with BioWorks 2.0 TuboSequest in the SwissProt database to generate the amino acid sequence, which was evaluated manually. Eleven nitrotyrosine sites were identified in eight proteins, including progestin and adipoQ receptor family member III, zinc finger protein 432, proteasome subunit alpha type 2, NADH-ubiquinone oxidoreductase B14, slit homolog 1 protein, lysozyme, aldose 1-epimerase, and PTS system lactose-specific EIICB component. Each nitrotyrosine site was located within a specific protein domain and motif. Those identified nitrated proteins could be involved in multiple functional metabolic systems, including transcriptional regulation, mitochondrial complex, immune system, and energy metabolism.
| S-EPMC3295046 | biostudies-literature
Project description:Global-scale examination of protein phosphorylation in human biological fluids by phosphoproteomics approaches is an emerging area of research with potential for significant contributions towards discovery of novel biomarkers. In this pilot work, we analyzed the phosphoproteome in human bronchoalveolar lavage fluid (BAL) from nondiseased subjects. The main objectives were to assess the feasibility to probe phosphorylated proteins in human BAL and to obtain the initial catalog of BAL phosphoproteins, including protein identities and exact description of their phosphorylation sites. We used a gel-free bioanalytical workflow that included whole-proteome digestion of depleted BAL proteins, enrichment of phosphopeptides by immobilized metal ion affinity chromatography (IMAC), LC-MS/MS analyses with a linear ion trap mass spectrometer, and searches of a protein sequence database to generate a panel of BAL phosphoproteins and their sites of phosphorylation. Based on sequence-diagnostic MS/MS fragmentation patterns, we identified a collection of 36 phosphopeptides that contained 26 different phosphorylation sites. These phosphopeptides mapped to 21 phosphoproteins including, for example, vimentin, plastin-2, ferritin heavy chain, kininogen-1, and others. The characterized phosphoproteins have diverse characteristics in terms of cellular origin and biological function. To the best of our knowledge, results of this study represent the first description of the human BAL phosphoproteome.
| S-EPMC3446654 | biostudies-literature