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Project description:Biospecimen Pre-analytical Variables (BPV)
| PRJNA376469 | ENA
Project description:The diagnostic and therapeutic use of extracellular vesicles (EV) is under intense investigation and may lead to societal benefits. Reference materials are an invaluable resource for developing, improving and assessing the performance of regulated EV applications and for quantitative and objective data interpretation. We have engineered recombinant extracellular vesicles (rEV) as a biological reference material. rEV have similar biochemical and biophysical characteristics as sample EV and function as an internal quantitative and qualitative control throughout analysis. Spike-in applications of rEV in bodily fluids prior to EV analysis map technical variability of EV applications and promote intra and inter laboratory studies. This protocol describes the production, recovery and quality assurance of rEV, their dilution and addition to bodily fluids, and the detection steps based on fluorescence-, nucleic acid- and protein measurements. Multiple application potentials for rEV are exemplified, including method development, big data normalization and assessment of pre-analytical variables. The protocol can be adopted by researchers with standard laboratory and basic EV separation/characterization experience and requires ~4–5 d.
2023-03-13 | PXD017542 | Pride
Project description:<p>The Biospecimen Pre-analytical Variables (BPV) Program is a National Cancer Institute-sponsored study to systematically assess the effects of pre-analytical factors on the molecular profile of biospecimens. A robust biospecimen collection infrastructure was established to prospectively collect biospecimens using rigorous standard operating procedures to control for most variables while introducing experimental conditions to study specific biospecimen handling issues, including the cold ischemic time (delay to formalin fixation), time in formalin, freezing methods, and storage temperatures and durations. RNA and DNA from biospecimens collected under these conditions was analyzed on multiple molecular platforms. The potential effects of these pre-analytical conditions on protein integrity and detection of metabolites were also examined. Data from this study will be used to develop evidence-based biospecimen standard operating procedures and best practices for fit-for-purpose collection, processing, and storage of biospecimens.</p> <p>The BPV Cohort is utilized in the following dbGaP substudies. To view genotypes, analysis, expression data, other molecular data, and derived variables collected in these substudies, please click on the following sub-studies below or in the "Substudies" box located on the right hand side of this top-level study page phs001304 BPV Cohort. The substudy links will be active once they are released by dbGaP.</p> <p> <ol> <li>Preanalytical Impacts on Global Metabolite Profiling - plasma (MassSpec by Metabolon) This study was to evaluate the impact of the storage temperature (s) (-80°C and LN2 vapor) and the length of storage on human plasma quality using LC-MS/MS (liquid-chromatography-mass spectrometry/mass spectrometry) based global metabolite profiling. The study includes 240 plasma samples collected from 40 donors.</li> <li>Investigate the effect of the delay to fixation on the proteome and phosphoproteome -FFPE (MassSpec by Caprion). The study is to do proteome and phosphoproteome analysis on Delay to fixation was carried out using FFPE tumor samples from colon and ovarian cancer patients comparing 2, 3, and 12hr delay to fixation to the 1hr time point. The study includes 100 samples 20 donors.</li> <li>Investigate the effect of storage conditions of tumor specimens on the proteome and phosphoproteome profiling- Frozen tissue and plasma (MassSpec by Caprion). The study was to evaluate the effects of storage conditions on tumor specimens. Plasma samples from 40 cancer patients stored at two different temperatures (-80°C and LN2) for a given period (0-2, 6-8, and 12-14 months) were evaluated. Frozen kidney tumor samples from 20 patients were compared for effects of different snap frozen (dry ice vs. LN2) and storage temperatures (-80°C and LN2). The study includes 100 tissue and 240 plasma samples from 60 donors.</li> <li>Preanalytical Impacts on Genomic Sequencing by Next Generation Sequencing (NGS) technology (mRNA/miRNA and WES by Expression Analysis). The goal of the study is to determine the effects of cold ischemic delay-to-fixation (4 time points) and formalin preservation (FFPE) on the nature and quality of genomic profiles using the matched freshly frozen sample as the gold standard, which including WES, RNAseq. The study includes 395 samples from 37 donors.</li> <li>Preanalytical Impacts on Copy Number Variation (CNV) Detection by aCGH technology (aCGH by Georgetown University). This study was to use aCGH to evaluate the effect of variation in cold ischemia time and time in formalin fixation on CNV in DNA extracted from kidney cancer specimens. The study includes 235 samples from 40 donors.</li> <li>Evaluation of frozen conditions on mRNA profiling by TaqMan assay (mRNA expression by Georgetown University). This study was to utilize gene expression profiling, using custom TaqMan arrays, to compare the molecular profiles of RNA from frozen tumor samples collected using two freezing methods (dry ice or LN2), two storage temperatures (-80°C or LN2 vapor), as well as Optimal Cutting Temperature (OCT) compound and non-OCT embedded. The study includes 100 samples from 20 donors.</li> <li>mRNA signature for stratification by cold ischemia time (mRNA expression by IBBL). The study was to determine the effects of cold ischemic time (delay-to-fixation) and formalin preservation (FFPE) on mRNA detection by Taqman assay using tumor tissue specimens from kidney, colon and ovarian cancer patients. There are160 samples from 40 donors.</li> </ol> </p> <p><b>The Biospecimen PV cohort is utilized in the following dbGaP individual studies.</b> To view molecular data, and derived variables collected in these individual studies, please click on the following individual studies below or in the "Sub-studies" box located on the right hand side of this top-level study page <a href="study.cgi?study_id=phs001304">phs001304</a> Biospecimen PV cohort. <ul> <li><a href="study.cgi?study_id=phs001634">phs001634</a> CIT mRNA</li> <li><a href="study.cgi?study_id=phs001635">phs001635</a> CNV aCGH</li> <li><a href="study.cgi?study_id=phs001636">phs001636</a> Fixation Delay</li> <li><a href="study.cgi?study_id=phs001637">phs001637</a> Global Metabolite Profiling</li> <li><a href="study.cgi?study_id=phs001638">phs001638</a> mRNA TaqMan</li> <li><a href="study.cgi?study_id=phs001639">phs001639</a> NGS</li> <li><a href="study.cgi?study_id=phs001640">phs001640</a> Tumor Storage</li> </ul> </p>
| phs001304 | dbGaP
Project description:Gene expression analyses of disseminated tumor cells can provide indispensable biological and clinical information at diagnosis, during therapy and at relapse. So far, however, disseminated tumor cells expression analyses have been hampered by insufficient infiltration rates and/or unclear consequences of transport duration, temperature and other pre-handling procedures. This study, using neuroblastoma as a model, focuses on the influences of pre-analytical procedures like tumor cell enrichment, transport time and temperature on the disseminated tumor cells expression profile.
2015-12-01 | GSE63294 | GEO
Project description:Our previous study identified 8 risk and 9 protective plasma miRNAs associated with progression to end-stage kidney disease (ESKD) in diabetes. This study aimed to elucidate pre-analytical factors that influence the quantification of circulating miRNAs. Using the EdgeSeq platform, which quantifies 2,002 miRNAs in plasma, including ESKD-associated miRNAs, we compared miRNA profiles in whole plasma vs. miRNAs profiles in RNA extracted from the same plasma specimens. Less than half of the miRNAs were detected in standard RNA extracted methods from plasma. Detection of individual and concentrations of miRNAs were much lower when RNA extracted from plasma was quantified by RNA sequencing (RNASeq) or RT-qPCR platforms compared to EdgeSeq. Plasma profiles of miRNAs determined by the EdgeSeq platform had excellent reproducibility in assessment, and had no variation with age, sex, HbA1c, BMI, and cryostorage time. The risk ESKD-associated miRNAs were detected and measured accurately only in whole plasma and using the EdgeSeq platform. Protective ESKD-associated miRNAs were detected by all platforms except RT-qPCR, however, correlations among concentrations obtained with different platforms were weak or non-existent. In conclusion, pre-analytical factors have a profound effect on detection and quantification of circulating miRNAs in ESKD in diabetes. Quantification of miRNAs in whole plasma and using the EdgeSeq platform may be the preferable method to study profiles of circulating cell-free miRNAs associated with ESKD, and possibly other diseases.
2024-05-07 | GSE266819 | GEO
Project description:Impact of pre-analytical handling on Bone Marrow (BM) mRNA Gene Expression: We have investigated the impact of RNA extraction protocols and time delays (24 and 48h) between sample aspiration and RNA extraction on RNA quality and gene expression profiles. Intact RNA can be extracted from BM samples stored at room temperature for up to 48h after aspiration. However, storage of BM has dramatic effects on mRNA expression of individual transcripts. Keywords: time-course
2004-04-28 | GSE1347 | GEO
Project description:Background: assays that account for the biological properties and fragmentation of cell-free DNA (cfDNA) can improve the performance of liquid biopsy. However, pre-analytic and physiological differences between individuals on fragmentomic analysis are poorly defined.
Methods: we analyzed the impact of collection tube, plasma processing time and physiology on the size distribution of cfDNA, their genome-wide representation and sequence diversity at the cfDNA fragment-ends using shallow Whole Genome Sequencing.
Results: we observed that using different stabilizing collection tubes, or processing times does not affect the cfDNA fragment sizes, but can impact the genome-wide fragmentation patterns and fragment-end sequences of cfDNA. In addition, beyond differences depending on the gender, the physiological conditions tested between 63 individuals (age, body mass index, use of medication and chronic conditions) minimally influenced the outcome of fragmentomic methods.
Conclusions: our results highlight that fragmentomic approaches have potential for implementation in the clinic, pending clear traceability of analytical and physiological factors.
| EGAS00001005748 | EGA
Project description:Tumor specimens are often preserved as formalin-fixed paraffin-embedded (FFPE) tissue blocks, the most common clinical source for DNA sequencing. Herein, we evaluated the effect of pre-sequencing parameters to guide proper sample selection for targeted gene sequencing. Data from 113 FFPE lung tumor specimens were collected, and targeted gene sequencing was performed. Libraries were constructed using custom probes and were paired-end sequenced on a next generation sequencing platform. A PCR-based quality control (QC) assay was utilized to determine DNA quality, and a ratio was generated in comparison to control DNA. We observed that FFPE storage time, PCR/QC ratio, and DNA input in the library preparation were significantly correlated to most parameters of sequencing efficiency including depth of coverage, alignment rate, insert size, and read quality. A combined score using the three parameters was generated and proved highly accurate to predict sequencing metrics. We also showed wide read count variability within the genome, with worse coverage in regions of low GC content like in KRAS. Sample quality and GC content had independent effects on sequencing depth, and the worst results were observed in regions of low GC content in samples with poor quality. Our data confirm that FFPE samples are a reliable source for targeted gene sequencing in cancer, provided adequate sample quality controls are exercised. Tissue quality should be routinely assessed for pre-analytical factors, and sequencing depth may be limited in genomic regions of low GC content if suboptimal samples are utilized.
| S-EPMC4659597 | biostudies-literature
Project description:This study tried to determine whether exposure of breast stem/progenitor cells to estrogen disrupts the epigenome of progeny epithelial cells. DNA methylation profiles were compared between control and pre-exposed epithelial cells using a genome-wide detection method called MeDIP-chip. Keywords: MeDIP-chip
2007-12-01 | GSE7844 | GEO
Project description:This SuperSeries is composed of the following subset Series: GSE14694: Computational and Analytical Framework for Small RNA Profiling by High-Throughput Sequencing (reproducibility) GSE14695: Computational and Analytical Framework for Small RNA Profiling by High-Throughput Sequencing (standards) Refer to individual Series
2009-06-02 | E-GEOD-14696 | biostudies-arrayexpress