Single Nucleus DNA Sequencing of Flow Sorted Archived Frozen and Formalin Fixed Paraffin Embedded Solid Tumors
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PROVIDER: PRJNA1119467 | ENA |
REPOSITORIES: ENA
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Project description:Archived tissues are a vast resource of annotated clinical samples. To expand their use for translational studies, we optimized flow sorting and sequencing of single nuclei from archived frozen and FFPE tumor tissues. Our methods, which include isolation and preparation of intact nuclei suitable for library preparations, quality control (QC) metrics for each step, and a single cell sequencing bioinformatic processing and analysis pipeline, were validated with flow sorted nuclei from matching frozen and FFPE ovarian cancer surgical samples and a sequencing panel of 553 amplicons targeting single nucleotide and copy number variants in genes of interest.
2024-06-20 | GSE270364 | GEO
Project description:We applied DNA content based flow cytometry methods to interrogate the genomes of clinical samples from 8 patients with ovarian cancer. These included 6 high grade serous ovarian carcinomas, a low grade serous carcinoma, and an endometriod carcinoma. Archived samples obtained from surgical resections from a University of California San Francisco (UCSF) tissue bank. Notably for this study we distinguished and sorted diploid and aneuploid tumors. We then profiled the the whole genome copy number for each sample. These data were used to provide standards for single cell analyses of the same tissues including FFPE samples.
2024-09-24 | GSE268921 | GEO
Project description:Profiling histone modifications in clinical archived formalin fixed paraffin embedded samples
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Project description:MicroRNAs regulate several aspects of tumorigenesis and cancer progression. Most cancer tissues are archived formalin-fixed and paraffin-embedded (FFPE). While microRNAs are a more stable form of RNA thought to withstand FFPE-processing and degradation there is only limited evidence for the latter assumption. We examined whether microRNA profiling can be successfully conducted on FFPE cancer tissues using SOLiD ligation based sequencing. Tissue storage times (3-9 years) appeared to not affect the number of detected microRNAs in FFPE samples compared to matched frozen samples (paired t-test p>0.7). Correlations of microRNA expression values were very high across microRNAs in a given sample (Pearson’s r=0.71-0.95). Higher variance of expression values among samples was associated with higher correlation coefficients between FFPE and frozen tissues. One of the FFPE samples in this study was degraded for unknown reasons with a peak read length of 17 nucleotides compared to 21 in all other samples. The number of detected microRNAs in this sample was within the range of microRNAs detected in all other samples. Ligation-based microRNA deep sequencing on FFPE cancer tissues is feasible and RNA degradation to the degree observed in our study appears to not affect the number of microRNAs that can be quantified.
2013-04-15 | GSE45740 | GEO
Project description:Tissue biopsies are most commonly archived in a paraffin block following tissue fixation with formaldehyde (FFPE) or as fresh frozen tissue (FFT). While both methods preserve biological samples, little is known about how they affect the quantifiable proteome. We performed a ‘bottom-up’ proteomic analysis (N=20) of short and long-term archived FFPE surgical samples of human meningiomas and compared them to matched FFT specimens. FFT facilitated identification of a slightly higher number of proteins compared with matched FFPE specimens (5735 vs 5670 proteins, respectively (p < 0.05), regardless of archival time. However, marked differences in the proteome composition were apparent between FFPE and FFT specimens. Twenty-three percent of FFPE-derived peptides and 8% of FFT-derived peptides contained at least one chemical modification. Methylation and formylation were most prominent in FFPE-derives peptides (36% and 17% of modified FFPE peptides, respectively) while, most of phosphorylation and iron modifications appeared in FFT-derived peptides (p<0.001). A mean 14% (2.9) of peptides identified in FFPE contained at least one modified Lysine residue. Importantly, larger proteins were significantly overrepresented in FFT specimens, while FFPE specimens were enriched with smaller proteins. This work cautions against comparing results of proteomic studies derived from different archival methods.
2024-02-12 | PXD032108 | Pride
Project description:Here, we present FACT-Seq, a highly sensitive method to decode the histone modifications in FFPE tissues. The FACT-Seq generates the high-quality chromatin profiles from both active and silent histone modifications with 1000 nuclei purified from a single FFPE tissue section, and reveals the disease-specific super enhancers in the FFPE archived human colorectal and human glioblastoma multiforme cancer tissue. In summary, the approach allows decoding the histone modifications that regulate gene expression in archival FFPE tissues with high sensitivity, in order to better understand epigenetic regulation in cancer and precision medicine.
2021-09-03 | GSE171758 | GEO
Project description:MicroRNAs regulate several aspects of tumorigenesis and cancer progression. Most cancer tissues are archived formalin-fixed and paraffin-embedded (FFPE). While microRNAs are a more stable form of RNA thought to withstand FFPE-processing and degradation there is only limited evidence for the latter assumption. We examined whether microRNA profiling can be successfully conducted on FFPE cancer tissues using SOLiD ligation based sequencing. Tissue storage times (3-9 years) appeared to not affect the number of detected microRNAs in FFPE samples compared to matched frozen samples (paired t-test p>0.7). Correlations of microRNA expression values were very high across microRNAs in a given sample (PearsonM-bM-^@M-^Ys r=0.71-0.95). Higher variance of expression values among samples was associated with higher correlation coefficients between FFPE and frozen tissues. One of the FFPE samples in this study was degraded for unknown reasons with a peak read length of 17 nucleotides compared to 21 in all other samples. The number of detected microRNAs in this sample was within the range of microRNAs detected in all other samples. Ligation-based microRNA deep sequencing on FFPE cancer tissues is feasible and RNA degradation to the degree observed in our study appears to not affect the number of microRNAs that can be quantified. we analyzed eight matched frozen and FFPE tissues of different histologies for which detailed processing and storage information.
2013-04-15 | E-GEOD-45740 | biostudies-arrayexpress
Project description:In recent years, the use of FFPE tissue in gene expression microarray (GEM) studies has become a topic of increased interest, because pathology departments worldwide contain an invaluable source of biological disease-specific FFPE- tissue material. Thus far, some published studies on FFPE tissue consider the processed tissue amenable, whereas other studies assert that the tissue is not useful in GEM. In general, pancreatic tissue is known to contain large amounts of nucleases, leading to high turnover rates and degradation of RNA in the tissue. This characteristic, in combination with FFPE processing time and its effect on the tissue, does not make FFPE pancreatic tissue the obvious choice for GEM. The basis for using FFPE pancreatic tissue in RNA-based assays seems suboptimal. Previous GEM studies have used different FFPE tissue types, but GEM studies on FFPE pancreatic tissue have not been published. In most studies, the tissue was specially processed for use with GEMs. Here we demonstrate the usefulness of randomly archived FFPE pancreatic tissues for GEMs. For this purpose we included FFPE pancreatic tissue from patients with congenital hyperinsulinism or insulinoma; In these patients, pancreas resection is performed when medical treatment is not adequate to prevent hyperinsulinemic hypoglycemia or when patients do not respond to medical treatment. Although ribonuclease-rich, we obtained biologically relevant and disease-specific, significant genes; cancer-related genes and genes involved in a) the regulation of insulin secretion and synthesis, b) amino acid metabolism, and c) calcium ion homeostasis. This application may extend the possibilities of gene expression studies to many tissue types, especially in rare diseases, for which fresh frozen tissue is not readily available. A total of 15 samples were included in this study using a 58K chip. 29,134 human 60mer oligonucleotide targets were arrayed on the chips in duplicates. Each chip is hybridized with sample material (labelled with Cy5) from one patient and reference material (labelled with Cy3). Reference material contains pooled RNA from four different cancer cell lines; HeLa (cervical epithelium), SK-BR-3 (mammary gland), HT29 (colon) and A431 (skin) cells. Seven patients with congenital hyperinsulinism (five FFPE and two frozen samples), one patient with insulinoma and five controls were included in the study. Inclusion criteria for controls was that their cause of death was not pancreas-related. Samples were obtained from archived FFPE pancreatic material. The insulinoma sample was included in the study, because the biopsy was originally divided with one part being frozen and the other part being routinely formalin-fixed and paraffin-embedded. In the study we therefore include a frozen insulinoma sample and a FFPE insulinoma sample that was processed in duplicate.
2012-09-30 | E-GEOD-32610 | biostudies-arrayexpress
Project description:DNA methylation is the most studied epigenetic modification due to its role in regulating gene expression and aberrations in methylation involved in the pathogenesis of cancer and several diseases. The method of choice to evaluate genome-wide methylation has been the Illumina HumanMethylation450 BeadChip (450K), but it was recently replaced with the MethylationEPIC BeadChip (EPIC). We therefore sought to validate the EPIC array in comparison to the 450K array for both fresh-frozen (FF) and formalin-fixed paraffin-embedded (FFPE) pediatric brain tumors. We also performed analysis on the EPIC array with paired FF and FFPE samples, to adapt to a clinical setting where FFPE is routinely used. Further, we compared two restoration methods, REPLI-g and Infinium, for FFPE-derived DNA on the EPIC array.
2017-04-01 | GSE92579 | GEO
Project description:DNA methylation is the most studied epigenetic modification due to its role in regulating gene expression and aberrations in methylation involved in the pathogenesis of cancer and several diseases. The method of choice to evaluate genome-wide methylation has been the Illumina HumanMethylation450 BeadChip (450K), but it was recently replaced with the MethylationEPIC BeadChip (EPIC). We therefore sought to validate the EPIC array in comparison to the 450K array for both fresh-frozen (FF) and formalin-fixed paraffin-embedded (FFPE) pediatric brain tumors. We also performed analysis on the EPIC array with paired FF and FFPE samples, to adapt to a clinical setting where FFPE is routinely used. Further, we compared two restoration methods, REPLI-g and Infinium, for FFPE-derived DNA on the EPIC array.
2017-04-01 | GSE92577 | GEO