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Protein structure files for the paper "Multiplexed identification of RAS paralog imbalance as a driver of lung cancer growth" in Nature Cell Biology by Tang et al.
<p>This archive contains models of HRAS, KRAS, and NRAS homo- and heterodimers with various mutations discussed in the paper, "Multiplexed identification of RAS paralog imbalance as a driver of lung cancer growth" in Nature Cell Biology by Tang et al.<br> as well as crystallographic dimers of these proteins as identified by the ProtCAD database, http://dunbrack2.fccc.edu/ProtCAD/Results/PfamArchClusterInfo.aspx?GroupId=8 (cluster 5). Several of the models are shown in Supp. Figure 11b and the crystallographic dimers of RAS that provide evidence for the possible biological relevance of these models are shown in Supp. Figure 11a.</p> <p>The crystallographic dimers were identified by clustering all possible interfaces generated by symmetry operators in crystals of HRAS, KRAS, and NRAS as described in the paper: Xu, Q., Dunbrack, R.L. ProtCID: a data resource for structural information on protein interactions. <em>Nat Commun</em> <strong>11</strong>, 711 (2020). https://doi.org/10.1038/s41467-020-14301-4.</p> <p>The models were created by superposing monomers of HRAS, KRAS, or NRAS onto the alpha4-alpha5 dimer present in the crystal of PDB entry 3k8y. Mutations were made in PyMOL. The structures were relaxed with the FastRelax protocol and the Ref2015 scoring function in the program Rosetta, which uses the backbone-dependent rotamer library of Shapovalov and Dunbrack to repack side chains.</p> <p>The crystallographic dimers are contained in a zipped PyMOL session. The mmCIF format for all the structures is present in a zip file, Tang_et_al_crystallographic_and_modeled_RAS_dimer_ciffiles.zip. The PyMOL session and zip file contains 87 HRAS dimers, 14 KRAS dimers, and 1 NRAS dimer, all having the interface consisting of the alpha4 and alpha5 helices. The PyMOL session also contains the modeled structures. Only Mg ions and GTP/GNP/GDP ligands are shown. Others are present but hidden and may be displayed by PyMOL ("show sticks, het").</p> <p> </p>
Cisplatin enhances cell stiffness and decreases invasiveness rate in prostate cancer cells by actin accumulation: Confocal and atomic force microscopy
<p><strong>Summary</strong></p> <p>Dataset of imaging data related to the publication Raudenska, M., Kratochvilova, M., Vicar, T., Gumulec, J., Balvan, J., Polanska, H. Pribyl, J. & Masarik, M.:Cisplatin enhances cell stiffness and decreases invasiveness rate in prostate cancer cells by actin accumulation. <em>Scientific Reports </em><strong>2019, </strong>9, 1660</p> <p>This dataset includes image data of <em>atomic force microcopy</em> (Young modulus) and <em>confocal microscopy</em>(staining of F-actin and β-tubulin) of prostate cell lines PNT1A, 22Rv1, and PC-3. </p> <p><strong>Materials and Methods</strong></p> <p><em>Cells, cell culture conditions</em></p> <p>Cells confluent up to 50–60% were washed with a FBS-free medium and treated with a fresh medium with FBS and required antineoplastic drug concentration (IC50 concentration for the particular cell line). The cells were treated with 93 µM (PC-3), 38 µM (PNT1A), and 24 µM (22Rv1) of cisplatin (Sigma-Aldrich, St. Louis, Missouri), respectively. IC50 concentrations used for treatment with docetaxel (Sigma-Aldrich, St. Louis, Missouri) were 200nM for PC-3, 70nM for PNT1A, and 150nM for 22Rv1. </p> <p><em>Long-term zinc (II) treatment of cell cultures</em></p> <p>Cells were cultivated in the constant presence of zinc(II) ions. Concentrations of zinc(II) sulphate in the medium were increased gradually by small changes of 25 or 50 µM. The cells were cultivated at each concentration no less than one week before harvesting and their viability was checked before adding more zinc. This process was used to select zinc resistant cells naturally and to ensure better accumulation of zinc within the cells (accumulation of zinc is usually poor during the short-term treatment of prostate cancer cells). Total time of the cultivation of cell lines in the zinc(II)-containing media exceeded one year. Resulting concentrations of zinc(II) in the media (IC50 for the particular cell line) were 50 µM for the PC-3 cell line, 150 µM for the PNT1A cell line, and 400 µM for the 22Rv1 cell line. The concentrations of zinc(II) in the media and FBS were taken into account. </p> <p><em>Actin and tubulin staining</em></p> <p>β-tubulin was labeled with anti- β tubulin antibody [EPR1330] (ab108342) at a working dilution of 1/300. The secondary antibody used was Alexa Fluor® 555 donkey anti-rabbit (ab150074) at a dilution of 1/1000. Actin was labeled with Alexa Fluor™ 488 Phalloidin (A12379, Invitrogen); 1 unit per slide. For mounting Duolink® In Situ Mounting Medium with DAPI (DUO82040) was used. The cells were fixed in 3.7% paraformaldehyde and permeabilized using 0.1% Triton X-100. </p> <p><em>Confocal microscopy</em></p> <p>The microscopy of samples was performed at the Institute of Biophysics, Czech Academy of Sciences, Brno, Czech Republic. Leica DM RXA microscope (equipped with DMSTC motorized stage, Piezzo z-movement, MicroMax CCD camera, CSU-10 confocal unit and 488, 562, and 714 nm laser diodes with AOTF) was used for acquiring detailed cell images (100× oil immersion Plan Fluotar lens, NA 1.3). Total 50 Z slices was captured with Z step size 0.3 μm.</p> <p><em>Atomic force microscopy</em></p> <p>We used the bioAFM microscope JPK NanoWizard 3 (JPK, Berlin, Germany) placed on the inverted optical microscope Olympus IX‑81 (Olympus, Tokyo, Japan) equipped with the fluorescence and confocal module, thus allowing a combined experiment (AFM‑optical combined images). The maximal scanning range of the AFM microscope in X‑Y‑Z range was 100‑100‑15 µm. The typical approach/retract settings were identical with a 15 μm extend/retract length, Setpoint value of 1 nN, a pixel rate of 2048 Hz and a speed of 30 µm/s. The system operated under closed-loop control. After reaching the selected contact force, the cantilever was retracted. The retraction length of 15 μm was sufficient to overcome any adhesion between the tip and the sample and to make sure that the cantilever had been completely retracted from the sample surface. Force‑distance (FD) curve was recorded at each point of the cantilever approach/retract movement. AFM measurements were obtained at 37°C (Petri dish heater, JPK) with force measurements recorded at a pulling speed of 30 µm/s (extension time 0.5 sec).</p> <p>The Young's modulus (E) was calculated by fitting the Hertzian‑Sneddon model on the FD curves measured as force maps (64x64 points) of the region containing either a single cell or multiple cells. JPK data evaluation software was used for the batch processing of measured data. The adjustment of the cantilever position above the sample was carried out under the microscope by controlling the position of the AFM‑head by motorized stage equipped with Petri dish heater (JPK) allowing precise positioning of the sample together with a constant elevated temperature of the sample for the whole period of the experiment. Soft uncoated AFM probes HYDRA-2R-100N (Applied NanoStructures, Mountain View, CA, USA), i.e. silicon nitride cantilevers with silicon tips are used for stiffness studies because they are maximally gentle to living cells (not causing mechanical stimulation). Moreover, as compared with coated cantilevers, these probes are very stable under elevated temperatures in liquids – thus allowing long-time measurements without nonspecific changes in the measured signal.</p> <p><em>Image analysis</em></p> <p>Fluorescence microscopy data were analyzed in ImageJ 1.52h and Python 3.7.1 as follows: cells were manually segmented using actin fluorescence channel, two regions were created for analysis: whole cell and cell periphery, lining a 4 μm thick region around cell border and including most of periphery actin cytoskeleton. In these two regions following parameters were measured for both actin and tubulin fluorescence: Integrated intensity, median intensity, and following regions were measured to describe cell morphology: Cell area, Maximum caliper (max feret diameter), roundness, and aspect ratio. Moreover, stress fibers were manually segmented in every cell and following parameters were measured: number of fibers per cell, feret angle of fiber, integrated intensity, fiber length, mean intensity. Next, a standard deviation of feret angles of individual fibers was calculated relatively to mean of feret angle using a circstd function from scipy package for Python.</p> <p><strong>Identification of files</strong></p> <p><em>Microscopy data</em></p> <p>Files are separated into individual zip files. The dataset of <em>confocal microscopy </em>is separated based on treatments: untreated control, docetaxel-treated cells, cisplatin-treated cells, zinc-treated cells. Filenames actin_tubulin_Zstack_cisplatin.zip, actin_tubulin_Zstack_untreated_control.zip, actin_tubulin_Zstack_zinc.zip, actin_tubulin_Zstack_docetaxel.zip. Files included in these ZIP archives are named as follows: "cellline_treatment_FOV". Files are 3-layer 16bit tiff files with layer sequence as follows: F-Actin (Phalloidin)/b-tubulin/Hoechst 33342. The dataset contains 242 FOVs of three cell line types/three treatments + one control, files are Z-stacks made of 50 slices.</p> <p>The dataset of <em>atomic force microscopy </em>(AFM) is included in one ZIP archive "AFM_YoungModulus_SetpointHeight.zip", which includes data on Young modulus and Setpoint Height of cell lines 22Rv1, PNT1A and PC-3 and treatments zinc, docetaxel, cisplatin (+control), i.e. identical like for confocal microscopy. The file naming is as follows: "AFM_cellline_treatment_FOV_Youngmodulus.tif" for Young modulus and "AFM_cellline_treatment_FOV_setpointheight.tif" for setpoint height. The data are filtered 32-bit tiff images, where the pixel value correspond to cell stiffness (young modulus) in Pa or setpoint height in m.</p> <p><em>Confocal microscopy analysis files</em></p> <p>Following files are csv tables including image analysis of actin/tubulin staining captured by confocal microscope:</p> <p>Cytoskeleton_fluo_analysis_Cell_Cell_periphery_morphology.csv: table includes analyzed data for actin and tubulin staining in following cellular regions: cell, cell periphery. Standard ImageJ parameters regarding intensity and morphology included.</p> <p>Cytoskeleton_fluo_analysis_Fibers.csv: table includes results of manual segmentation and consequent analysis of actin stres fibers in the cells. Apart from standard ImageJ parameters, also number of stress fibers per cell and standard deviation of fiber angle relative to the cell mean angle (for details see methods) are included.</p>
RNA datasets to derive predictors for immune checkpoint inhibitor therapy of non-small cell lung cancer
<p>Nanostring nCounter datasets and corresponding clinical data of tumor samples of patients with advanced NSCLC who received anti-PD-1 immuntherapy. Prospectively divided into a discovery and a validation cohort.</p> <p>Please cite the corresponding publication in Annals of Oncology (10.1093/annonc/mdz049)</p>
Ergolide Mediates Anti-Cancer Effects on Metastatic Uveal Melanoma Cells and Modulates their Cellular and Extracellular Vesicle proteomes
<p>Underlying dataset and extended dataset of the results described in the article "Ergolide Mediates Anti-Cancer Effects on Metastatic Uveal Melanoma Cells and Modulates their Cellular and Extracellular Vesicle proteomes".</p>
Spatial immunophenotyping of the tumor microenvironment in non-small cell lung cancer
<p>A dataset with spatial immune cell information on a lung cancer cohort from Uppsala University Hospital, Sweden, with anonymized clinical data. For more information please refer to the 'readme' file and the original study (https://doi.org/10.1016/j.ejca.2023.02.012).</p>
Systematic investigation of mitochondrial transfer between cancer and T cells at singlecell level
<p>The benmark datasets about MT transfer, including mtSNV profile, coverage information, cell information and expression information.</p>
Detection of HER2+ Breast Cancer Cells using Bioinspired DNA-Based Signal Amplification
<p>Circulating tumor cells (CTC) are promising biomarkers for metastatic cancer detection and monitoring progression. However, CTC detection remains challenging due to their low frequency and heterogeneity. Herein, we report a bioinspired approach to detect individual cancer cells, based on a signal amplification cascade using a programmable DNA hybridization chain reaction (HCR) circuits. We applied this approach to detect HER2+ cancer cells using the anti-HER2 antibody (trastuzumab) coupled to initiator DNA eliciting a HCR cascade that leads to a fluorescent signal at the cell surface. At 4°C, this HCR detection scheme resulted in highly efficient, specific and sensitive signal amplification of the DNA hairpins specifically on the membrane of the HER2+ cells in a background of HER2- cells and peripheral blood leukocytes, which remained almost non-fluorescent. The results indicate that this system offers a new strategy that may be further developed toward an in vitro diagnostic platform for the sensitive and efficient detection of CTC.</p>
Raw read counts and phased SNP counts for every single cell in the sequencing datasets of the breast cancer patient S1 from "Characterizing allele- and haplotype-specific copy numbers in single cells with CHISEL"
<p>This dataset contains the raw read counts and phased SNP counts for every single cell in the sequencing datasets of breast cancer patient S1 from “Characterizing allele- and haplotype-specific copy numbers in single cells with CHISEL” [Zaccaria & Raphael, 2020]. These data enable the full reproduction of all the results in the related manuscript for breast cancer patient S1. Specifically, the data are provided in two files for every dataset <em>DAT</em> of patient S1 with the following format:</p> <ol> <li><em>DAT.raw</em>_<em>read</em>_<em>counts.bed.gz </em>is a multi-cell BED file containing the raw read counts in the following fields: <ul> <li>CHROMOSOME: the name of a human chromosome</li> <li>START: the starting genomic position of a genomic bin in the chromosome</li> <li>END: the ending genomic position of the genomic bin in the chromosome</li> <li>CELL: the cell barcode that uniquely identifies a cell</li> <li>NORMAL: the raw read count for the specified bin from a matched-normal sample</li> <li>COUNT: the raw read count for the specified bin in the specified cell</li> <li>RDR: the estimated read-depth ratio for the specified bin in the specified cell</li> </ul> </li> <li><em>DAT.phased</em>_<em>snps</em>_<em>counts.pos.gz </em>is a multi-cell POS file containing the phased SNP counts in the following fields: <ul> <li>CHROMOSOME: the name of a human chromosome</li> <li>POS: the genomic position in the chromosome of a germline SNP</li> <li>CELL: the cell barcode that uniquely identifies a cell</li> <li>COUNT_HAPLOTYPE_A: the count of reads that cover the SNP and that belong to haplotype A in the specified cell</li> <li>COUNT_HAPLOTYPE_B: the count of reads that cover the SNP and that belong to haplotype B in the specified cell</li> </ul> </li> </ol> <p>All the files have been compressed using standard <em>gzip</em>.</p>
Raw read counts and phased SNP counts for every single cell in the sequencing datasets of the breast cancer patient S0 from "Characterizing allele- and haplotype-specific copy numbers in single cells with CHISEL"
<p>This dataset contains the raw read counts and phased SNP counts for every single cell in the sequencing datasets of breast cancer patient S0 from “Characterizing allele- and haplotype-specific copy numbers in single cells with CHISEL” [Zaccaria & Raphael, 2020]. These data enable the full reproduction of all the results in the related manuscript for breast cancer patient S0. Specifically, the data are provided in two files for every dataset <em>DAT</em> of patient S0 with the following format:</p> <ol> <li><em>DAT.raw</em>_<em>read</em>_<em>counts.bed.gz </em>is a multi-cell BED file containing the raw read counts in the following fields: <ul> <li>CHROMOSOME: the name of a human chromosome</li> <li>START: the starting genomic position of a genomic bin in the chromosome</li> <li>END: the ending genomic position of the genomic bin in the chromosome</li> <li>CELL: the cell barcode that uniquely identifies a cell</li> <li>NORMAL: the raw read count for the specified bin from a matched-normal sample</li> <li>COUNT: the raw read count for the specified bin in the specified cell</li> <li>RDR: the estimated read-depth ratio for the specified bin in the specified cell</li> </ul> </li> <li><em>DAT.phased</em>_<em>snps</em>_<em>counts.pos.gz </em>is a multi-cell POS file containing the phased SNP counts in the following fields: <ul> <li>CHROMOSOME: the name of a human chromosome</li> <li>POS: the genomic position in the chromosome of a germline SNP</li> <li>CELL: the cell barcode that uniquely identifies a cell</li> <li>COUNT_HAPLOTYPE_A: the count of reads that cover the SNP and that belong to haplotype A in the specified cell</li> <li>COUNT_HAPLOTYPE_B: the count of reads that cover the SNP and that belong to haplotype B in the specified cell</li> </ul> </li> </ol> <p>All the files have been compressed using standard <em>gzip</em>.</p>
Patient-derived and artificial ascites have minor effects on MeT-5A mesothelial cells and do not facilitate ovarian cancer cell adhesion
<p>Raw data of "Patient-derived and artificial ascites have minor effects on MeT-5A mesothelial cells and do not facilitate ovarian cancer cell adhesion".</p>
Supplementary files for Molecular Differences Between Squamous Cell Carcinoma and Adenocarcinoma Cervical Cancer Subtypes: Potential Prognostic Biomarkers
<p>Supplementary files for Molecular Differences Between Squamous Cell Carcinoma and Adenocarcinoma Cervical Cancer Subtypes: Potential Prognostic Biomarkers</p>
Fluorescent Confocal Laser Scanning Microscopy of White Blood Cells, Cancer Cell Line MCF7, and Mixtures of these Cells: A Model System for Circulating Tumor Cell Biomarker Evaluation V.1
<p>This is a confocal laser scanning microscopy data set of white blood cells (leukocytes), the cancer cell line MCF7, and mixtures of these cells acquired on a Zeiss LSM 780 microscope in the University of Colorado Anschutz Medical Campus Advanced Light Microscopy Core. Cells are fluorescently labeled for DNA with DAPI (Sigma D9542), lipids with Bodipy 495/503 (Thermo Fisher D3922), the filament protein cytokeratin (CK) with pan-cytokertain-alexa555 antibodies (Cell Signaling Technologies 3478S) and the surface membrane antigen CD45 with CD45-alexa647 antibodies (Biolegend 304020). Bodipy was excited with a continuous wave (CW) 488 nm laser, alexa555 was excited with CW 561 nm laser, and alexa647 was excited with a CW 633 nm laser. The acquiring instrument does not have a CW 405 nm source so DAPI was excited by two photon process using a Coherent Cameleon ultrafast pulsed laser tuned to 765 nm. The objective used was a Zeiss Plan-Apochromat 20x, 0.8 NA, air.</p> <p>The data consists of 4 channel 8x8 mosaic z-stacks. The Zeiss software performed stitching of the mosaics. These stitched data images are included and marked with _Stitched at the end. Those interested in performing the stitching themselves can do this with the raw data files (without the _Stitched). The jpeg images are processed from the stitched LSM images. The LSM files contain additional meta data on the experiment including power levels and acquisition settings.</p> <p>The _Stiched .lsm files will load in ImageJ (tested with V.1.49) as 4 channel 3 stack images.</p> <p>This data is a model system for evaluating the DNA/Lipids/CK/CD45 biomarker panel to identify circulating tumor cells (CTCs). The D- population of the model is the WBCs and the D+ population is the MCF7 cancer cell line. The amount of separation the biomarker panel plus analysis algorithm can produce between these populations (D+/D-) is an estimate the sensitivity and specificity of the biomarker panel plus algorithm to CTCs.</p> <p>Experiments generating the data were performed over the course of 15 days. Peripheral blood samples were collected from the Gynecological Tissue and Fluid Bank (COMIRB 07-0935 / COMIRB 05-1081) from consenting patients undergoing surgery at the University of Colorado Hospital. Blood samples were used the same day they were collected. Blood samples were collected from 3 patients with benign conditions, labeled WBBN#, and 3 patients with ovarian cancer, labeled WBCA#. We do not expect there to be any difference in the isolated white blood cells samples prepared from the cancer and benign patients. Samples were stored at room temperature until white blood cells were isolated. Mixed samples were prepared by passaging a MCF7 flask and mixing it with isolated white blood cells before fixation. A schedule showing the time duration between collection, processing and imaging is included as “experimental schedule.gif”.</p> <p>The MCF7 cancer cell line was a kind gift from Dr. Heide Ford. Genomic DNA was isolated from the MCF7 cell line after the experiment and sent for cell line authentication. The gDNA was a match to MCF7. The authentication report and data are included in this submission.</p> <p>CD45 antibodies were exhausted on day 7. New antibody was purchased and received on day 8. The day 7 images only has labels for DAPI and Bodipy. The samples prepared with the old antibodies on days 4 and 7 were relabeled and imaged with the new antibodies on days 14 and 15. This labeling was also done to confirm the pan-CK antibodies remained good since they are dim in the MCF7 cells imaged on days 12 and 13. The pan-CK on days 14 and 15 looks the same as it did on days 5 and 7 confirming the antibodies are good.</p> <p>Four of the filters containing cells were not sufficiently flat to be acquired with a 3 slice z-stack so a 5 slice z-stack was used. These files have been zipped to compress them under the 2 GB limit permitted by zenodo.org</p> <p>Further information on how these samples were prepared, processed, and analyzed can be found in our associated 2016 SPIE Photonics West BIOS conference proceeding titled, “Quantitative image cytometry measurements of lipids, DNA, CD45 and cytokeratin for circulating tumor cell identification in a model system”, http://dx.doi.org/10.1117/12.2222317.</p> <p>This work was supported by funding provided to the University of Colorado Cancer Center by the American Cancer Society and awarded as Institutional Research Grant Number 57-001-53, by funding provided by the Defense Advanced Research Projects Agency under grant number N66001-10-4035, and by funding provided by NIH/NCATS Colorado CTSI Grant Number TL1 TR001081. The University of Colorado Anschutz Medical Campus Advanced Light Microscopy Core is also supported in part by NIH/NCATS Colorado CTSI Grant Number UL1 TR001082. The funders had no role in the study design, data collection, analysis, or decision to publish.</p>
Suplementary data, results and scripts: "Reconstruction of Cell-specific Models Capturing the Influence of Metabolism on DNA methylation in Cancer"
<p>This repository contains supplementary data, models and scripts associated with "Reconstruction of Cell-specific Models Capturing the Influence of Metabolism on DNA methylation in Cancer".</p><p>Folders content:</p><p>'data_results_matlabscripts': data, result files and scripts (original python scripts and adapted MATLAB scripts)</p><p>'supplementary_figures': supplementary figures</p><p>'supplementary_tables': supplementary tables</p>
Supplementary material - Optical Diffraction Tomography and Raman Confocal Microscopy for the Investigation of Vacuoles Associated with Cancer Senescent Engulfing Cells
<p>Supplementary material containing the data used in the manuscript "Optical Diffraction Tomography and Raman Confocal Microscopy for the Investigation of Vacuoles Associated with Cancer Senescent Engulfing Cells"</p>
Identification of biomarkers for the early detection of non-small cell lung cancer: a systematic review and meta-analysis
<p>We sought to identify the best biomarkers for the early diagnosis of LC, using a systematic review of seven databases. We identified 79 articles that focused on the identification and assessment of diagnostic biomarkers and then performed a meta-analysis. This work has been submitted for publication.</p>
NanoString dataset for study: Impairment of cancer-associated fibroblasts promotes CD8+ T cell infiltration and enhances sensitivity to immune checkpoint blockade
<p>Pre-processed NanoString mRNA abundance data and associated sample sheet for study:</p> <p>Impairment of cancer-associated fibroblasts promotes CD8+ T cell infiltration and enhances sensitivity to immune checkpoint blockade</p>
Data of FigS7, "The Potential Tumor-Suppressor DHRS7 Inversely Correlates with EGFR Expression in Prostate Cancer Cells and Tumor Samples"
<p>Data of FigS7, “The Potential Tumor-Suppressor DHRS7 Inversely Correlates with EGFR Expression in Prostate Cancer Cells and Tumor Samples”</p> <p>The Dataset (original publication: doi: 10.3390/cancers14133074) contains the original figure as PNG-format (10.3390-cancers14133074_FigS7.PNG). The Corresponding raw data and subsequent data analysis obtained contains one file in txt-format (31003A-179400_10.3390-cancers14133074_SSDHRS7_23_1_M .txt) and one file as csv-format (31003A-179400_10.3390-cancers14133074_SSDHRS7_12_1.csv).</p>
Impact of paclitaxel treatment on the Triple Negative Breast Cancer Cell line HCC1143
<div>Data and code related to Zenodo repository: 10.5281/zenodo.11237850</div> <div> </div> <div>Experimental goal:</div> <div>Evaluate the impact of escalating paclitaxel dose on cell count, nuclear morphology and cellular outcome.</div> <div> </div> <div>Methods:</div> <div>Cells were plated at 3000 cells in 100ul of complete media per well in a 96 well plate (#08-772-225, FisherScientific). After 24 hours, an additional 100ul of either vehicle (0.1% DMSO) or paclitaxel containing complete media was added. After 72 hours cells were fixed with 4% Formaldehyde (#28908, ThermoFisher Scientific) for 15 minutes at room temperature, then permeabilized with 0.3% Triton X-100 (#X100-100ML, Sigma Aldrich) for 10 minutes at room temperature, then washed twice with PBS. Fixed cells were blocked with 1% BSA (A7906-100G, Millipore Sigma) in PBS for 1 hour at room temperature and then stained overnight with 1:100 anti-CDKN2A/p16INK4A+CDKN2B/p15INK4B-AF644 (#ab199756, Abcam), and 1:100 anti-cPARP-AF647 (#6987S, Cell Signaling Technology) or 1:500 anti-TUBB3-AF647 (#ab190575, Abcam) overnight at 4C. Each well was washed twice with room temp PBS then stained with 0.5ug/mL DAPI (4083S, Cell Signaling Technology) in PBS for 15 minutes at room temperature. Following DAPI staining, wells were washed once with PBS, then stained with 1:20,000 HCS CellMask in PBS (Orange: #H32713, Green: #H32714, Invitrogen) for 15 minutes at room temperature. Wells were washed twice with room temperature PBS and then 4 fields of view per well imaged on an InCell 6000 (GE Healthcare). Images were segmented with two custom Cellpose models to segment the nucleus (using parameters: diameter = 45, chan = DAPI, chan2 = Cellmask Orange) and cytoplasm (using parameters: diameter = 90, chan = Cellmask Orange, chan2 = DAPI). Image quantification was performed in R (v4.3.1) using EBImage (v4.42.0), and cells were annotated based on the number of distinct nuclei segmented within each cytoplasmic mask. </div> <div> </div> <div>Included files:</div> <div>row_#_level_1.zip : 6 zip file containing original images from InCell 6000, one zip per row</div> <div>level_2.csv : Data quantified to the nuclear level (cytoplasmic quantification is duplicates across multiplet nuclei)</div> <div>level_3.csv: Data quantified at the cellular level including number of nuclei and stain intensities for segmented compartments</div> <div>platemap.csv: Description of each well from the stained plate</div> <div>cellpose_modelz.zip: Zip file containing the two CellPose models used for segmentation</div> <div>image_quantification.rmd : R markdown file containing code for extracting and quantifying image intensities using the raw images (level_1) and segmentation masks created from cellpose.</div>
Robust estimation of cancer and immune cell-type proportions from bulk tumor ATAC-Seq data.
<p>Bulk ATAC-seq data of tumour samples result in an averaged signal across different cell-types (cancer, stromal, vascular and immune cells). We propose a deconvolution framework called EPIC-ATAC (<a href="https://doi.org/10.7554/eLife.94833.1">https://doi.org/10.7554/eLife.94833.1</a>), which relies on newly identified cell-type specific ATAC-Seq marker peaks and reference profiles for all major cancer-relevant cell-types to predict the proportions of each cell-type.</p> <p>To evaluate EPIC-ATAC, we generated a bulk ATAC-Seq dataset from peripheral blood mononuclear cells (PBMCs) samples, from which the number of cells in each cell-type has been estimated using flow cytometry, as ground truth for cell proportions. The data provided in this Zenodo deposit correspond to:</p> <p>- The raw counts matrix for each peak called in this ATAC-Seq dataset: PBMC_counts.txt</p> <p>- The normalized (TPM-like) counts matrix for each peak called in this ATAC-Seq dataset: PBMC_counts_norm.txt</p> <p>- The cell fractions of each cell type in each sample: PBMC_cell_fractions.txt</p> <p>- The peaks called in each sample using MACS2 (*narrow.peaks): *_normalized.narrowPeak</p> <p>- Bed files listing ATAC-Seq fragments for each sample: *.bed</p> <p>We also evaluated EPIC-ATAC on multiple pseudobulks generated from single-cell ATAC-Seq data. We provide rds files containing the pseudobulks data used in our work for the evaluation of EPIC-ATAC. The rds files are located in the zip file "pseudobulks.zip".</p> <p>The file "additional_data.zip" contains additional files used to generate the reference profiles in EPIC-ATAC and to reproduce the main analyses performed in the manuscript: <a href="https://doi.org/10.7554/eLife.94833.1">https://doi.org/10.7554/eLife.94833.1</a>. These files are required to run the code available on the following GitHub repository: GfellerLab/EPIC-ATAC_manuscript. </p>
Single-cell RNA-seq of breast cancer infiltrating T cells (case 1)
<p>Single cell suspensions were generated from two individual TNBC primary tumor samples (this entry contains case 2) and the viable cells were FACS sorted for CD3<sup>+</sup> T cells. Sorted cells were then counted and assessed for viability. Single cell library preparation was carried out as per the 10X Genomics Chromium Single cell protocol for the v2 reagent kit (10X Genomics, Pleasanton, CA, USA). Cell suspensions were loaded onto a Chromium Single Cell Chip along with the reverse transcription (RT) mastermix and single cell 3’ gel beads. Following generation of single cell gel bead-in-emulsions (GEMs), reverse transcription was performed using a C1000 Touch Thermal Cycler with a Deep Well Reaction Module (Bio-Rad Laboratories, Hercules, CA, USA). Amplified cDNA was purified using SPRIselect beads (Beckman Coulter, Lane Cove, NSW, Australia) and sheared to approximately 200bp with a Covaris S2 instrument (Covaris, Woburn, MA, USA) using the manufacturer’s recommended parameters. Sequencing libraries were generated with unique sample indices (SI) for each sample. Libraries were sequenced on an Illumina HiSeq 2500 High Output Mode using V4 clustering and sequencing chemistry.</p> <p>This dataset contains the raw .bcl files.</p>
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These curated guides explain access requirements, typical timelines, costs, and reuse considerations for widely used research datasets.
Allen Brain Atlas
Allen Brain Atlas is an Allen Institute collection of brain map atlases, datasets, APIs, and analysis tools covering mouse, human, and non-human primate brain resources.
Annotated Behaviour and Observability Dataset (ABODe)
ABODe is a University of Edinburgh DataShare dataset for behavior classification in group-housed mice using home-cage video, identities, bounding boxes, ground-plate positions, and annotator labels.
DANDI Archive for NWB datasets
DANDI is a BRAIN Initiative archive for publishing and sharing neurophysiology data, including electrophysiology, optophysiology, and behavioral data packaged as NWB and related standards.
International Brain Laboratory public data
The International Brain Laboratory public data releases expose standardized mouse decision-making experiments, including Neuropixels recordings, widefield calcium imaging, behavior, and session metadata accessed through the ONE API.
OpenNeuro
OpenNeuro is a free, open platform for sharing neuroimaging datasets, with public search, dataset pages, and download paths for web, S3, DataLad, and the OpenNeuro CLI.