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55 results for “immunofluorescence”
Antibody Validation for Cyclical Immunofluorescence Microscopy of Human Kidneys (Part 1, Figures 31 and 33)
<p>This dataset includes Cyclical Immunofluorescnce (CyCIF) images (15-20 channels) of frozen human kidney sections interogated with a panel of 12 validated antibodies designed to evaluate renal tubular cell segmentation (Figures 31 and 32), an a panel of 12 validated antibodies and a lectin designed to evaluate glomerular and glomerulus-associated structures in the normal human kidney (Figure 33). We have attached an excel file (Supplemental Table 2) that includes all of the experimental and (de-identified) patient metadata associated with these images, with information and comments about each of the channel images, the antibodies used, CyCIF cycles, and the cell types and extracellular matrix compartments identified with these combinations of antibodies. Because of the sixze of the images, we have divided this into two separate datasets (Antibody Validation for Cyclical Immunofluorescence Parts 1 and 2). </p>
Antibody Validation for Immunofluorescence Microscopy of Human Kidneys (Part 2, Figures 16-30)
<p>This dataset includes multiplex immunofluorescence images (mostly 3+1 channels) of frozen human kidney sections that have been used to validate a panel of 27 antibodies and 1 lectin designed to define the main cellular and extracellular matrix (ECM) comparments in the normal human kidney. We have attached an excel file (Supplemental Table 1) that includes all of the experimental and (de-identified) patient metadata associated with these images with information, and comments about each of the images, the antibodies used, and the cell types and ECM compoartments identified using these antibodies. Because of the size and number of images used for these studies, we have divided this into two separate datasets (Antibody validation studies Parts 1 and 2). </p> <p>A subset of these antibodies have also been evaluated for both 2D and 3D cyclical immunofluorescence studies that have been included in separate datasets under this umbrella "community". These are identified in the "Antibodies used" tab in Supplemental Table 1 (CyCIF Cycles). </p>
Antibody Validation for Immunofluorescence Microscopy of Human Kidneys (Part 1, Figures 1-15)
<p>This dataset includes multiplex immunofloresence images (mostly 3+1 channels) of frozen human kidney sections that have been used to validate a panel of 27 antibodies and 1 lectin designed to define the main cellular and extracellular matrix compartments in the normal human kidney. We have attached an excel file (Supplemental Table 1) that includes all of the experimental and (deidentified) patient metadata associated with these images with information and comments about each of the images, the antibodies used, and the cell types and ECM compartments identified using these antibodies. Because of the size and number of images used for these validation studies, we have divided this into two separate datasets (Antibody validation studies Parts 1 and 2). </p> <p>A subset of these antibodies have also been evaluated for both 2D and 3D cyclical immunofluoresecnce studies that have been included in separate datasets under this umbrella "community". These are identfied in the "Antibodies used" tab in Supplemental Table 1 (CyCIF Cycles). </p> <p> </p> <p> </p> <p> </p>
msiFlow: Automated Workflows for Reproducible and Scalable Multimodal Mass Spectrometry Imaging and Immunofluorescence Microscopy Data Processing and Analysis
<p>This record contains example and result data of msiFlow.</p> <p>msiFlow is a collection of automated workflows for reproducible and scalable multimodal mass spectrometry imaging (MSI) and immunofluorescence microscopy (IFM) data processing and analysis. Using an experimental mouse model for urinary tract infection, induced by uropathogenic E.coli (UPEC), we generated data by</p> <ul> <li>matrix-assisted laser desorption ionisation mass spectrometry imaging with laser-induced postionisation (MALDI-2 MSI) using the Bruker timsTOFfleX instrument</li> <li>transmission-mode MALDI-2 MSI (t-MALDI-2)</li> <li>immunofluorescence microscopy (IFM) using the MACSima system from Miltenyi </li> </ul> <p>msiFlow was tested on MALDI-2 MSI, t-MALDI-2 MSI and IFM data of control and UPEC-infected mouse bladder sections. In IFM we used Ly6G and actin for staining neutrophils and the muscle layer. We validated msiFlow on MALDI MSI data of bone marrow (BM)-derived neutrophils. Tentative lipid annotations were validated by MALDI DDA MSI and MALDI MS/MS. All data used and results generated by msiFlow are included in this dataset (besides the intermediate results of the MALDI-2 preprocessing due to data size).</p> <p>The dataset contains the following zip files:</p> <table> <tbody> <tr> <td><strong>zip file</strong></td> <td><strong>description</strong></td> </tr> <tr> <td>ly6g_heterogeneity.zip</td> <td>example and result data (Ly6G clusters) for molecular_heterogeneity_flow</td> </tr> <tr> <td>if_segmentation.zip</td> <td>example and result data (Ly6G segmentation) for if_segmentation_flow</td> </tr> <tr> <td>ly6g_heterogeneity_signatures.zip</td> <td>example and result data (lipids for Ly6G clusters) for molecular_signatures_flow</td> </tr> <tr> <td>ly6g_molecular_signatures.zip</td> <td>example and result data (lipids for Ly6G) for molecular_signatures_flow</td> </tr> <tr> <td>msi_if_registration.zip</td> <td>example and result data for msi_if_registration_flow</td> </tr> <tr> <td>msi_segmentation.zip</td> <td>example and result data (segmented MSI bladder data) for msi_segmentation_flow</td> </tr> <tr> <td>region_group_analysis.zip</td> <td>example and result data (regulated lipids in different bladder tissue regions) for region_group_analysis_flow</td> </tr> <tr> <td>macsima.zip</td> <td>raw IFM data of UPEC-infected bladders containing Ly6G, actin and autofluorescence images</td> </tr> <tr> <td>maldi-bm-neutrophils.zip</td> <td>raw and pre-processed MALDI MSI data of BM-derived neutrophils</td> </tr> <tr> <td>t-maldi-2.zip</td> <td>raw t-MALDI-2 MSI data of a UPEC-infected bladder section</td> </tr> <tr> <td>maldi-2-<em>group-sampleno</em>.zip</td> <td>raw MALDI-2 MSI data of a control/UPEC bladder section</td> </tr> <tr> <td>MALDI_DDA_MSI.zip</td> <td>raw MALDI MSI data acquired in DDA mode</td> </tr> <tr> <td>TIMS_MS_MS.zip</td> <td>raw MALDI TIMS MS/MS data</td> </tr> </tbody> </table> <p> </p>
Multiplex immunofluorescence ROIs related to Van Hijfte et al. 2024
<p>Multiplex immunofluorescence ROIs from IDH1-R132H mutant astrocytomas. </p> <p>For more information see associated publication. </p> <p>Panel:</p> <ul> <li>Opal 520: CD3</li> <li>Opal 540: CD68</li> <li>Opal 570: CD8</li> <li>Opal 620: CD56</li> <li>Opal 650: CD20</li> <li>Opal 690: IDH1-R132H</li> </ul>
Raw microscopy data of split-GFP and immunofluorescence experiments
<p><span>Mitochondria critically rely on protein import and its tight regulation. Here, we found </span><span>that the complex I assembly factor NDUFAF8 follows a two-step import pathway </span><span>linking IMS and matrix import systems. A weak targeting sequence drives TIM23-</span><span>dependent NDUFAF8 matrix import, and</span> <span>en route</span> <span>allows exposure to the IMS </span><span>disulphide relay which oxidizes NDUFAF8. Import is closely surveyed by proteases: </span><span>YME1L prevents accumulation of excess NDUFAF8 in the IMS, while CLPP </span><span>degrades reduced NDUFAF8 in the matrix. Therefore, NDUFAF8 can only fulfil its </span><span>function in complex I biogenesis if both oxidation in the IMS and subsequent matrix </span><span>import work efficiently. We propose that the two-step import pathway for NDUFAF8 </span><span>allows to integrate the activity of matrix complex I biogenesis pathways with the </span><span>activity of the mitochondrial disulphide relay system in the IMS. Such coordination </span><span>might not be limited to NDUFAF8 as we identified further proteins that can follow </span><span>such a two-step import pathway. The raw microscopy data provided in this dataset were obtained to visualize the localization of NDUFAF8 and further proteins following a two-step import pathway.</span><span><br></span></p>
Immunofluorescence_bladder tissue
<p>Immunofluorescence images of bladder tissue</p>
Immunofluorescence pictures related to Figure 4B-C
<p>This dataset includes the original images obtained by immunofluorescence against E-cadherin and vimentin on somite progenitors derived from human induced pluripotent stem cells. The E-cadherin antibody is coupled to Alexa Fluor 488 (green fluorescence), and the vimentin antibody is coupled to eFluor 570 (red fluorescence). Images were acquired with a Nikon A1 RSi confocal microscope and a 20x objective. The .nd2 format can be opened with standard image processing softwares, such as the open source and free software imageJ, available here: https://imagej.nih.gov/ij/download.html</p> <p>The dataset also includes a .csv file providing the key between the image_ID, cell line and quantification of E-cadherin-positive cells as percentages of total cells.</p>
Crosstalk assessment for multi-colour immunofluorescence of Histone 3 and Polymerase II post-translational modifications in pluripotent zebrafish embryos
<p>Microscopy images recorded to assess the extent of crosstalk from the detection channels of H3K27ac and recruited RNA polymerase II (Serine 5 phosphorylation of the C-terminal domain heptad repeat of subunit 1) to the detection channel of elongating RNA polymerase II (Serine 5 phosphorylation of the C-terminal domain heptad repeat of subunit 1). Sample preparation and image recording was carried out jointly by Süheyla Eroğlu-Kayikci, Elisa Kämmer, and Lennart Hilbert.</p>
Immunofluorescence images related to Figure 3A-B
<p>This dataset includes the native images obtained after immunofluorescence against myosin heavy chain and alpha-actinin on myotubes derived from hiPSCs in which the DMD gene was mutated with CRISPR Cas9. Images were acquired with a Nikon A1 RSi confocal microscope and a 20x objective. The .nd2 format can be opened with standard image processing softwares, such as the open source and free software imageJ, available here: https://imagej.nih.gov/ij/download.html, or the QuPath software, available here: https://qupath.github.io/</p>
High-content imaging, immunoblot and immunofluorescence data related to: Caprin-1 binding to the critical stress granule protein G3BP1 is influenced by pH
G3BP is the central node within stress-induced protein–RNA interaction networks known as stress granules (SGs). The SG-associated proteins Caprin-1 and USP10 bind mutually exclusively to the NTF2 domain of G3BP1, promoting and inhibiting SG formation, respectively. Herein, we present the crystal structure of G3BP1-NTF2 in complex with a Caprin-1-derived short linear motif (SLiM). Caprin-1 interacts with His-31 and His-62 within a third NTF2-binding site outside those covered by USP10, as confirmed using biochemical and biophysical-binding assays. Nano-differential scanning fluorimetry revealed reduced thermal stability of G3BP1-NTF2 at acidic pH. This destabilization was counterbalanced significantly better by bound USP10 than Caprin-1. The G3BP1/USP10 complex immunoprecipated from human U2OS cells was more resistant to acidic buffer washes than G3BP1/Caprin-1. Acidification of cellular condensates by approximately 0.5 units relative to the cytosol was detected by ratiometric fluorescence analysis of pHluorin2 fused to G3BP1. Cells expressing a Caprin-1/FGDF chimera with higher G3BP1-binding affinity had reduced Caprin-1 levels and slightly reduced condensate sizes. This unexpected finding may suggest that binding of the USP10-derived SLiM to NTF2 reduces the propensity of G3BP1 to enter condensates.
Raw microscopy data of split-GFP and immunofluorescence experiments
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High-content imaging, immunoblot and immunofluorescence data related to: Caprin-1 binding to the critical stress granule protein G3BP1 is influenced by pH
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Immunofluorescence on (Frozen and Paraffin) Renal Biopsies
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3-D Cyclical Immunofluorescence Microscopy of the Human Kidney
<p>This dataset includes 3-D images that have been generated by co-registration of CyCIF images from 58 sequential sections obtained from a frozen human kidney block that were interrogated with a panel of 12 validated antibodies and a lectin designed to evaluate glomerular and glomerulus-associated structures in the human kidney. We have attached an excel file (Supplemental Table 3) that includes all of the experimental and (de-identified) patient metadata associated with these images, with information and comments about each of the channel images, information aboiut each of the tissue sections stained, including documention of instances where tissues were folded, cycles were out of focus, or the tissues had been damaged during the CyCIF cycles, as well as the antibodies used, and the cell types and extracellular matrix compartments identified with these combinations of antibodies. GIF images were generated using four antibody channels (a-SMA, CD31, NaK-ATPase, and Podocalyxin) from the panel of 13 markers to best visualize 3-D glomerular, tubular (TAL), and vascular segmentation of the human kidney (Figures 34 and 35).</p> <p> </p> <p> </p> <p> </p> <p> </p> <p> </p> <p> </p>
Figure S3: Immunofluorescence of mitochondria alongside LC3B [FINAL]
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STED immunofluorescence imaging of histone protein H3K27 in a 2-cell stage mice embryo
<p>In either confocal or STED mode, immunofluorescence was performed as previously described in [1], using an upright Zeiss microscope with a mounted STEDYCON module and a Zeiss 100x 1.46 NA objective. Primary antibodies used were anti-H3K27ac (Active Motif, 39034) and anti-H3K27me3 (Abcam, ab6002), both at 1:200 dilution. Secondary anti-mouse and anti-rabbit antibodies used were labeled with STAR Red (Sigma-Aldrich, 52283) and STAR Orange (Sigma-Aldrich, 41367), respectively. Excitation was provided by 640 nm (@3%) and 775 nm (@96.5%) lasers for the STAR Red channel, whereas 561 nm (@7.8%) and 775 nm (@100%) lasers were used for the STAR Orange channel. Imaging of the samples was performed with 5 µs pixel dwell time, 64 µm pinhole aperture, 15-line accumulations and a pixel size of 20 nm.</p> <p>Experimental procedures were approved by the EMBL Rome Animal Facility in accordance with European and Italian legislations.</p> <p>1. Bošković, A., Bender, A., Gall, L., Ziegler-Birling, C., Beaujean, N., & Torres-Padilla, M.-E. Analysis of active chromatin modifications in early mammalian embryos reveals uncoupling of h2a.z acetylation and h3k36 trimethylation from embryonic genome activation. Epigenetics 7, 747–757 (2012).</p>
ROS-Specific Huntingtin Interactions: Optimization of Poly ADP Ribose Detection by Immunofluorescence
<p>Optimization of conditions for the detection of poly ADP ribose by immunofluorescence.</p>
CD3/CD31/LAMA2 immunofluorescence multiplex data related to Van Hijfte et al.
<p>CD3/CD31/LAMA2 immunofluorescence multiplex images and GeoJSON files used for analysis in R. For more information see related publication.</p>
Immunofluorescence images: Inflammasome activation leads to cDC1-independent cross-priming of CD8 T cells by epithelial cell derived antigen
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