AutoRELACS: Automated Generation And Analysis of Ultra-parallel ChIP-seq
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens Drosophila melanogaster
PROVIDER: GSE147042 | GEO | 2020/07/28
REPOSITORIES: GEO
Similar Datasets
Project description:A collection of digital automated proteomic sample preparation protocols detail three optimized step-by-step methods to: (A) lyse Gram-negative bacteria and fungal cells; (B) quantify the amount of protein extracted; and (C) normalize the amount of protein and set up tryptic digestion. These protocols have been developed to facilitate rapid, low variance sample preparation of hundreds of samples, be easily implemented on widely-available Beckman-Coulter Biomek automated liquid handlers, and allow flexibility for future protocol development
2022-02-14 | PXD029122 | Pride
Project description:High-throughput has become a substantial need in many omics investigations. In proteomics the sample preparation workflow consists of multiple steps, whereby with each additional manual step more bias is added to the sample. Especially for label-free quantification experiments this drastically reduces the outcome. Here, a positive pressure workstation was evaluated to increase automation of sample preparation and decrease input time and consumables. The semi-automated workflow with the Resolvex A200 workstation including the sorbents Maestro and WWP2 performed equally well to the routinely conducted manual workflow, with similar technical variability in MSMS-based identifications of peptides and proteins. Both sorbents produced highly reproducible results within a range of 10-300 µg of peptide starting material. A direct, on-column sample preparation using the NBE columns - without conditioning and equilibration of the sorbents -works generally robust, whereby consumables and time are drastically reduced.
2021-01-25 | PXD018444 | Pride
Project description:The submitted files contain ChIP-seq data for the p300 transcriptional coactivator in GM12878 cells and for the NRSF transcription factor in GM12878 and Jurkat cells generated using a fully automated robotic chromatin immunoprecipitation protocol. Cells were fixed using 1% formaldehyde (NRSF samples) or 1% formaldehyde at 37C (p300 samples).
2014-06-01 | GSE53366 | GEO
Project description:A new approach to sample preparation and enzymatic digestion in bottom-up proteomics has been developed using alginate-based hydrogel entrapment of enzymes. This hydrogel facilitates rapid and room-temperature digestions with multienzyme capabilities. Three methodologies were tested: within microcentrifuge tubes, in situ pipette tips, and automated robotic liquid handling. Factorial experimental design identified a 1 h, room temperature, pepsin–trypsin dual-enzyme digestion as optimal for sequence coverage and protein group identification, comparable to a gold-standard overnight proteomic protocol. This method promises significant advancements in proteomic analysis by enhancing reusability, speed, throughput, convenience, and cost-effectiveness, without hindering digestion efficiency.
2024-11-12 | PXD052369 | Pride
Project description:Recent advances in stem cell technology have led to the development of three-dimensional (3D) culture systems called organoids, which have fueled hopes to bring about the next generation of more physiologically relevant high throughput screens (HTS). However, the adaptation of established organoid protocols for HTS applications has so far been elusive. Here, we present a fully scalable, HTS-compatible workflow for the automated generation, maintenance, whole mount staining, clearing, and optical analysis of human neural organoids generated from neural precursor cells in a standard 96-well format. By combining organoid generation and analysis steps in an automated fashion, we can perform quantitative whole-organoid high content imaging with single cell resolution. The resulting organoids are highly homogeneous with regard to their morphology, size, global gene expression, cellular composition, and structure. Calcium imaging suggests organoid-wide synchronized functional coupling. The scalability of our approach has the potential to form the basis for 3D tissue-based screening in a variety of applications including drug development, toxicology studies, and disease modeling.
2020-09-28 | GSE119060 | GEO
Project description:BACKGROUND: Array Comparative Genomic Hybridization (aCGH) is a rapidly evolving technology that still lacks complete standardization. Yet, it is of great importance to obtain robust and reproducible data to enable meaningful multiple hybridization comparisons. Special difficulties arise when aCGH is performed on archival formalin-fixed, paraffin-embedded (FFPE) tissue due to its variable DNA quality. Recently, we have developed an effective DNA quality test that predicts suitability of archival samples for BAC aCGH. METHODS: In this report, we first used DNA from a cancer cell-line (SKBR3) to optimize the aCGH protocol for automated hybridization, and subsequently optimized and validated the procedure for FFPE breast cancer samples. We aimed for highest throughput, accuracy, and reproducibility applicable to FFPE samples, which can also be important in future diagnostic use. RESULTS: Our protocol of automated array-CGH on archival FFPE ULS-labeled DNA showed very similar results compared with published data and our previous manual hybridization method. CONCLUSION: This report combines automated aCGH on unamplified archival FFPE DNA using non-enzymatic ULS labeling, and describes an optimized protocol for this combination resulting in improved quality and reproducibility.
2007-02-23 | GSE7122 | GEO
Project description:BACKGROUND: Array Comparative Genomic Hybridization (aCGH) is a rapidly evolving technology that still lacks complete standardization. Yet, it is of great importance to obtain robust and reproducible data to enable meaningful multiple hybridization comparisons. Special difficulties arise when aCGH is performed on archival formalin-fixed, paraffin-embedded (FFPE) tissue due to its variable DNA quality. Recently, we have developed an effective DNA quality test that predicts suitability of archival samples for BAC aCGH. METHODS: In this report, we first used DNA from a cancer cell-line (SKBR3) to optimize the aCGH protocol for automated hybridization, and subsequently optimized and validated the procedure for FFPE breast cancer samples. We aimed for highest throughput, accuracy, and reproducibility applicable to FFPE samples, which can also be important in future diagnostic use. RESULTS: Our protocol of automated array-CGH on archival FFPE ULS-labeled DNA showed very similar results compared with published data and our previous manual hybridization method. CONCLUSION: This report combines automated aCGH on unamplified archival FFPE DNA using non-enzymatic ULS labeling, and describes an optimized protocol for this combination resulting in improved quality and reproducibility. In this study, we optimized the BAC araay-CGH protocol for automated hybridization for FFPE breast cancer samples. We have tested hybridization temperature and duration, different hybridization buffer conditions, and post-hybridization washing.
2010-06-12 | E-GEOD-7122 | biostudies-arrayexpress
Project description:Existing protocols for full-length single-cell RNA sequencing (scRNA-seq) produce libraries of high complexity (thousands of distinct genes) with outstanding sensitivity and specificity of transcript quantification. These full-length libraries have the advantage of allowing probing of transcript isoforms, are informative regarding single nucleotide polymorphisms, and allow assembly of the VDJ region of the T- and B-cell receptor sequences. Since full length protocols are mostly plate-based at present, they are also suited to profiling cell types where cell numbers are limiting, such as rare cell types during development for instance. A disadvantage of these methods has been the scalability and cost of the experiments, which has limited their popularity as compared to droplet-based and nanowell approaches. Here, we describe an automated protocol for full-length scRNA-seq, including both an in-house automated SMART-seq2 protocol, and a commercial kit-based workflow. We discuss these two protocols in terms of ease-of-use, equipment requirements, running time, cost per sample and sequencing quality. By benchmarking the lysis buffers, reverse transcription enzymes and their combinations, we propose an optimized in-house automated protocol with dramatically reduced cost. These pipelines have been employed successfully for several research projects allied with the Human Cell Atlas initiative (www.humancellatlas.org) and are available on protocols.io.
2020-12-31 | E-MTAB-9345 | biostudies-arrayexpress
Project description:Spatial tissue proteomics integrating whole-slide imaging, laser microdissection and ultrasensitive mass spectrometry is a powerful approach to link cellular phenotypes to functional proteome states in (patho)physiology. To be applicable to large patient cohorts and low sample input amounts, including single-cell applications, loss-minimized and streamlined end-to-end workflows are key. We here introduce an automated sample preparation protocol for laser microdissected samples utilizing the cellenONE® robotic system, which has the capacity to process 192 samples in three hours. Following laser microdissection collection directly into the proteoCHIP LF 48 or EVO 96 chip, our optimized protocol facilitates lysis, formalin de-crosslinking and tryptic digest of low-input archival tissue samples. The seamless integration with the Evosep ONE LC system by centrifugation allows ‘on-the-fly’ sample clean-up, particularly pertinent for laser microdissected workflows. We validate our method in human tonsil archival tissue, where we profile proteomes of spatially-defined B-cell, T-cell and epithelial microregions of 4,000 µm2 to a depth of ~2,000 proteins and with high cell type specificity. We finally provide detailed equipment templates and experimental guidelines for broad accessibility.
2024-03-22 | PXD047762 | Pride
Project description:To identify insulin responsive genes in humans, in the first protocol, skeletal muscle biopsies from six non-diabetic subjects were obtained before and after a two-hour of hyperinsulinaemic (infusion rate 40 mU/m2/min) euglycemic clamp. A variable infusion of glucose (180 g/l) enriched with tritiated glucose (100 μCi/500 ml) maintained euglycemia during insulin infusion, with monitoring of plasma glucose concentration every 5 to 10 min during the basal and clamp periods using an automated glucose oxidation method (Glucose Analyzer 2, Beckman Instruments, Fullerton, CA). In the second protocol, skeletal muscle biopsies from six non-diabetic subjects were obtained before and after a 3-hour hyperinsulinemic (infusion rate 40 mU/m2/min) euglycemic clamp in order to increase the effects of insulin on gene expression. A variable infusion of glucose (180 g/l) was used to maintain euglycemia during insulin infusion with monitoring of plasma glucose concentration every 5 to 10 min using an automated glucose oxidation method (Glucose Analyzer 2, Beckman Instruments, Fullerton, CA). Keywords: dose response
2007-03-01 | GSE7146 | GEO