4/3/2023 0 Comments Flowjo panel analysisLeveraging this template and command line, preprocessing analysis can be rapidly and consistently applied to all samples in this study. This gating analysis was applied to all samples stained with the T-cell cytokine reagent panel ( Figure 2), generating reports for QA that flagged outliers, and gated T-cell data (or further subsetting to analyze CD4+ and CD8+ T cells), and exported this pre-processed data for clustering analysis. In this case T cells of interest were examined by setting up a hierarchy of gates (in order) on singlet cells, a simple clean-up gate to remove the cells on the axis, and a “magnetic” CD3+ gate. Apply automated gating at another level of the analysis to mine the data for additional informationīy creating a preprocessing template, some simple and very common actions to the data can be applied. Apply manual expert gates in a traditional manner.Process many datasets in the same manner.In this case, FlowJo is combined with two freely available clustering tools to perform a high-throughput analysis of T-cell intracellular cytokine assays after simian immunodeficiency virus (SIV) infection in order to: Automating flow cytometry analysis with FlowJo templates and command line: FCS files generated by a cytometer are analyzed using a FlowJo template through command line: Reports and analysis workspaces are saved to a user-set location for review.Ĭase Study 2: HT Analysis Combining Manual Gating and ClusteringĪutomated analysis of cytometry data has been advanced significantly by a variety of flow cytometry-specific clustering algorithms, which can be integrated as part of a workflow leveraging FlowJo’s gating, templates, and visualizations. With a few lines of text, command line on Mac, PC, or LINUX is used to point FlowJo to the files to be analyzed, perform auto-compensation (signal processing), calculate gates and statistics, and produce output files such as batch spreadsheet and graph reports ( Figure 1).įigure 1. The final piece of the automation puzzle comes through the ability to launch FlowJo without a user interface or human intervention. Thus, when data is placed into a template, samples are organized into groups, analyses executed, reports generated, and analysis workspaces created and saved for review.Ĭommand Line Reduces Time from Collection to Reports Templates Execute Consistent Analysis for Repeated SOPįlowJo has the ability to generate workspace templates, which saves sample organization, analysis strategy, and report output options without the flow cytometry data (FCS) files. Data organization and management is a critical component in any data-analysis strategy, and groups organize related samples in a database-like structure, add metadata, apply analysis, and provide the underlying organization for batched reports. Groups Organize Samples and Enable Batch AnalysisĪn intuitive user interface facilitates rapid setup of an analysis strategy, including statistics, hierarchical cellular subsetting through “gating” (boundaries which isolate events), and batch application to related samples using a patented groups structure. Increase lab capability for high-throughput and high-content assays.Reduce downtime caused by manual analysis and report generation.Maintain consistent analysis for repeated SOP.However, with three key tools described in this article-group organization, templates, and command line-FlowJo provided an automation solution, addressing the bottleneck with a scalable solution that can: Manual organization, analysis, and report generation with these FCS files would produce a large bottleneck, slowing the progress of trial reporting and publication. The four-arm, randomized, double-blind, placebo-controlled study had nine time points, eight panels examining immune cell activation and one negative control panel-producing 10,991 flow cytometry standard (FCS) data files. We recently analyzed a large dataset from a clinical trial of anti-IgE (omalizumab), which examined whether combination therapy with omalizumab and allergen immunotherapy for ragweed reduced seasonal allergic rhinitis compared with immunotherapy alone. The newest version of Tree Star’s FlowJo visualization and analysis software for flow cytometry advances research in flow cytometry by providing robust tools for analysis automation and a bridge to repository/cloud-based data.Ĭase Study 1: Automating Analysis of Clinical Trial Data As the throughput, applications, and content of flow cytometry have increased exponentially over the past several years, cytometrists are beginning to embrace cloud-based data-management solutions. Clinical trials and high-throughput flow cytometry produce large amounts of data, leading to an analysis bottleneck unaddressed by commercially available cytometry analysis software.
0 Comments
Leave a Reply. |
AuthorWrite something about yourself. No need to be fancy, just an overview. ArchivesCategories |