Clonotype count and frequency measures are used for clonality calculations. Immune repertoire measuresįirst, clonotypes are defined as clonally related cells derived from a common progenitor cell. For example, differences in the samples of healthy and diseased individuals can be identified. In addition, these two measures can provide information about the composition of the adaptive immune response. Therefore, several different immune repertoire measures are used in the community as the analysis of clonality and diversity of immune repertoires are of fundamental interest. Sequencing of the TCR and IG repertoire for deciphering the V(D)J gene segments and CDR3 region allows for the quantitative description of the immune repertoire and its clonal composition. Owing to the increased speed of DNA and RNA sequencing and continuous improvement of read length, usage of such high-throughput systems results in large amounts of data. The advantages of this methodology, including lower costs and effort, supersede the automated Sanger method in clinical and scientific research.
Next-generation sequencing (NGS) is the current state of the art high-throughput technology for DNA sequencing. Modern sequencing methods allow determining the V(D)J gene segment nucleotide sequences. V(D)J rearrangement in early T and B cell development contributes to the diversity of the immune system. These peptides are commonly presented by major histocompatibility complexes (MHC) and belong to potentially pathogenic microorganisms or endogenous molecules. These unique receptors and especially the third complementary-determining region (CDR3) are necessary to recognize and bind different peptides. This V(D)J recombination is important for the unique antigen receptors such as T cell receptors (TCR) and immunoglobulins (IG). An essential aspect of the major lymphocyte types, T and B cells, is the ability of random rearrangements of the variable (V), diversity (D), and joining (J) gene segments of the lymphocyte receptor. Amongst other aspects, lymphocytes protect us from potentially pathogenic microorganisms and cancer cells. Lymphocytes play an essential role in the human immune system. The implementation is open-source and available on.
In summary, IMDA reduces the effort usually required for immune repertoire data analysis by providing an automated workflow for processing raw NGS data into immune repertoires and subsequent analysis. IMDA further supports the detection of sample swaps and cross-sample contamination that potentially occurred during sample preparation. The provided output directly facilitates the interpretation of input data and includes information about clonality, diversity, clonotype overlap as well as similarity, and V(D)J gene segment usage. The IMDA workflow focus on quality control and ease of use for non-computer scientists. IMDA constructs TCR and IG repertoire data from raw NGS reads and facilitates descriptive data analysis and comparison of immune repertoires. IMDA further generates an output file including key figures for all samples, designed to serve as input for machine learning frameworks to find models for differentiating between specific traits of samples. IMDA reports all relevant information in a compact summary containing visualizations, calculations, and sample details, all of which serve for a more detailed overview. These include commonly used clonality and diversity measures, as well as indicators for V(D)J gene segment usage and between sample similarity. It provides methods for automated pre-processing of barcoded and UMI tagged immune repertoire NGS data, facilitates the assembly of clonotypes and calculates key figures for describing the immune repertoire.
Lossius a immune repertoire software#
IMDA unites the functionality from carefully selected immune repertoire analysis software tools and covers the whole spectrum from initial quality control up to the comparison of multiple immune repertoires. ImmunoDataAnalyzer (IMDA) is a pipeline we have developed for automatizing the analysis of immunological NGS data. Accurate processing, evaluation and visualization of immune repertoire NGS data is important for better understanding immune responses and immunological behavior. A main objective is the analysis of the V(D)J recombination defining the structure and specificity of the immune repertoire. Research in the field of T cell receptor (TCR) and immunoglobulin (IG) repertoires aids in understanding immunological diseases. Among others NGS enables the in-depth analysis of immune repertoires. Next-generation sequencing (NGS) is nowadays the most used high-throughput technology for DNA sequencing.
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