b'Next-Generation Sequencing (NGS) | CE-IVD AssaysTRB Assay Next-Generation Sequencing (NGS) | CE-IVD AssaysLymphoTrack Dx TRB AssayAssay Description testing for TRG gene rearrangements only. As a result, combining The LymphoTrack Dx TRB Assay for the Illumina MiSeq is an inthe analysis of TRB and TRG loci increases the sensitivity of clonality vitro diagnostic product intended for next-generation sequencingdetection. (NGS) based determination of the frequency distribution of TRB geneThe human T-cell receptor beta (TRB) gene locus on chromosome rearrangements in patients suspected of having lymphoproliferative7 (7q34) includes 65 V (variable) gene segments, followed by two disease. This assay aids in the identification of lymphoproliferativeseparate clusters of genes each containing a D (diversity) gene, disorders. several J (joining) genes, and a C (constant) region spread over 685 kilobases. The 2 C genes, TRBC1 and TRBC2, encode highly Summary and Explanation of the Test homologous products with no functional difference.During lymphoid cell development, antigen receptor genes undergo This assay utilizes a single multiplex master mix to target conservedsomatic gene rearrangements. 1Specifically, during T-cell development V and J regions of the human TRB gene locus described in lymphoidgenes encoding TRB molecules are assembled from multiple malignancies, thereby reducing sample DNA requirements andpolymorphic gene segments that generate V - D - J combinations simplifying the testing workflow. The LymphoTrack Dx TRB masterunique in both length and sequence. 2mix primers are designed with Illumina adapters and 8 (Kit A) or 24 (Panel) different indices. This allows amplicons generated fromSince leukemias and lymphomas originate from the malignant different indexed TRB master mixes to be pooled into a single librarytransformation of individual lymphoid cells, all leukemias and for loading onto one MiSeq flow cell. lymphomas generally share one or more cell-specific or clonal The associated LymphoTrack Dx Software is capable of sortingantigen receptor gene rearrangements. Therefore, tests that detect complex NGS data by gene target, offering a second layer ofTRB clonal rearrangements can be useful in the study of B- andmultiplexing. This provides users the ability to reduce per sampleT-cell malignancies. testing costs by sequencing amplicons from any LymphoTrack Dx MiSeq Assay (e.g. TRB, TRG, IGH, IGK) at the same time. In addition,Note: For a more thorough explanation of the locus and the targeted deep sequencing strategy, please refer to Principle of Immunoglobulin the LymphoTrack Dx Software provides an easy and streamlinedand T-Cell Receptor Gene Rearrangement. 2method for visualization of data. Guidelines to interpret samples for evidence or no evidence of clonality are included in the instructionsfor use provided with each kit. Specimen RequirementPositive clonal and negative polyclonal DNA controls are included in kits. 50 ng of high-quality genomic DNA.Background ReferencesThe LymphoTrack Dx TRB Assays represent a significant improvement1. Tonegawa, S. (1983). Nature 302, 575-581. over fragment analysis methods for clonality testing by providing two2. Miller JE. (2013) Molecular Genetic Pathologyimportant and complementary uses:(2nd Edition.,sections 30.2.7.13 and 30.2.7.18).1.Detection of initial clonal populations.3. JE Miller et al., Molecular Genetic Pathology (2nd ed.). Springer Science & Business Media. 2013: 30.2.7.13.2. Identification of sequence information required to track clonalrearrangements in subsequent samples.Analysis of the rearranged TRB locus increases the probability of identifying T cell receptor gene rearrangements, as compared to Simplified Representation of the TRB GeneV D1 J1 C1 D2 J2 C2Depicted are the variable (V), diversity (D), and joining (J) gene regions involved in TRB gene rearrangements, in addition to the downstream consensus (C) gene regions. 36'