b'Next-Generation Sequencing (NGS) | Research Use Only (RUO) AssaysIGH Assay Next-Generation Sequencing (NGS) | RUO AssaysLymphoTrack IGH FR1/FR2/FR3 AssaysAssay UsesThese research use only (RUO) assays for next-generation sequencingPositive clonal (SHM negative) and negative polyclonal DNA controls (NGS), identify clonal IGH V H -J Hrearrangements, the associated V H -J H are included in kits. A clonal SHM positive control can be purchased region DNA sequences, the frequency distribution of V Hregion and J H separately (4-088-0008).region segment utilization. The LymphoTrack FR1 can also identify the degree of somatic hypermutation (SHM) of rearranged genes using theBackground Illumina MiSeq, Thermo Fisher Scientific Ion PGM or Ion S5. TheThe human immunoglobulin heavy chain (IGH) gene locus on LymphoTrack IGH FR1, IGH FR2, and IGH FR3 Assays contain primerschromosome 14 (14q32.3) includes 46-52 functional and 30 that target the conserved framework 1 (FR1), framework 2 (FR2), andnonfunctional variable (V H ) gene segments, 27 functional diversity framework 3 (FR3) regions, respectively. The LymphoTrack IGH FR1/2/3(D H ) gene segments, and 6 functional joining (J H ) gene segments Assay kits contain the master mixes of all three frameworks. spread over 1250 kilobases.Summary and Explanation of the TestDuring development of lymphoid cells, the antigen receptor genes go through somatic gene rearrangements. For example, during B-cell The LymphoTrack IGH Assays represent a significant improvementdevelopment, genes encoding the IGH molecules are assembled from over clonality assays that utilize fragment analysis by providing fourmultiple polymorphic gene segments that undergo rearrangements important and complementary uses: and selection, generating V H -D H -J Hcombinations. Since leukemias and lymphomas originate from the malignant transformation of 1.Detection of clonal populations. individual lymphoid cells, all leukemias and lymphomas generally 2.dentification of sequence information and V H -J Hshare one or more cell-specific or clonal antigen receptor gene I rearrangements. Therefore, tests that detect IGH clonal populations segment utilization.can be useful in the study of B- and T-cell malignancies. 3. The LymphoTrack IGH FR1 Assays provide the degree of SHM of the immunoglobulin variable heavy chain (IGHV). Specimen Requirement4. The ability to track sequences in subsequent samples with theInvivoscribe LymphoTrack MRD Software. 50 ng of high-quality genomic DNA.Primers included in the master mixes are designed with Illumina adapters and indices (8 included in Kit A, 24 included in the Panel, andReferencesan independent 24 included in the Panel B) or Thermo Fisher adaptors1.S Tonegawa. Nature 302: 575-581 (1983).and indices (12 indices per framework kits). This allows for a one-step PCR reaction to generate sequence-ready amplicons and pooling of2. JE Miller et al., Molecular Genetic Pathology (2nd ed.). Springerseveral different samples on the same Illumina MiSeq cell, Ion S5 orScience & Business Media. 2013: 30.2.7.13 and 30.2.7.18.Ion PGM chip. The LymphoTrack bioinformatics software enables easy3. KJ Trainor et al., Blood 75: 2220-2222 (1990).analysis and visualization of data and the LymphoTrack MRD Software allows sequences to be tracked in subsequent samples. Please see4. P Ghia. Leukemia 21: 1-3 (2007).the LymphoTrack MRD software section to learn how the LymphoTrack Assays can be applied to MRD studies, or email [email protected] representation of the organization of the immunoglobulin heavy chain (IGH) D H J H gene locus on chromosome 14. Depicted are the variable region (V H ) genes and V L V H -FR1 V H -FR2 V H -FR3J Hprimersdownstream consensus joining region genes segments (J H ) that are involved in rearrangements. V Hfamily primers38'