12 Invivoscribe 2019 | 13 Test Algorithm for Suspect Lymphoproliferations Developed in concert with the EuroClonality/BIOMED-2 group for PCR-based clonality assessment of suspected B- and T-cell lymphoproliferative disorders. PCR & NGS-Based Assessment of Clonality in Hematologic Malignancies For more than 20 years, Invivoscribe has developed, manufactured, and commercialized the gold-standard molecular hematopathology assays and reagents for gel and capillary electrophoresis detection, and most recently, next-generation sequencing instruments. These standardized, cGMP manufactured assays and reagents were developed and validated using standardized workflow and optimized primer sets, reagents and controls. A number of our products were developed in collaboration with studies conducted by the EuroClonality BIOMED-2 concerted action and these capillary based products have provided reliable methods for clonality detection that have withstood the test of time. Importantly, we have never accepted the status quo, so our comprehensive menu of clonality assays continues to evolve. All of our NGS-based clonality assays were developed in-house together with accompanying bioinformatics software by our Invivoscribe R&D team. Developed under full ISO 13485 design control, these assays and bioinformatics software were designed to run on several next- generation sequencing platforms. These NGS-based assays are several generations ahead of capillary-based products. Our comprehensive bioinformatics software not only provides critical information on the presence of clonality, but also identifies the sequence information required to track clones in subsequent samples. The unique process of gene rearrangement that occurs within the immunoglobulin (Ig) and T-cell receptor (TCR) gene loci during immune cell development and maturation generates a vast pool of genetically distinct cells. The resulting diverse population of lymphocytes displays an astonishing number of diverse antigen receptors, each coded in the DNA by a unique sequence, and each displayed on the cell surface, or as antibodies in the blood unique to a given cell 1,2 . This diversity allows the adaptive immune system to carry out its role in protecting the human body by recognizing the infinite number of pathogens it might encounter during a lifetime. In sum, lymphoid malignancies are characterized by size- and sequence-specific rearrangements within these loci, which result from the transformation and subsequent proliferation from a single cell. The associated cellular population typically shares one or more cell-specific or “clonal” antigen-receptor gene rearrangements. The detection of these clonal cells forms the basis for clonality assessment in leukemia, lymphoma, and hematologic disease. These methods can also be used to assess somatic hypermutation (SHM) and to study minimal residual disease (MRD). Malignant cells that remain in the bone marrow following treatment are a major cause of disease relapse. MRD testing by NGS offers enhanced sensitivity and specificity (compared to MRD testing by flow cytometry), and allows residual cells to be identified at very low levels and monitored throughout the different stages of disease. Invivoscribe can provide you with the necessary tools to accommodate your needs. From gel detection to NGS, we can help you accurately identify and track hematologic biomarkers. For additional information on the detection methods available and the biomarkers offered, please refer to the respective product sections of this catalog. Immunoglobulin and T-Cell Receptor Gene Rearrangements and the Principle and Method of Clonality Testing The adaptive vertebrate immune system produces a repertoire of immunoglobulin and T-cell receptor molecules using a relatively limited number of heritable germline gene segments. Somatic gene rearrangement is the fundamental mechanism used to generate different immunoglobulin and T-cell receptor molecules, each with unique binding specificity. Lymphocytes undergo gene rearrangements to assemble CDR3 coding regions that are unique in both size and DNA sequence. Since leukemias and lymphomas arise from the malignant transformation of a single cell, they share clonal rearrangement(s) of the antigen receptor genes. This is the basis for clonality testing3 . References: 1.  Tonegawa, S. Somatic Generation of Antibody Diversity. Nature 302:575-581 (1983) 2.  Expression of T-cell receptor genes during early T-cell development. Immunol Cell Biol. 2008 Feb;86(2):166-74. Epub 2007 Oct 23. 3.  Miller, J. E. (2013). Principle of Immunoglobulin and T cell Receptor Gene Rearrangement. In Cheng, L., Zhang, D., Eble, J. N. (Eds), Molecular Genetic Pathology (2nd Ed., Sections 30.2.7.13 and 30.2.7.18). pp825 – 856. New York, USA: Springer Science & Business Media. Background *Previously known as BCL1/JH Results should be considered in the context of all available clinical, histological and immunophenotypic data. If a definitive diagnosis is not possible following: Morphology Immunohistochemistry Flow Cytometry Suspected B-cell proliferations Suspected T-cell proliferations IGH (DH-JH) preferably with IGL Suspected lymphoid proliferations of unknown origin (B or T) Clonal (generally multiple clonal results) Clonal No evidence of clonality TCRƴ δ+ proliferations and immature T-cells IGH (VH-JH) preferably with IGK Mantle Cell Lymphoma IGH- CCND1* Follicular Lymphoma IGH- BCL2 No clonality but still suspected TRD AND TRB preferably with TRG