Next-Generation Sequencing Technology to Identify Minimal Residual Disease in Lymphoid Malignancies

Methods Mol Biol. 2021:2185:95-111. doi: 10.1007/978-1-0716-0810-4_7.


Next-generation sequencing (NGS) of immunoglobulin (IG) and T cell receptor (TR) rearrangements represents a modern alternative to classical RQ-PCR-based minimal residual disease (MRD) detection. The same primer sets and conditions can be used for all patients, which is undoubtedly one of the most important benefits of NGS, not only reducing the labor required to perform the analysis but also enabling the assay to comply with the upcoming EU IVD regulation. So far, only one standardized academic protocol for this task has been published, developed, and validated within the EuroClonality-NGS working group. In this chapter we describe the materials and methods for amplicon library preparation for sequencing on Illumina MiSeq, and the bioinformatic pipeline for this protocol.

Keywords: Amplicon sequencing; Bioinformatics; Immunoglobulin; Library preparation; Minimal residual disease; Next-generation sequencing; Rearrangement; T cell receptor.

MeSH terms

  • Gene Rearrangement*
  • Genes, Immunoglobulin*
  • Hematologic Neoplasms* / blood
  • Hematologic Neoplasms* / genetics
  • High-Throughput Nucleotide Sequencing*
  • Humans
  • Lymphoproliferative Disorders* / blood
  • Lymphoproliferative Disorders* / genetics
  • Neoplasm, Residual
  • Receptors, Antigen, T-Cell / genetics*


  • Receptors, Antigen, T-Cell