Bone Marrow-Free Sequencing of M Protein Genes in Monoclonal Gammopathies

Nevone, Alice; Mazzini, Giulia; Milani, Paolo; Mangiacavalli, Silvia; Melati, Anna; Scanavino, Giulia; Girelli, Maria Elisa; Caminito, Serena; Basset, Marco; Cartia, Claudio Salvatore; Benvenuti, Pietro; Nanci, Martina; Bellofiore, Claudia; Foli, Andrea; Merlini, Giampaolo; Arcaini, Luca; Palladini, Giovanni

doi:10.1182/blood-2022-168615

Blood

2022

DOI: 10.1182/blood-2022-168615

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Bone Marrow-Free Sequencing of M Protein Genes in Monoclonal Gammopathies

Alice Nevone

Giulia Mazzini

Paolo Milani

et al.

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2024

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SMaRT M-Seq: an optimized step-by-step protocol for M protein sequencing in monoclonal gammopathies

Nevone,

Lattarulo,

Russo

et al. 2024

Biology Methods and Protocols

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In patients with monoclonal gammopathies, tumor B cells or plasma cells secrete a monoclonal antibody (M protein) that not only serves as a biomarker for tumor tracking but can also cause potentially life-threatening organ damage. This damage is driven by the patient-specific sequence of the M protein. Methods for sequencing M proteins have been limited by their high cost and time-consuming nature, and while recent approaches using next-generation sequencing or mass spectrometry offer advancements, they may require tumor cell sorting, are limited to subsets of immunoglobulin genes or patients, and/or lack extensive validation. To address these limitations, we have recently developed a novel method, termed Single Molecule Real-Time Sequencing of the M protein (SMaRT M-Seq), which combines the unbiased amplification of expressed immunoglobulin genes via inverse PCR from circularized cDNA with long-read DNA sequencing, followed by bioinformatic and immunogenetic analyses. This approach enables the unambiguous identification of full-length variable sequences of M protein genes across diverse patient cohorts, including those with low tumor burden. Our protocol has undergone technical validation and has been successfully applied to large patient cohorts with monoclonal gammopathies. Here we present the step-by-step protocol for SMaRT M-Seq. By enabling the parallel sequencing of M proteins from a large number of samples in a cost-effective and time-efficient manner, SMaRT M-Seq facilitates access to patient-specific M protein sequences, advancing personalized medicine approaches and enabling deeper mechanistic studies in monoclonal gammopathies.

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SMaRT M-Seq: an optimized step-by-step protocol for M protein sequencing in monoclonal gammopathies

Nevone,

Lattarulo,

Russo

et al. 2024

Biology Methods and Protocols

View full text Add to dashboard Cite

show abstract

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Bone Marrow-Free Sequencing of M Protein Genes in Monoclonal Gammopathies

Cited by 1 publication

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SMaRT M-Seq: an optimized step-by-step protocol for M protein sequencing in monoclonal gammopathies

SMaRT M-Seq: an optimized step-by-step protocol for M protein sequencing in monoclonal gammopathies

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