Abstract
Although new techniques (i.e., droplet digital-PCR, next-generation sequencing, advanced flow cytometry) are being developed, DNA-based allele-specific real-time quantitative (RQ)-PCR is still the gold standard for sensitive and accurate immunoglobulin/T cell receptor (IG/TR)-based minimal residual disease (MRD) monitoring, allowing the detection of up to 1 leukemic cell in 100,000 normal lymphoid cells. We herewith describe the standard PCR procedure which has been developed and standardized (with minor modification in single labs) through the last 20 years of activity of the EuroMRD Consortium, a volunteer activity of expert laboratories that is continuously providing education, standardization, quality control rounds, and guidelines for interpretation of RQ-PCR data.
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References
Cazzaniga G, Biondi A (2005) Molecular monitoring of childhood acute lymphoblastic leukemia using antigen receptor gene rearrangements and quantitative polymerase chain reaction technology. Haematologica 90:382–390
Van Dongen JJ, Seriu T, Panzer-Grümayer ER, Biondi A, Pongers-Willemse MJ, Corral L et al (1998) Prognostic value of minimal residual disease in acute lymphoblastic leukaemia in childhood. Lancet 352:1731–1738
Conter V, Bartram CR, Valsecchi MG, Schrauder A, Panzer-Grümayer R, Möricke A et al (2010) Molecular response to treatment redefines all prognostic factors in children and adolescents with B-cell precursor acute lymphoblastic leukemia: results in 3184 patients of the AIEOP-BFM ALL 2000 study. Blood 115:3206–3214
Schrappe M, Valsecchi MG, Bartram CR, Schrauder A, Panzer-Grümayer R, Möricke A et al (2011) Late MRD response determines relapse risk overall and in subsets of childhood T-cell ALL: results of the AIEOP-BFM-ALL 2000 study. Blood 118:2077–2084
Brüggemann M, Raff T, Flohr T, Gökbuget N, Nakao M, Droese J et al (2006) Clinical significance of minimal residual disease quantification in adult patients with standard-risk acute lymphoblastic leukemia. German Multicenter Study Group for Adult Acute Lymphoblastic Leukemia. Blood 107:1116–1123
Nunes V, Cazzaniga G, Biondi A (2017) An update on PCR use for minimal residual disease monitoring in acute lymphoblastic leukemia. Expert Rev Mol Diagn 17:953–963
Pongers-Willemse MJ, Verhagen OJ, Tibbe GJ, Wijkhuijs AJ, de Haas V, Roovers E et al (1998) Real-time quantitative PCR for the detection of minimal residual disease in acute lymphoblastic leukemia using junctional region specific TaqMan probes. Leukemia 12:2006–2014
Van der Velden VH, Cazzaniga G, Schrauder A, Hancock J, Bader P, Panzer-Grumayer ER et al (2007) European Study Group on MRD detection in ALL (ESG-MRD-ALL). Analysis of minimal residual disease by Ig/TCR gene rearrangements: guidelines for interpretation of real-time quantitative PCR data. Leukemia 21:604–611
van der Velden VH, Panzer-Grümayer ER, Cazzaniga G, Flohr T, Sutton R, Schrauder A et al (2007) Optimization of PCR-based minimal residual disease diagnostics for childhood acute lymphoblastic leukemia in a multi-center setting. Leukemia 21:706–713
Aubin J, Davi F, Nguyen-Salomon F, Leboeuf D, Debert C, Taher M et al (1995) Description of a novel FR1 IgH PCR strategy and its comparison with three other strategies for the detection of clonality in B cell malignancies. Leukemia 9:471–479
Szczepanski T, Pongers-Willemse MJ, Langerak AW, Harts WA, Wijkhuijs AJM, van Wering ER et al (1999) Ig heavy chain gene rearrangements in T-cell acute lymphoblastic leukemia exhibit predominant DH6-19 and DH7-27 gene usage, can result in complete V-D-J rearrangements, and are rare in T-cell receptor alpha beta lineage. Blood 93:4079–4085
Pongers-Willemse MJ, Seriu T, Stolz F, d’Aniello E, Gameiro P, Pisa P et al (1999) Primers and protocols for standardized detection of minimal residual disease in acute lymphoblastic leukemia using immunoglobulin and T cell receptor gene rearrangements and TAL1 deletions as PCR targets report of the BIOMED-1 CONCERTED ACTION: investigation of minimal residual disease in acute leukemia. Leukemia 13:110–118
van Dongen JJM, Langerak AW, Bruggemann M, Evans PAS, Hummel M, Lavender FL et al (2003) Design and standardization of PCR primers and protocols for detection of clonal immunoglobulin and T-cell receptor gene recombinations in suspect lymphoproliferations: report of the BIOMED-2 Concerted Action BMH4-CT98-3936. Leukemia 17:2257–2317
Szczepanski T, van der Velden VHJ, Hoogeveen PG, de Bie M, Jacobs DCH, Elisabeth R et al (2004) Vd2-Ja rearrangements are frequent in precursor-B–acute lymphoblastic leukemia but rare in normal lymphoid cells. Blood 103:3798–3804
Boone E, Verhaaf B, Langerak AW (2013) PCR-based analysis of rearranged immunoglobulin or T-cell receptor genes by GeneScan analysis or heteroduplex analysis for clonality assessment in lymphoma diagnostics. Methods Mol Biol 971:65–91
van der Velden VHJ, Wijkhuijs JM, Jacobs DCH, van Wering ER, van Dongen JJM (2002) T cell receptor gamma gene rearrangements as targets for detection of minimal residual disease in acute lymphoblastic leukemia by real-time quantitative PCR analysis. Leukemia 16:1372–1380
Bruggemann M, van der Velden VHJ, Raff T, Droese J, Ritgen M, Pott C et al (2004) Rearranged T-cell receptor beta genes represent powerful targets for quantification of minimal residual disease in childhood and adult T-cell acute lymphoblastic leukemia. Leukemia 18:709–719
Verhagen OJHM, Willemse MJ, Breunis WB, Wijkhuijs AJM, Jacobs DCH, Joosten SA et al (2000) Application of germline IGH probes in real-time quantitative PCR for the detection of minimal residual disease in acute lymphoblastic leukemia. Leukemia 14:1426–1435
van der Velden VHJ, de Bie M, van Wering ER, van Dongen JJM (2006) Immunoglobulin light chain gene in precursor-B-acute lymphoblastic leukemia: characteristics and applicability for the detection of minimal residual disease. Haematologica 91:679–682
Brüggemann M, Kotrová M, Knecht H, Bartram J, Boudjogrha M, Bystry V et al (2019) EuroClonality-NGS Working Group. Standardized next-generation sequencing of immunoglobulin and T-cell receptor gene recombinations for MRD marker identification in acute lymphoblastic leukaemia; a EuroClonality-NGS validation study. Leukemia 33:2241–2253
Knecht H, Reigl T, Kotrová M, Appelt F, Stewart P, Bystry V et al (2019) EuroClonality-NGS Working Group. Quality control and quantification in IG/TR next-generation sequencing marker identification: protocols and bioinformatic functionalities by EuroClonality-NGS. Leukemia 33:2254–2265
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Cazzaniga, G., Songia, S., Biondi, A., On Behalf of the EuroMRD Working Group. (2021). PCR Technology to Identify Minimal Residual Disease. In: Cobaleda, C., Sánchez-García, I. (eds) Leukemia Stem Cells. Methods in Molecular Biology, vol 2185. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0810-4_6
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DOI: https://doi.org/10.1007/978-1-0716-0810-4_6
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