Application of Microfluidic Technology to the BIOMED-2 Protocol for Detection of B-Cell Clonality

Zamò, Alberto; Bertolaso, Anna; van Raaij, Annemiek W.M.; Mancini, Francesca; Scardoni, Maria; Montresor, Marina; Menestrina, Fabio; van Krieken, Johan H.J.M.; Chilosi, Marco; Groenen, Patricia J.T.A.; Scarpa, Aldo

doi:10.1016/j.jmoldx.2011.07.007

« Previous Next »The Journal of Molecular Diagnostics
Volume 14, Issue 1 , Pages 30-37, January 2012

Application of Microfluidic Technology to the BIOMED-2 Protocol for Detection of B-Cell Clonality

Alberto Zamò
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
- Address reprint requests to Alberto Zamò, M.D., Ph.D., Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Piazzale L.A. Scuro, 10, 37134 Verona, Italy
Anna Bertolaso
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Annemiek W.M. van Raaij
- Department of Pathology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands
- Address reprint requests to Alberto Zamò, M.D., Ph.D., Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Piazzale L.A. Scuro, 10, 37134 Verona, Italy
Francesca Mancini
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Maria Scardoni
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Marina Montresor
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Fabio Menestrina
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Johan H.J.M. van Krieken
- Department of Pathology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands
Marco Chilosi
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy
Patricia J.T.A. Groenen
- Department of Pathology, Radboud University Nijmegen Medical Center, Nijmegen, The Netherlands
Aldo Scarpa
- Department of Pathology and Diagnostics, Section of Anatomic Pathology, University of Verona, Verona, Italy

Accepted 22 July 2011. published online 24 October 2011.

The BIOMED-2 protocol is widely used for detecting clonality in lymphoproliferative disorders. The protocol requires multiple PCR reactions, which are analyzed by either capillary electrophoresis (GeneScan analysis) or heteroduplex PAGE analysis. We tested a microfluidic chip-based electrophoresis device (Agilent 2100 Bioanalyzer) for the analysis of B-cell clonality using PCR for the three framework subregions (FR) of the Ig heavy chain gene (IGH) and PCR for two rearrangements occurring in the Ig κ chain gene (IGK-VJ and IGK-DE). We analyzed 62 B-cell lymphomas (33 follicular and 29 nonfollicular) and 16 reactive lymph nodes. Chip-based electrophoresis was conclusive for monoclonality in 59/62 samples; for 20 samples, it was compared with GeneScan analysis. Concordant results were obtained in 45/55 IGH (FR1, FR2, and FR3) gene rearrangements, and in 34/37 IGK gene rearrangements. However, when the chip device was used to analyze selected IGK gene rearrangements (biallelic IGK rearrangements or IGK rearrangements in a polyclonal background), its performance was not completely accurate. We conclude, therefore, that this microfluidic chip-based electrophoresis device is reliable for testing cases with dominant PCR products but is less sensitive than GeneScan in detecting clonal peaks in a polyclonal background for IGH PCR, or with complex IGK rearrangement patterns.