Alberto Zamò, Anna Bertolaso, Annemiek W. M

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Application of Microfluidic Technology to the BIOMED-2 Protocol for Detection of B-Cell Clonality Alberto Zamò, Anna Bertolaso, Annemiek W.M. van Raaij, Francesca Mancini, Maria Scardoni, Marina Montresor, Fabio Menestrina, Johan H.J.M. van Krieken, Marco Chilosi, Patricia J.T.A. Groenen, Aldo Scarpa The Journal of Molecular Diagnostics Volume 14, Issue 1, Pages 30-37 (January 2012) DOI: 10.1016/j.jmoldx.2011.07.007 Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 1 Comparison of GeneScan analysis and microfluidic analysis electropherograms. Representative clonality profiles are shown at the left for IGH FR1, FR2, and FR3 and at the right for IGK VJ and DE. Sample numbering corresponds to that of Table 1. The electropherograms of the IGH (FR1, FR2, and FR3) PCR product typically exhibit a curve that is easy to interpret. By contrast, the electropherograms of the IGK (KVJ and KDE) products have distinct patterns (due to the difference in primer design), which make it difficult to interpret the electropherograms and detect clonality. The sizing resolution of the microfluidic chip-based electrophoresis is lower, compared with the GeneScan capillary electrophoresis; however, as seen from sample 12, the Bioanalyzer resolution improves as the PCR amplicon gets smaller. The Bioanalyzer detected both of the biallelic rearrangements for sample 12 (differing by 10 bp) only in IGH FR3 PCR. The Journal of Molecular Diagnostics 2012 14, 30-37DOI: (10.1016/j.jmoldx.2011.07.007) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions

Figure 2 Comparison of IGK rearrangement electropherograms acquired by microfluidic chip-based electrophoresis and by GeneScan analysis for 3 of the 10 additional samples with complex patterns. Case numbering corresponds to that of Table 2. Detection of complex IGK-VJ rearrangements was poorly successful using the microfluidic chip-based electrophoresis readout. This is evident for IGK tube A especially when the sample has a low-intensity signal, which is frequently interpreted as polyclonal. The Journal of Molecular Diagnostics 2012 14, 30-37DOI: (10.1016/j.jmoldx.2011.07.007) Copyright © 2012 American Society for Investigative Pathology and the Association for Molecular Pathology Terms and Conditions