Capillary Electrophoresis Artifact Due to Eosin

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Presentation transcript:

Capillary Electrophoresis Artifact Due to Eosin Kathleen M. Murphy, Karin D. Berg, Tanya Geiger, Michael Hafez, Katie A. Flickinger, Lisa Cooper, Patrick Pearson, James R. Eshleman The Journal of Molecular Diagnostics Volume 7, Issue 1, Pages 143-148 (February 2005) DOI: 10.1016/S1525-1578(10)60021-9 Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 A–B and D–E: Examples of capillary electrophoresis electropherograms. x axis is size in bases, y axis is fluorescence intensity. For all colors, the color of the fluorescence is depicted by the color of the tracing, except for yellow fluorescence, depicted by a black tracing. IGH PCR amplification products are in blue. Red peaks are internal size standards. A: The original CE results from the IGH gene rearrangement assay of the reported case. Note the peak at 71 bases that is predominantly blue (blue arrow), but also fluoresces to a lesser extent in green (green arrow) and yellow (black arrow). B: CE analysis of the genomic DNA sample without PCR amplification. Both the major peak at 71 bases and the minor peak at 54 bases are consistent with the peaks seen in (A). C: The paraffin-embedded tissue from which DNA was isolated and analyzed. Note the pink color due to the tissue stain eosin. D: Capillary electrophoresis electropherogram of eosin diluted 1:1,000,000 in H2O. The peak at 71 bases fluoresces predominantly blue, but also fluoresces to a lesser extent in green, and even less in yellow. Also note the minor peak at 54 bases fluorescing in blue and green. These findings are consistent with those generated by CE analysis of the original genomic DNA sample without PCR amplification (B). E: CE analysis of the repeat IGH gene rearrangement PCR after additional purification of the DNA sample. The pattern is consistent with polyclonality with no evidence of the 71 base contaminant peak. The Journal of Molecular Diagnostics 2005 7, 143-148DOI: (10.1016/S1525-1578(10)60021-9) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions