Exp Hematol 2017 May 29;49:39-47.e5. Epub 2017 Jan 29.
Cyto-Molecular Diagnostic Research Group, Victorian Clinical Genetics Services and Murdoch Childrens Research Institute, Royal Children's Hospital, Parkville, Victoria, Australia; Department of Paediatrics, University of Melbourne, Parkville, Victoria, Australia. Electronic address:
Chimerism analysis has an important role in the management of allogeneic hematopoietic stem cell transplantation. It informs response to disease relapse, graft rejection, and graft-versus-host disease. We have developed a method for chimerism analysis using ubiquitous copy number variation (CNV), which has the benefit of a "negative background" against which multiple independent informative markers are quantified using digital droplet polymerase chain reaction. A panel of up to 38 CNV markers with homozygous deletion frequencies of approximately 0.4-0.6 were used. Sensitivity, precision, reproducibility, and informativity were assessed. CNV chimerism results were compared against established fluorescence in situ hybridization, single nucleotide polymorphism, and short tandem repeat-based methods with excellent correlation. Using 30 ng of input DNA per well, the limit of detection was 0.05% chimerism and the limit of quantification was 0.5% chimerism. High informativity was seen with a median of four informative markers detectable per individual in 39 recipients and 43 donor genomes studied. The strength of this approach was exemplified in a multiple donor case involving four genomes (three related). The precision, sensitivity, and informativity of this approach recommend it for use in clinical practice.