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Int. J. Mol. Sci. 2016, 17(9), 1515; doi:10.3390/ijms17091515

Digital PCR Panel for Sensitive Hematopoietic Chimerism Quantification after Allogeneic Stem Cell Transplantation

1
Department of Stem Cell Transplantation, University Medical Center Hamburg-Eppendorf, 20246 Hamburg, Germany
2
Biotype Diagnostic GmbH, 01109 Dresden, Germany
*
Author to whom correspondence should be addressed.
Academic Editor: Maurizio Muraca
Received: 13 July 2016 / Revised: 10 August 2016 / Accepted: 31 August 2016 / Published: 9 September 2016
(This article belongs to the Special Issue Advances in Cell Transplantation)
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Abstract

Accurate and sensitive determination of hematopoietic chimerism is a crucial diagnostic measure after allogeneic stem cell transplantation to monitor engraftment and potentially residual disease. Short tandem repeat (STR) amplification, the current “gold standard” for chimerism assessment facilitates reliable accuracy, but is hampered by its limited sensitivity (≥1%). Digital PCR (dPCR) has been shown to combine exact quantification and high reproducibility over a very wide measurement range with excellent sensitivity (routinely ≤0.1%) and thus represents a promising alternative to STR analysis. We here aimed at developing a whole panel of digital-PCR based assays for routine diagnostic. To this end, we tested suitability of 52 deletion/insertion polymorphisms (DIPs) for duplex analysis in combination with either a reference gene or a Y-chromosome specific PCR. Twenty-nine DIPs with high power of discrimination and good performance were identified, optimized and technically validated. We tested the newly established assays on retrospective patient samples that were in parallel also measured by STR amplification and found excellent correlation. Finally, a screening plate for initial genotyping with DIP-specific duplex dPCR assays was designed for convenient assay selection. In conclusion, we have established a comprehensive dPCR system for precise and high-sensitivity measurement of hematopoietic chimerism, which should be highly useful for clinical routine diagnostics. View Full-Text
Keywords: chimerism; minimal residual disease; digital PCR; allogeneic stem cell transplantation chimerism; minimal residual disease; digital PCR; allogeneic stem cell transplantation
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

Stahl, T.; Rothe, C.; Böhme, M.U.; Kohl, A.; Kröger, N.; Fehse, B. Digital PCR Panel for Sensitive Hematopoietic Chimerism Quantification after Allogeneic Stem Cell Transplantation. Int. J. Mol. Sci. 2016, 17, 1515.

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