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Quantitative analysis of chimerism after allogeneic stem cell transplantation by PCR amplification of microsatellite markers and capillary electrophoresis with fluorescence detection: the Tuebingen experience

Leukemia volume 17, pages 237–240 (2003)Cite this article

In recent years, considerable progress has been made in the analysis of hematopoietic chimerism after allogeneic stem cell transplantation (SCT). The introduction of serial and quantitative PCR-based methods for the characterization of ‘variable number of tandem repeats’ (VNTR) and ‘short tandem repeats’ (STR) offered the potential for close monitoring of engraftment and for studying the kinetics of graft failure and graft rejection.1,2 Moreover, these methods also offered the possibility to identify patients at high risk of developing relapse after allo-SCT.3,4,5,6,7 In the meantime, analysis of chimerism has become a routine diagnostic instrument at many centers performing allo-SCT. Nevertheless standardized techniques and unified terminology in the field are still missing. This, however, is a prerequisite for quality control and may assist in optimizing techniques to achieve best diagnostic support for clinical decision-making.8 As a contribution to the debate started in Leukemia6,8,9,10,11 we describe here the PCR approach currently used in our laboratory.

The method is based on the discrimination of donor and recipient alleles using STR (short tandem repeat) PCR with markers established for forensic purposes (genetic fingerprinting). STRs express a high degree of size polymorphism, thus giving rise to PCR products of various lengths. In total, a set of nine markers (see Table 1) has been established. To obtain an informative marker, initial screening is performed with patient and donor samples pre-transplant using three of the listed primers. The separation of the PCR products and fluorescence detection are performed on a capillary electrophoresis instrument (ABI 310 Genetic Analyzer; Applied Biosystems, Weiterstadt, Germany) and the final calculation of recipient chimerism is based on the ratio of informative donor and recipient signals.

Table 1 Primer characteristics
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Acknowledgements

This work was supported by the ‘Deutsche Krebshilfe’.

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  1. University Children's Hospital, Tuebingen, Germany

    H Kreyenberg, W Hölle, S Möhrle, D Niethammer & P Bader

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Kreyenberg, H., Hölle, W., Möhrle, S. et al. Quantitative analysis of chimerism after allogeneic stem cell transplantation by PCR amplification of microsatellite markers and capillary electrophoresis with fluorescence detection: the Tuebingen experience. Leukemia 17, 237–240 (2003). https://doi.org/10.1038/sj.leu.2402761

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