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Detection of Residual Donor Erythroid Progenitor Cells after Hematopoietic Stem Cell Transplantation for Patients with Hemoglobinopathies
Published on: September 6, 2017
An internal validation approach and quality control on hematopoietic chimerism testing after allogeneic hematopoietic
Miguel Waterhouse1, Regina Kunzmann, Marta Torres
1Department of Hematology/Oncology, University of Freiburg, Hugstetter Strasse 55, 79106 Freiburg, Germany. miguel.waterhouse@uniklinik-freiburg.de
Background:
Hematopoietic chimerism analysis is important in the follow-up of patients undergoing allogeneic stem cell transplantation. PCR of short tandem repeats is mainly used for monitoring chimerism after transplantation. Validation studies and precision of assay's performance with respect to different mixed chimerism stages is not fully addressed. The aim of the present study was to assess the impact of several microsatellite analytical parameters in the quantification of hematopoietic chimerism after allogeneic hematopoietic stem cell transplantation and to analyze the overall analytical process through the application of internal quality control procedures.
Methods:
Artificial DNA mixtures prepared in known proportions and patients samples were analyzed using three microsatellites, together with amplification of amelogenin gene and fluorescence in situ hybridization (FISH) for X and Y chromosomes. Limit of detection, analytical and clinical sensitivity, stochastic threshold and precision profiling was established. Levey-Jennings charts and Westgard rules were applied for quality control evaluation.
Results:
Analytical and clinical sensitivity of the microsatellite markers was between 0.5% and 1.6%. Amelogenin detection and FISH for X and Y chromosomes showed a similar sensitivity. Severe allelic imbalance resulted in up to 50% difference between the calculated and corrected mixed chimerism. Systematic errors were identified using Levey-Jennings charts and Westgard rules.
Conclusions:
Analysis of hematopoietic chimerism performance is a critical step to better understand potential intrinsic errors that may impact the final hematopoietic chimerism results. Implementing quality control tools, such as Levey-Jennings charts together with Westgard rules can identify systematic and random errors so corrective actions can be performed.
