Sequence polymorphism systems for quantitative real-time polymerase chain reaction to characterize hematopoietic chimerism-high informativity and sensitivity as well as excellent reproducibility and precision of measurement.
Sequence polymorphisms (SPs) can serve as genetic markers for quantitative polymerase chain reactions (qPCR) for chimerism analysis, providing a significantly higher sensitivity compared to short tandem repeat PCR. In this study, a panel of 29 selected markers was evaluated in 317 patients with leukemia and myelodysplastic syndrome, who received allogeneic stem cell transplantation. In total, 5415 posttransplantation samples were analyzed. Recipient genotype discrimination was possible in 96% with a mean number of 2.5 (1-7) informative markers per recipient/donor pair. Marker specific standard dilution series from volunteers' DNA served as standard for quantification of chimerism. Sensitivity of the method was < or =1 x 10-3 (0.1% of recipient cells) in 83.3% of the assays. By this method, it was possible to very accurately detect autologous signals in the range from 0% to 0.5% (95% confidence interval [CI] +/-0.2), from 0.5% to 1% (95% CI +/-0.4), from 1% to 2% (95% CI +/-0.6) and from 2% to 5% (95% CI +/-1.2). Reproducibility of the quantified autologous signals was independent from the amount of DNA. This is the first report on a SP-based chimerism system allowing for the performance of chimerism analyses for virtually all patients with high sensitivity, excellent reproducibility, and precision of measurement.
Willasch
Andre
A
J. W. Goethe University Children's Hospital of Frankfurt, Frankfurt, Germany.
Schneider
Gaby
G
Reincke
Britta Sofia
BS
Shayegi
Nona
N
Kreyenberg
Hermann
H
Kuci
Selim
S
Weber
Gerrit
G
Van Der Reijden
Bert
B
Niethammer
Dietrich
D
Klingebiel
Thomas
T
Bader
Peter
P
D000293:Adolescent; D000328:Adult; D002648:Child; D002675:Child, Preschool; D019411:Clinical Laboratory Techniques / Q000379:methods; D005260:Female; D005819:Genetic Markers / Q000235:genetics*; D018380:Hematopoietic Stem Cell Transplantation; D006801:Humans; D007223:Infant; D007231:Infant, Newborn; D008297:Male; D016133:Polymerase Chain Reaction / Q000379:methods*; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D018183:Transplantation Chimera / Q000235:genetics*; D014184:Transplantation, Homologous / Q000502:physiology
D005819:Genetic Markers