Systematic analysis of interference due to stutter in estimating chimerism following hematopoietic cell transplantation

Stutter formed during amplification of short tandem repeats (STRs) interfere with accurate engraftment monitoring following hematopoietic cell transplantation (HCT). We describe a mathematical approach to minimize the contribution of stutter when estimating chimerism following HCT. Pretransplant DNA samples from 409 donors and recipients were used to define marker-specific stutter cut-off values for all makers used in the AMPFℓSTR Profiler Plus amplification kit. Mock chimerism samples (5, 20, and 50%) were used to evaluate the contribution of stutter in estimating chimerism. Three markers, vWA, D13S317, and D18S51, had overlapping stutter in the mock chimerism samples. Only D18S51 had stutter from a shared allele, whereas the other two markers had stutter from nonshared alleles. Without adjustment for stutter, D18S51 showed a 8, 6, and 4% difference from expected chimerism values (5, 20, and 50%) and after adjusting for laboratory-defined marker-specific stutter cut-offs the corresponding difference from expected chimerism values was 1, 0, and 0%. Difference from expected chimerism values in the vWa and D13S317 markers were similar before and after adjustment of stutter. Adjustment for stutter from shared alleles may improve accuracy of estimated chimerism.