Molecular tracking of immunodominant clonotypes in BM biopsies and peripheral blood CD8⁺ cells from AA/MDS patients. (A) Independent sequencing of CDR3 regions derived from BM biopsy DNA and peripheral blood CD8⁺ RNA revealed identical or highly homologous clonotypes. Three examples are shown: in patients no. 37 and no. 39, identical expanded clonotypes were detected in both archived tissue and peripheral blood; patient no. 34 harbors 2 highly homologous minor clonotypes. PBMC indicates peripheral blood mononuclear cell. (B) The presence of clonotypes identified in archival BM tissue was tested on RNA from peripheral blood CD8⁺ cells using a quantitative Taqman assay. The presence and quantity of each shown clonotype was tested in the original patient and 2 healthy control samples obtained from our laboratory. Expression level in controls is shown as fold decrease in comparison to patient's values, which served as calibrator. ND indicates not detected. (C) Detection of blood-derived clonotypes in BM biopsies using nonquantitative clonotypic PCR. We were able to amplify peripheral blood-specific clonotypes in patient BM biopsies. Each clonotype was tested on genomic DNA from patient BM and genomic DNA from healthy controls. The primer set VB forward - JB reverse was used as an endogenous amplification control. The clonotypic primer derived from patient no. 40 also amplifies a nonspecific product of unexpected length in Ctrl1, possibly suggesting a partially rearranged TCR with high homology to patient's TCR sequence.