Error bars depict mean+SD with n = 4 (HD1-4) or n = 11 (pat1-11). marrow samples were assigned to the different subclassesIgA1 and IgA2 as well as IgG1-4. Total numbers of sequences assigned to the different subclasses are outlined in the table. (c) Heatmaps illustrate CDR3 overlap between Ig-subclasses of HD4. Related clones of each subclass repertoire were clustered (4000 sequences if available, otherwise sequence figures according to the above table; 95% CDR3 sequence identity; same VJ-usage), and sorted according Carotegrast to the 100 most abundant clonotypes present. MHI-values of pairwise comparisons are outlined in the table.(TIF) pone.0168096.s002.tif (593K) GUID:?D4946FDF-1EDE-40D5-9D2F-32BD1F775E97 S3 Fig: IgG repertoire dynamics of AML-patients treated by allogeneic HSCT. Matches Fig 2 with data of the remaining 9 patients outlined in Table 1. Evaluation is based on 4000 clustered sequences each (only 2000 sequences for patient 9). Heatmaps illustrate shared clonotypes for the 100 most frequent clonotypes before and after HSCT. Pre- and post-HSCT repertoire similarity is definitely quantified as Morisita-Horn index (MHI).(TIF) pone.0168096.s003.tif (408K) GUID:?22F8A630-0483-4C4B-B8DD-C890525CF579 S4 Fig: IgA repertoire dynamics of AML-patients treated by allogeneic HSCT. IgA sequences of eleven individuals pre- and post-HSCT (patient 1 and patient 9: 2000 sequences; Carotegrast additional individuals: 4000 sequences) were assigned to clonotypes (observe Fig 1 for details). Heatmaps display CDR3-overlap of the 100 most abundant IgA clonotypes. Repertoire similarity is definitely described as Morisita-Horn index (MHI).(TIF) pone.0168096.s004.tif (429K) GUID:?2A32B80A-06F1-44BB-AAC2-DDAA9D75E943 S5 Fig: Exponent Shannon as function of quantity of input sequences. (a, b) For each patient clonotypes were sorted by rate of recurrence and their large quantity displayed IgG and IgA repertoires pre- (blue collection, filled area) and post-HSCT (reddish line, non-filled area). Numbers show the amount of unique clones (x-axis) and quantity sequences assigned to each individual clonotype (y-axis). Graphs display the clonal composition of IgG (a) and IgA repertoires (b). The pub graph depicts the percentage of unique CDR3 sequences pre- and post-transplantation in individual patients; percentage above 1 shows a reduced and a percentage below 1 an increased sample richness post-HSCT (c) Exponent Shannon, expressing sample diversity, was determined for varying numbers of clustered input IgG sequences (95% CDR3 sequence identity; same VJ-usage) derived from healthy donor 3 (HD3) and patient 1 before transplantation (1pre HSCT).(TIF) pone.0168096.s005.tif (585K) GUID:?9CDDDDDD-674D-401F-843F-25BAB3A6A961 S6 Fig: Overall mutation frequency is not impaired after transplantation. (a) IgA and IgG repertoires of four healthy donor (HD1-4) and eleven patient samples pre- as well as post-HSCT (pat 1C11) were analyzed for frequencies of silent and alternative mutations within framework work areas 2 and 3 (FR 2, 3) and complementarity determining areas 1 and 2 (CDR1, 2). Error bars depict mean+SD with n = 4 (HD1-4) or n = 11 (pat1-11). (b) Average quantity of mutations for each patient, listed separately for IgA and IgG repertoires pre- and post-HSCT. Matched samples are connected by lines, all sequences were taken into account. (c) Graphs depict the frequencies of IgG and IgA sequences comprising the indicated quantity of somatic hypermutations (d) For patient 9 Ig repertories pre- and post- transplantation were compared to the respective donor-repertoire; evaluation is based on 1900 clustered sequences. Heatmaps display overlap of the 100 most frequent clonotypes, compared to their frequencies in the additional repertoires. Morisita-Horn indices (MHI) were determined to quantify overall repertoire similarity.(TIF) pone.0168096.s006.tif (616K) GUID:?8DB406C4-BE84-4976-AD98-4F988A326D25 S1 Table: Additional clinical information of AML-patients investigated for repertoire analysis before and after allogeneic HSCT. (DOCX) pone.0168096.s007.docx (18K) GUID:?25FB0F7B-CDA2-44F1-9E19-6687F63B38F5 Data Availability StatementAll relevant data are within the paper and its Supporting Info files. Abstract After allogeneic hematopoietic stem cell transplantation (HSCT), recovery of humoral immunity is essential to protect from life-threatening infections. However, monitoring Carotegrast the humoral immune system after transplantation with standard techniques in the medical routine is definitely imprecise. Here, we performed sequencing of mononuclear bone marrow cells to characterize the VH1-repertoire of switched B cells of healthy volunteers and individuals undergoing HSCT. Analysis of healthy bone marrow donors and individuals showed virtually no clonally related sequences between individuals. Interestingly, clonally related sequences were present in pre- and post-transplantation bone marrow of individuals undergoing HSCT for acute myeloid leukemia treatment. We consistently observed such related B cell clones, irrespective of conditioning regimen, donor resource or time post transplantation. In general, repertoire PI4KB diversity was reduced post-HSCT as compared to pre-HSCT samples. However, post-HSCT repertoires retained highly mutated sequences, despite immunosuppressive therapy and presence of T cell deficiency after HSCT. These.