A comparative study of two routinely used protocols for ex vivo erythroid differentiation.

BACKGROUND: Erythropoiesis is a complex developmental process in which a hematopoietic stem cell undergoes serial divisions and differentiates through well-defined stages to give rise to red blood cells. Over the last decades, several protocols have been developed to perform ex vivo erythroid differentiation, allowing investigation into erythropoiesis and red cell production in health and disease. RESULTS: In the current study, we compared the two commonly used protocols by assessing the differentiation kinetics, synchronisation, and cellular yield, using molecular and cellular approaches. Peripheral blood CD34+ cells were cultured in a two-phase (2P) or a four-phase (4P) liquid culture (LC) and monitored for 20 days. Both protocols could recapitulate all stages of erythropoiesis and generate reticulocytes, although to different extents. Higher proliferation and viability rates were achieved in the 4P-LC, with a higher degree of terminal differentiation and enucleation, associated with higher levels of the erythroid-specific transcription factors GATA-1, KLF-1, and TAL-1. Although the 2P-LC protocol was less efficient regarding terminal erythroid differentiation and maturation, it showed a higher yield of erythroid progenitors in the erythropoietin (EPO)-free expansion phase. CONCLUSIONS: We provide data supporting the use of one protocol or the other to study the biological processes occurring in the early or late stages of erythroid differentiation, depending on the physiological process or pathological defect under investigation in a given study.

Comparison of HLA antibody identification methods for the selection of platelet products for HLA-mediated platelet refractory patients.

In an ineffective transfusion context, solid-phase immunoassays using the Luminex platform for the detection and characterization of HLA antibodies are currently used to select HLA-compatible platelet products. A new HLA antibody identification method, the HISTO SPOT® HLA AB test (BAG Health care GmbH, Lich, Germany), based on the detection of antibodies directed against a recombinant single antigen (SA) by colored spots detected by HISTO MATCH HLA AB module software, runs fully automated on the MR.SPOT®. The aim of this study was to compare the ability of the HISTO SPOT HLA AB and C1qScreen™ (C1q SAB) assays with that of the Labscreen single antigen class I (OL SAB) assay to detect anti-HLA class I antibodies in 56 serum samples from 54 platelet refractory acute myeloid leukemia patients who received HLA mismatch platelet concentrates at a single oncohematology center. In total, 1414 class I specificities, 433 HLA-A and 981 HLA-B, were detected by the OL SAB test. The mean fluorescence intensity (MFI) was >5000 for 874 antigens and <5000 for 655 antigens. The HISTO SPOT® HLA AB and C1q SAB tests identified 85% and 79% of OL SA-detected antigens with an MFI >5000, respectively, but did not identify 34% and 44% of OL SAB-detected antigens, highlighting the lower sensitivity of these techniques. Interestingly, the donor-specific antibodies (DSAs) identified by the HISTO SPOT® HLA AB and C1q SAB assays reacted against HLA mismatch platelet concentrates with the same specificity (86%) and positive predictive (77%) value as in the OL SAB test when the MFI threshold was >2000 for DSA detection. Although the HISTO SPOT® HLA AB test is less sensitive than the OL SAB test, this test could be used for the selection of HLA-compatible platelet products.

New methods for the quantification of mixed chimerism in transplantation.

Background: Quantification of chimerism showing the proportion of the donor in a recipient is essential for the follow-up of hematopoietic stem cell transplantation but can also be useful to document an immune tolerance situation after solid organ transplantation. Historically, chimerism has been quantified from genomic DNA, but with technological advances, chimerism from donor-derived cell-free DNA seems particularly relevant in solid organ transplantation. Methods: The reference method was until recently the short tandem repeat technique, but new innovative techniques as digital PCR (dPCR) and NGS, have revolutionized the quantification of chimerism, such as the so-called microchimerism analysis. After a short review of chimerism methods, a comparison of chimerism quantification data for two new digital PCR systems (QIAcuity™ dPCR (Qiagen®) and QuantStudio Absolute Q (ThermoFisher®) and two NGS-based chimerism quantification methods (AlloSeq HCT™ (CareDx®) and NGStrack™ (GenDX®)) was performed. Results: These new methods were correlated and concordant to routinely methods (r²=0.9978 and r²=0.9974 for dPCR methods, r²=0.9978 and r²=0.9988 for NGS methods), and had similar high performance (sensitivity, reproductibility, linearity). Conclusion: Finally, the choice of the innovative method of chimerism within the laboratory does not depend on the analytical performances because they are similar but mainly on the amount of activity and the access to instruments and computer services.

Therapeutic efficacy of platelet transfusion treated with amotosalen/UVA pathogen inactivation technology (INTERCEPTTM Blood System) in acute myeloid leukemia patients undergoing chemotherapy with curative intent: a single center experience.

BACKGROUND: The INTERCEPTTM Blood System (Intercept Blood System, Cerus Europe BV, Amersfoort, the Netherlands) has been used to reduce or inactivate pathogen load in platelet concentrates in France for three years. MATERIALS AND METHODS: After comparing the transfusion efficiency between pathogen-reduced platelets (PR_PLT) and untreated platelet products (U_PLT), our single-center observational study assessed the effectiveness of PR_PLT for the prevention of bleeding and for therapeutic treatment of WHO grade 2 bleeding in 176 patients undergoing chemotherapy with curative intent for acute myeloid leukemia (AML). The main endpoints were the 24-hour (h) corrected count increment (24h_CCI) after each transfusion, and time to next transfusion. RESULTS: Whereas the transfused doses tended to be higher in the PR_PLT group compared to U_PLT, there was a significant difference in intertransfusion interval (ITI) and 24h_CCI. In prophylactic transfusions, PR_PLT transfusions of >0.65×1011/10 kg, regardless of the age of the product (day 2 to day 5), resulted in a 24h_CCI similar to that of the untreated platelet product; this meant the patient could be transfused at least every 48h. In contrast, most PR_PLT transfusions of <0.55×1011/10 kg did not achieve a transfusion interval of 48h. In the context of WHO grade 2 bleeding, PR_PLT transfusions >0.65×1011/10 kg and storage of less than 4 days seems more effective in stopping bleeding. DISCUSSION: These results, which must be confirmed by prospective studies, indicate the need for vigilance regarding the quantity and quality of PR_PLT products used to treat patients at risk of bleeding crisis. Future prospective studies are needed to confirm these findings.

Mobilisation of HLA-F on the surface of bronchial epithelial cells and platelets in asthmatic patients.

Uncontrolled inflammation of the airways in chronic obstructive lung diseases leads to exacerbation, accelerated lung dysfunction and respiratory insufficiency. Among these diseases, asthma affects 358 million people worldwide. Human bronchial epithelium cells (HBEC) express both anti-inflammatory and activating molecules, and their deregulated expression contribute to immune cell recruitment and activation, especially platelets (PLT) particularly involved in lung tissue inflammation in asthma context. Previous results supported that HLA-G dysregulation in lung tissue is associated with immune cell activation. We investigated here HLA-F expression, reported to be mobilised on immune cell surface upon activation and displaying its highest affinity for the KIR3DS1-activating NK receptor. We explored HLA-F transcriptional expression in HBEC; HLA-F total expression in PBMC and HBEC collected from healthy individuals at rest and upon chemical activation and HLA-F membrane expression in PBMC, HBEC and PLT collected from healthy individuals at rest and upon chemical activation. We compared HLA-F transcriptional expression in HBEC from healthy individuals and asthmatic patients and its surface expression in HBEC and PLT from healthy individuals and asthmatic patients. Our results support that HLA-F is expressed by HBEC and PLT under healthy physiological conditions and is retained in cytoplasm, barely expressed on the surface, as previously reported in immune cells. In both cell types, HLA-F reaches the surface in the inflammatory asthma context whereas no effect is observed at the transcriptional level. Our study suggests that HLA-F surface expression is a ubiquitous post-transcriptional process in activated cells. It may be of therapeutic interest in controlling lung inflammation.

HLA-H*02:07 Is a Membrane-Bound Ligand of Denisovan Origin That Protects against Lysis by Activated Immune Effectors.

The biological relevance of genes initially categorized as "pseudogenes" is slowly emerging, notably in innate immunity. In the HLA region on chromosome 6, HLA-H is one such pseudogene; yet, it is transcribed, and its variation is associated with immune properties. Furthermore, two HLA-H alleles, H*02:07 and H*02:14, putatively encode a complete, membrane-bound HLA protein. Here we thus hypothesized that HLA-H contributes to immune homeostasis similarly to tolerogenic molecules HLA-G, -E, and -F. We tested if HLA-H*02:07 encodes a membrane-bound protein that can inhibit the cytotoxicity of effector cells. We used an HLA-null human erythroblast cell line transduced with HLA-H*02:07 cDNA to demonstrate that HLA-H*02:07 encodes a membrane-bound protein. Additionally, using a cytotoxicity assay, our results support that K562 HLA-H*02:07 inhibits human effector IL-2-activated PBMCs and human IL-2-independent NK92-MI cell line activity. Finally, through in silico genotyping of the Denisovan genome and haplotypic association with Denisovan-derived HLA-A*11, we also show that H*02:07 is of archaic origin. Hence, admixture with archaic humans brought a functional HLA-H allele into modern European and Asian populations.

Evaluation of Next-Generation Sequencing and Crystal Digital PCR for Chimerism Monitoring of Post-Allogeneic Hematopoietic Stem Cell Transplantation.

Hematopoietic stem cell transplantation (HSCT) is a curative treatment for most hematologic diseases. To evaluate the level of donor engraftment, chimerism must be carefully monitored after HSCT. Short tandem repeats, quantitative PCR (qPCR), and, more recently, digital PCR (dPCR) are widely used to determine the proportions of donor and recipient cells after HSCT. The screening and quantification of chimerism have been evaluated by 2 new methods: a ready-to-use next-generation sequencing (NGS)-based method using the Devyser ChimerismNGS kit and an original combination of the Stilla crystal digital PCR (cdPCR) platform with 3-color multiplexing capacity using GenDX KMRtrack reagents. The genotyping of 4 HSCT pairs by cdPCR using 11 triplex mixes of the GenDX KMRtype kit was consistent at 98.8% with qPCR. Informative samples (n = 20) from 6 donor-recipient pairs and 1 external proficiency test demonstrated the reliability of the results (0.1% to 50%) for the 2 methods. The methods are also highly sensitive (0.1%) and accurate. The chimerism values of the 2 methods are correlated and concordant with those of the reference methods. In addition, the ADVYSER software (Devyser) is user-friendly and well adapted to chimerism monitoring. In conclusion, these 2 innovative methods are easy to perform and user-friendly in all molecular, hematology, and immunogenetic laboratories and allow the genotyping and monitoring of chimerism with high performance and sensitivity.

Identification of 11 novel HLA-A, -B, -C, -DRB1, and -DQB1 alleles in the 1000 Genomes Project panel.

We isolated 11 novel HLA alleles in the 1000 Genomes Project using PolyPheMe software.

Analysis of a large single institution cohort of related donors fails to detect a relation between SDF1/CXCR4 or VCAM/VLA4 genetic polymorphisms and the level of hematopoietic progenitor cell mobilization in response to G-CSF.

We studied a cohort of 367 healthy related donors who volunteered to donate their hematopoietic stem cells for allogeneic transplantation. All donors were homogeneously cared for at a single institution, and received rhG-CSF as a mobilization treatment prior to undergoing apheresis. Peripheral blood CD34+ cell counts were used as the main surrogate marker for rhG-CSF induced mobilization. We searched whether inter-individual variations in known genetic polymorphisms located in genes whose products are functionally important for mobilization, could affect the extent of CD34+ mobilization, either individually or in combination. We found little or no influence of individual SNPs or haplotypes for the SDF1, CXCR4, VCAM and VLA4 genes, whether using CD34+ cell counts as a continuous or a categorical variable. Simple clinical characteristics describing donors such as body mass index, age and possibly sex are more potent predictors of stem cell mobilization. The size of our cohort remains relatively small for genetic analyses, however compares favorably with cohorts analyzed in previously published reports suggesting associations of genetic traits to response to rhG-CSF; notwithstanding this limitation, our data do not support the use of genetic analyses when the choice exists of several potential donors for a given patient.

A transcriptional signature associated with non-Hodgkin lymphoma in the blood of patients with Q fever.

Coxiella burnetii, the agent causing Q fever, has been associated with B-cell non-Hodgkin lymphoma (NHL). To better clarify this link, we analysed the genetic transcriptomic profile of peripheral blood leukocytes from patients with C. burnetii infection to identify possible links to lymphoma. Microarray analyses revealed that 1189 genes were expressed differently (p <.001 and fold change ≥4) in whole blood of patients with C. burnetii infection compared to controls. In addition, 95 genes expressed in patients with non-Hodgkin lymphoma (NHL) and in patients with C. burnetii persistent infection have allowed us to establish the 'C. burnetii-associated NHL signature'. Among these, 33 genes previously found modulated in C. burnetii-associated -NHL by the microarray analysis were selected and their mRNA expression levels were measured in distinct C. burnetii-induced pathologies, namely, acute Q fever, focalized persistent infection, lymphadenitis and C.burnetii-associated NHL. Specific genes involved in anti-apoptotic process were found highly expressed in leukocytes from patients with C. burnetii associated-NHL: MIR17HG, REL and SP100. This signature differed from that found for NHL-control group. Patients with C. burnetii lymphadenitis presented significant elevated levels of BCL2 and ETS1 mRNAs. Altogether, we identified a specific transcriptionnal signature for NHL during C. burnetii infection reflecting the up-regulation of anti-apoptotic processes and the fact that lymphadenitis might constitute a critical step towards lymphomagenesis.

FCGR3A and FCGR2A Genotypes Differentially Impact Allograft Rejection and Patients' Survival After Lung Transplant.

Fc gamma receptors (FcγRs) play a major role in the regulation of humoral immune responses. Single-nucleotide polymorphisms (SNPs) of FCGR2A and FCGR3A can impact the expression level, IgG affinity and function of the CD32 and CD16 FcγRs in response to their engagement by the Fc fragment of IgG. The CD16 isoform encoded by FCGR3A [158V/V] controls the intensity of antibody-dependent cytotoxic alloimmune responses of natural killer cells (NK) and has been identified as a susceptibility marker predisposing patients to cardiac allograft vasculopathy after heart transplant. This study aimed to investigate whether FCGR2A and FCGR3A polymorphisms can also be associated with the clinical outcome of lung transplant recipients (LTRs). The SNPs of FCGR2A ([131R/H], rs1801274) and FCGR3A ([158V/F], rs396991) were identified in 158 LTRs and 184 Controls (CTL). The corresponding distribution of genotypic and allelic combinations was analyzed for potential links with the development of circulating donor-specific anti-HLA alloantibodies (DSA) detected at months 1 and 3 after lung transplant (LTx), the occurrence of acute rejection (AR) and chronic lung allograft dysfunction (CLAD), and the overall survival of LTRs. The FCGR3A [158V/V] genotype was identified as an independent susceptibility factor associated with higher rates of AR during the first trimester after LTx (HR 4.8, p < 0.0001, 95% CI 2.37-9.61), but it could not be associated with the level of CD16- mediated NK cell activation in response to the LTR's DSA, whatever the MFI intensity and C1q binding profiles of the DSA evaluated. The FCGR2A [131R/R] genotype was associated with lower CLAD-free survival of LTRs, independently of the presence of DSA at 3 months (HR 1.8, p = 0.024, 95% CI 1.08-3.03). Our data indicate that FCGR SNPs differentially affect the clinical outcome of LTRs and may be of use to stratify patients at higher risk of experiencing graft rejection. Furthermore, these data suggest that in the LTx setting, specific mechanisms of humoral alloreactivity, which cannot be solely explained by the complement and CD16-mediated pathogenic effects of DSA, may be involved in the development of acute and chronic lung allograft rejection.

HLAIb worldwide genetic diversity: New HLA-H alleles and haplotype structure description.

The classical HLA class I genes (HLA Ia) were extensively studied because of their implication in clinical fields and anthropology. Less is known about worldwide genetic diversity and linkage disequilibrium for non-classical HLA class I genes (HLA Ib) and HLA pseudogenes. Notably, HLA-H, which is deleted in a fraction of the population, remains scarcely explored. The aims of this study were 1/ to get further insight into HLA-H genetic diversity and into how this variability potentially affects its expression and 2/ to define HLA Ib worldwide allelic diversity and linkage. Exome sequence data from the 1000 Genomes Project were used to define second field HLA-A, -E, -F, -G and -H typing using PolyPheMe software. Allelic and two-loci haplotype frequencies were estimated using Gene[Rate] software both at worldwide and continental levels. Eleven novel HLA-H alleles identified in exome data were validated by NGS performed on 25 genomic DNA samples from the same cohort. Phylogenetic analysis and frequency distribution of HLA-H alleles revealed three clades, each predominantly represented in Admixed American, European and East Asian populations, African populations and South Asian populations. Among these eleven novel alleles, two potentially encode complete transmembrane HLA proteins. We confirm the high LD between HLA-H and -A, and between HLA-H and -G, and show the three genes have distinct worldwide allelic distribution. Conversely, HLA-E and HLA-F both showed little LD, displayed restricted allelic diversity and practically no difference in their distribution across the planet. Our work thus reveals an unexpectedly high HLA-H genetic diversity, with alleles highly represented in Asia possibly encoding a functional HLA protein. Functional implication of these results remains to be explored, both in physiological and pathological contexts.

Defining KIR and HLA Class I Genotypes at Highest Resolution via High-Throughput Sequencing.

The physiological functions of natural killer (NK) cells in human immunity and reproduction depend upon diverse interactions between killer cell immunoglobulin-like receptors (KIRs) and their HLA class I ligands: HLA-A, HLA-B, and HLA-C. The genomic regions containing the KIR and HLA class I genes are unlinked, structurally complex, and highly polymorphic. They are also strongly associated with a wide spectrum of diseases, including infections, autoimmune disorders, cancers, and pregnancy disorders, as well as the efficacy of transplantation and other immunotherapies. To facilitate study of these extraordinary genes, we developed a method that captures, sequences, and analyzes the 13 KIR genes and HLA-A, HLA-B, and HLA-C from genomic DNA. We also devised a bioinformatics pipeline that attributes sequencing reads to specific KIR genes, determines copy number by read depth, and calls high-resolution genotypes for each KIR gene. We validated this method by using DNA from well-characterized cell lines, comparing it to established methods of HLA and KIR genotyping, and determining KIR genotypes from 1000 Genomes sequence data. This identified 116 previously uncharacterized KIR alleles, which were all demonstrated to be authentic by sequencing from source DNA via standard methods. Analysis of just two KIR genes showed that 22% of the 1000 Genomes individuals have a previously uncharacterized allele or a structural variant. The method we describe is suited to the large-scale analyses that are needed for characterizing human populations and defining the precise HLA and KIR factors associated with disease. The methods are applicable to other highly polymorphic genes.

HLA-E(⁎)01:03 Allele in Lung Transplant Recipients Correlates with Higher Chronic Lung Allograft Dysfunction Occurrence.

Lung transplantation (LTx) is a valid therapeutic option for selected patients with end-stage lung disease. HLA-E seems to play a major role in the immune response to different viral infections and to affect transplantation outcome, in Hematopoietic Stem Cell Transplantation, for example. Two nonsynonymous alleles, HLA-E(⁎)01:01 and HLA-E(⁎)01:03, have functional differences, involving relative peptide affinity, cell surface expression, and potential lytic activity of NK cells. The aim of this retrospective study was to determine the impact of these two alleles for LTx recipients on anti-HLA alloimmunization risk, overall survival, and chronic rejection (CLAD). HLA-E was genotyped in 119 recipients who underwent LTx from 1998 to 2010 in a single transplantation center. In univariate analysis, both HLA-E homozygous states were associated with impaired overall survival compared to heterozygous HLA-E alleles (p = 0.01). In multivariate analysis, HLA-E(⁎)01:03 allele showed increased CLAD occurrence when compared to homozygous HLA-E(⁎)01:01 status (HR: 3.563 (CI 95%, 1.016-12), p = 0.047). HLA-E allele did not affect pathogen infection or the production of de novo DSA. This retrospective study shows an uninvestigated, deleterious association of HLA-E alleles with LTx and requires verification using a larger cohort.

A comprehensive survey of both RHD and RHCE allele frequencies in sub-Saharan Africa.

BACKGROUND: The RH system is one of the most polymorphic blood group systems with numerous allele variants affecting Rh polypeptides expression. This complexity is at the origin of difficulties for transfusion of African patients especially sickle cell disease patients requiring chronic transfusion therapy with high risk of immunization. As a complete survey of RH variants is lacking in African populations, we performed red blood cell genotyping to determine the type and frequency of RHD and RHCE alleles in sub-Saharan African populations. STUDY DESIGN AND METHODS: A total of 347 blood samples were collected from individuals of six nonpygmoid and three pygmoid populations. RH typing was performed using two single-tube multiplex polymerase chain reaction amplifications (BioArray Solutions, Immucor). RESULTS: All six sub-Saharan nonpygmoid populations exhibited constant variety in both type and frequency of aberrant RHD and RHCE alleles. Predicted partial RH1 (1.8%) and RH5 (0.9%) phenotypes were less than expected. Conversely, predicted partial phenotype RH2 (5.5%) was frequent. Data confirmed the high frequency of samples positive for the non-clinically significant RH10/RH20 antigens (39.5%) and revealed a high frequency of RH54 (DAK, 8.1%). The pygmoid groups showed higher percentages of predicted partial RH antigens and greater heterogeneity reflecting wide genetic differentiation. CONCLUSION: Our data show that frequencies of aberrant RHD and RHCE alleles were similar, irrespective of location and ethnicity. In view of the predicted frequencies and relative clinical significance of both private antigens and high-prevalence antigens absent, the most relevant assays for individuals of African descent in a transfusion setting are for 1) partial RH2 in the patient and 2) RH54 (DAK) in the donor.

RHCE*cE734C allele encodes an altered c antigen and a suppressed E antigen not detected with standard reagents.

BACKGROUND: The RH blood group system has many RHCE variant alleles that have arisen through gene conversion or nucleotide changes. Two probands, with red blood cells (RBCs) that were D+C+E-c+(w) e+ were sent to our laboratories to resolve the weak c expression. STUDY DESIGN AND METHODS: Hemagglutination tests were performed by automated and manual procedures. Genomic DNA analysis was performed by sequencing of Exons 1 to 10 of RHCE and RHD. RESULTS: The probands' RBCs did not react with standard monoclonal anti-E reagents from Bio-Rad, Diagast, DiaMed, Immucor, Ortho, and Quotient. The RBCs reacted variably with anti-c reagents from Diagast, DiaMed, Immucor, or Ortho and did not react with the Quotient anti-c reagent. Surprisingly, sequencing results of RHCE showed the presence of C/G at Position 676 (E/e polymorphism) and the association of the E polymorphism with a 734T>C transition in Exon 5 of the RHCE, encoding a Leu245Pro amino acid substitution in the mature RhcE polypeptide. Replacement of leucine 245 by proline in the eighth transmembrane domain of the RhcE protein may have a steric effect on the protein such that most anti-E reagents do not bind and the interaction between anti-c and c antigen is also affected. CONCLUSION: We report a novel RHCE*cE allele, RHCE*cE734C, which was assigned the provisional ISBT allele name RHCE*cE.14 or RHCE*03.14. It was found in two probands whose RBCs had weakened c expression and typed E- with conventional anti-E reagents. These data, once again, highlight the fact that the genotype does not always reflect the phenotype.

A genetic strategy to control expression of human blood group antigens in red blood cells generated in vitro.

BACKGROUND: The ability to generate red blood cells of a chosen blood group phenotype would be a major advance in transfusion when considering low- and high-frequency blood group antigens. STUDY DESIGN AND METHODS: Cord blood CD34+ cells undergoing erythroid differentiation in vitro were genetically manipulated with human immunodeficiency virus Type 1-derived lentiviral vectors expressing hUT-B1 cDNA (overexpression strategy) or bicistronic vectors expressing both enhanced green fluorescent protein and a short-hairpin RNA (shRNA) designed to silence SLC14A1(JK) gene that encodes hUT-B1 protein (silencing strategy). Resulting cell populations were analyzed by fluorescent-activated cell sorting and gel affinity column assay. RESULTS: When transduced with hUT-B1 cDNA lentiviral vectors encoding JK*B and JK*A alleles, respectively, CD34+ cell-derived erythroid populations from Jk(a+b-) and Jk(a-b+) donors exhibited a Jk(a+b+) phenotype different from the original phenotype. In concomitant tests, Jk(a+b+) donor cells transduced with lentiviral vectors carrying a shRNA designed to interfere with hUT-B1 transcription showed a marked decrease in hUT-B1 expression and were assessed as null for Jk antigen by a routine assay. CONCLUSION: In this work focusing on the Kidd blood group system that relies on expression of hUT-B1 glycoprotein under the Jk(a) or Jk(b) antigenic configurations, we demonstrated that hematopoietic progenitors could be genetically modified to exhibit a chosen Kidd phenotype. Beyond production of atypical Kidd phenotypes, this genetic strategy could allow generation of rare blood phenotypes from hematopoietic stem cells regardless of initial donor phenotype. Potential applications for genetically modified blood include production of control samples for immunohematologic testing and for resolution of antibody detection in multiply transfused patients.