SEROLOGICAL AND MOLECULAR CHARACTERIZAZATION OF RHD VARIANTS
(Abstract release date: 05/19/16)
EHA Library. Foglieni B. 06/09/16; 135383; LB2272
Dr. Barbara Foglieni
Contributions
Contributions
Abstract
Abstract: LB2272
Type: Eposter Presentation
Background
RhD is the most important, immunogenic and polymorphic Rh antigen, which plays a key role in transfusion medicine. Anti-D antibodies remain the leading cause of the hemolytic disease of the newborn, and antigen D compatible units are chosen when transfusions are needed. Screening tests are based on panels of monoclonal antibodies developed to identify the majority of D variants, but Rh D typing is a constant challenge in donor routine testing, since in presence of RHD blood group polymorphisms of RH partial D or weak D phenotypes may vary according to reagent and method used.In January 2015, the Lombardy transfusion system has been deeply reorganized, and screening tests performed by 27 transfusion centers have been centralized into 8 centers. In our department, we are now testing for ABD about 85,000 units per year.
Aims
The aim of the study was to describe the findings of ABD typing performed with a different system based on agglutination on solid phase technology, different from the ones previously used, based on gel-card technology.
Methods
From March 2015 to March 2016, AB0/Rh typing of blood donations were performed by solid phase technology with a completely automated system (Capture-R Ready-Screen, Immucor). Results were compared with data obtained by gel-card.Samples with negative or weak anti-D reactivity were screened for the presence of RhD variants with different anti-D sera and advanced serological kits such as ID-Partial RhD Typing (Biorad), and, furthermore, they were analyzed for DAT testing. Discrepant or inconclusive samples were selected for further investigation with molecular techniques, based on allele specific PCR for the detection of 14 RhD weak and 48 RhD partial variants (Inno-Train, Essemedical).
Results
A total of 82,000 blood donations, collected from 38,515 donors, were analyzed for AB0 and RhD blood groups. In 130 donors (0.3%) a weak or discrepant RhD typing with different anti-sera were obtained. All of them were analyzed for the presence of RhD variants by molecular approach, which identified: 117 weak D alleles (91 type 1, 7 type 2, 6 type 3, 1 type 4, 1 type 5, 10 type 11, 2 compound heterozygote type 1+4 and type 2+4) and 12 partial D alleles (9 DFR, 1 DNB, 2 DV). In 2 subjects none of the RhD variants analyzed were found.All RhD variants were identified by both gel-card and solid phase technologies, except the weak D type 11 (885G>T) variant carried by 10 subjects, which showed a completely negative results with all monoclonal gel-card antisera used, and only the D weak cells analysis on Immucor microplates revealed a weak reactivity. This RHD variant is commonly classified among the Del phenotype, since it can be only identified by adsorption and elution techniques. The RhD typing of blood component of these 10 blood donors were changed from Rh negative to Rh positive.Moreover, all the RhD variants identified showed a strong correlation between serological reactivity obtained with different anti-sera and molecular results. All RHD variants were identified in subjects with Cc or Ee phenotype, and a linkage disequilibrium between RHD variants and RHCE phenotypes were observed.
Conclusion
Solid phase methods were highly sensitive in detecting very weak RhD expression variants, such as DEL, which is important for the prevention of anti-D post-transfusion or newborn immunizations.Molecular methods help in the differentiation and definition of partial D and weak D types, providing additional information for transfusion procedures.
Session topic: E-poster
Keyword(s): ABO blood group, Molecular, Transfusion
Type: Eposter Presentation
Background
RhD is the most important, immunogenic and polymorphic Rh antigen, which plays a key role in transfusion medicine. Anti-D antibodies remain the leading cause of the hemolytic disease of the newborn, and antigen D compatible units are chosen when transfusions are needed. Screening tests are based on panels of monoclonal antibodies developed to identify the majority of D variants, but Rh D typing is a constant challenge in donor routine testing, since in presence of RHD blood group polymorphisms of RH partial D or weak D phenotypes may vary according to reagent and method used.In January 2015, the Lombardy transfusion system has been deeply reorganized, and screening tests performed by 27 transfusion centers have been centralized into 8 centers. In our department, we are now testing for ABD about 85,000 units per year.
Aims
The aim of the study was to describe the findings of ABD typing performed with a different system based on agglutination on solid phase technology, different from the ones previously used, based on gel-card technology.
Methods
From March 2015 to March 2016, AB0/Rh typing of blood donations were performed by solid phase technology with a completely automated system (Capture-R Ready-Screen, Immucor). Results were compared with data obtained by gel-card.Samples with negative or weak anti-D reactivity were screened for the presence of RhD variants with different anti-D sera and advanced serological kits such as ID-Partial RhD Typing (Biorad), and, furthermore, they were analyzed for DAT testing. Discrepant or inconclusive samples were selected for further investigation with molecular techniques, based on allele specific PCR for the detection of 14 RhD weak and 48 RhD partial variants (Inno-Train, Essemedical).
Results
A total of 82,000 blood donations, collected from 38,515 donors, were analyzed for AB0 and RhD blood groups. In 130 donors (0.3%) a weak or discrepant RhD typing with different anti-sera were obtained. All of them were analyzed for the presence of RhD variants by molecular approach, which identified: 117 weak D alleles (91 type 1, 7 type 2, 6 type 3, 1 type 4, 1 type 5, 10 type 11, 2 compound heterozygote type 1+4 and type 2+4) and 12 partial D alleles (9 DFR, 1 DNB, 2 DV). In 2 subjects none of the RhD variants analyzed were found.All RhD variants were identified by both gel-card and solid phase technologies, except the weak D type 11 (885G>T) variant carried by 10 subjects, which showed a completely negative results with all monoclonal gel-card antisera used, and only the D weak cells analysis on Immucor microplates revealed a weak reactivity. This RHD variant is commonly classified among the Del phenotype, since it can be only identified by adsorption and elution techniques. The RhD typing of blood component of these 10 blood donors were changed from Rh negative to Rh positive.Moreover, all the RhD variants identified showed a strong correlation between serological reactivity obtained with different anti-sera and molecular results. All RHD variants were identified in subjects with Cc or Ee phenotype, and a linkage disequilibrium between RHD variants and RHCE phenotypes were observed.
Conclusion
Solid phase methods were highly sensitive in detecting very weak RhD expression variants, such as DEL, which is important for the prevention of anti-D post-transfusion or newborn immunizations.Molecular methods help in the differentiation and definition of partial D and weak D types, providing additional information for transfusion procedures.
Session topic: E-poster
Keyword(s): ABO blood group, Molecular, Transfusion
Abstract: LB2272
Type: Eposter Presentation
Background
RhD is the most important, immunogenic and polymorphic Rh antigen, which plays a key role in transfusion medicine. Anti-D antibodies remain the leading cause of the hemolytic disease of the newborn, and antigen D compatible units are chosen when transfusions are needed. Screening tests are based on panels of monoclonal antibodies developed to identify the majority of D variants, but Rh D typing is a constant challenge in donor routine testing, since in presence of RHD blood group polymorphisms of RH partial D or weak D phenotypes may vary according to reagent and method used.In January 2015, the Lombardy transfusion system has been deeply reorganized, and screening tests performed by 27 transfusion centers have been centralized into 8 centers. In our department, we are now testing for ABD about 85,000 units per year.
Aims
The aim of the study was to describe the findings of ABD typing performed with a different system based on agglutination on solid phase technology, different from the ones previously used, based on gel-card technology.
Methods
From March 2015 to March 2016, AB0/Rh typing of blood donations were performed by solid phase technology with a completely automated system (Capture-R Ready-Screen, Immucor). Results were compared with data obtained by gel-card.Samples with negative or weak anti-D reactivity were screened for the presence of RhD variants with different anti-D sera and advanced serological kits such as ID-Partial RhD Typing (Biorad), and, furthermore, they were analyzed for DAT testing. Discrepant or inconclusive samples were selected for further investigation with molecular techniques, based on allele specific PCR for the detection of 14 RhD weak and 48 RhD partial variants (Inno-Train, Essemedical).
Results
A total of 82,000 blood donations, collected from 38,515 donors, were analyzed for AB0 and RhD blood groups. In 130 donors (0.3%) a weak or discrepant RhD typing with different anti-sera were obtained. All of them were analyzed for the presence of RhD variants by molecular approach, which identified: 117 weak D alleles (91 type 1, 7 type 2, 6 type 3, 1 type 4, 1 type 5, 10 type 11, 2 compound heterozygote type 1+4 and type 2+4) and 12 partial D alleles (9 DFR, 1 DNB, 2 DV). In 2 subjects none of the RhD variants analyzed were found.All RhD variants were identified by both gel-card and solid phase technologies, except the weak D type 11 (885G>T) variant carried by 10 subjects, which showed a completely negative results with all monoclonal gel-card antisera used, and only the D weak cells analysis on Immucor microplates revealed a weak reactivity. This RHD variant is commonly classified among the Del phenotype, since it can be only identified by adsorption and elution techniques. The RhD typing of blood component of these 10 blood donors were changed from Rh negative to Rh positive.Moreover, all the RhD variants identified showed a strong correlation between serological reactivity obtained with different anti-sera and molecular results. All RHD variants were identified in subjects with Cc or Ee phenotype, and a linkage disequilibrium between RHD variants and RHCE phenotypes were observed.
Conclusion
Solid phase methods were highly sensitive in detecting very weak RhD expression variants, such as DEL, which is important for the prevention of anti-D post-transfusion or newborn immunizations.Molecular methods help in the differentiation and definition of partial D and weak D types, providing additional information for transfusion procedures.
Session topic: E-poster
Keyword(s): ABO blood group, Molecular, Transfusion
Type: Eposter Presentation
Background
RhD is the most important, immunogenic and polymorphic Rh antigen, which plays a key role in transfusion medicine. Anti-D antibodies remain the leading cause of the hemolytic disease of the newborn, and antigen D compatible units are chosen when transfusions are needed. Screening tests are based on panels of monoclonal antibodies developed to identify the majority of D variants, but Rh D typing is a constant challenge in donor routine testing, since in presence of RHD blood group polymorphisms of RH partial D or weak D phenotypes may vary according to reagent and method used.In January 2015, the Lombardy transfusion system has been deeply reorganized, and screening tests performed by 27 transfusion centers have been centralized into 8 centers. In our department, we are now testing for ABD about 85,000 units per year.
Aims
The aim of the study was to describe the findings of ABD typing performed with a different system based on agglutination on solid phase technology, different from the ones previously used, based on gel-card technology.
Methods
From March 2015 to March 2016, AB0/Rh typing of blood donations were performed by solid phase technology with a completely automated system (Capture-R Ready-Screen, Immucor). Results were compared with data obtained by gel-card.Samples with negative or weak anti-D reactivity were screened for the presence of RhD variants with different anti-D sera and advanced serological kits such as ID-Partial RhD Typing (Biorad), and, furthermore, they were analyzed for DAT testing. Discrepant or inconclusive samples were selected for further investigation with molecular techniques, based on allele specific PCR for the detection of 14 RhD weak and 48 RhD partial variants (Inno-Train, Essemedical).
Results
A total of 82,000 blood donations, collected from 38,515 donors, were analyzed for AB0 and RhD blood groups. In 130 donors (0.3%) a weak or discrepant RhD typing with different anti-sera were obtained. All of them were analyzed for the presence of RhD variants by molecular approach, which identified: 117 weak D alleles (91 type 1, 7 type 2, 6 type 3, 1 type 4, 1 type 5, 10 type 11, 2 compound heterozygote type 1+4 and type 2+4) and 12 partial D alleles (9 DFR, 1 DNB, 2 DV). In 2 subjects none of the RhD variants analyzed were found.All RhD variants were identified by both gel-card and solid phase technologies, except the weak D type 11 (885G>T) variant carried by 10 subjects, which showed a completely negative results with all monoclonal gel-card antisera used, and only the D weak cells analysis on Immucor microplates revealed a weak reactivity. This RHD variant is commonly classified among the Del phenotype, since it can be only identified by adsorption and elution techniques. The RhD typing of blood component of these 10 blood donors were changed from Rh negative to Rh positive.Moreover, all the RhD variants identified showed a strong correlation between serological reactivity obtained with different anti-sera and molecular results. All RHD variants were identified in subjects with Cc or Ee phenotype, and a linkage disequilibrium between RHD variants and RHCE phenotypes were observed.
Conclusion
Solid phase methods were highly sensitive in detecting very weak RhD expression variants, such as DEL, which is important for the prevention of anti-D post-transfusion or newborn immunizations.Molecular methods help in the differentiation and definition of partial D and weak D types, providing additional information for transfusion procedures.
Session topic: E-poster
Keyword(s): ABO blood group, Molecular, Transfusion
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