Molecular background of RH in Bastiaan, the RH:-31,-34 index case, and two novel RHD alleles.

Anti-hr(B) (-RH31) and anti-Hr(B) (-RH34) were found nearly 40 years ago in the serum of a South African woman. The anti-hr(B) was revealed after adsorption with DcE/DcE red blod cells (RCBs). Numerous anti-hr(B), in the absence of anti-Hr(B), have since been identified. We obtained a sample of blood from this index case (Bastiaan) and report the molecular basis of her D+C-E-c+e+/-, V-VS+Hr+hr(S)+hr(B)-Hr(B)- phenotype as well as results of testing her RBCs using currently available regents. We tested a cohort of African Americans to estimate the frequency of the RHCE*ce 48C,733G,1006T allele, and in addition found two novel RHD alleles. Hemagglutination tests and DNA analyses were performed by standard methods. Analyses revealed homozygosity for RHCE*ce 48C,733G,1006T in Bastiaan. RBCs from Bastiaan were strongly agglutinated by three commercial anti-e reagents. Testing RBCs from people homozygous for RHCE*ce 48C,733G,1006T showed that anti-e MS16, MS17, and MS63 were weakly reactive or non reactive, MS21 was strongly reactive, and HIRO38, HIRO41, and HIRO43 were non reactive. Results show that Bastiaan has RHD*DIIIa150C and RHD*DIIIa-CE(4-7)-D. Tests on 605 samples from random African Americans revealed a frequency of 0.036 for RHCE*ce 48C,733G,1006T and revealed two novel alleles: RHD*186T and RHD*DIIIa150C. The Bastiaan phenotype is encoded by RHD*DIIIa150C-RHCE*ce 48C,733G,1006T and RHD*DIIIa-CE(4-7)-D-RHCE*ce 48C,733G,1006T ; thus, this genotype is the gold standard for the hr(B)-Hr(B)-phenotype. The r'(s) complex encodes VS, which explains why most hr(B)-RCBs are VS+.

An alloantibody to a high-prevalence MNS antigen in a person with a GP.JL/Mk phenotype.

The low-prevalence MNS blood group antigenTSEN is located at the junction of glycophorin A (GPA) to glycophorin B (GPB) in several hybrid glycophorin molecules. Extremely rare people have RBCs with a double dose of the TSEN antigen and have made an antibody to a high-prevalence MNS antigen. We report the first patient who is heterozygous for GYP.JL and Mk. During prenatal tests,an alloantibody to a high-prevalence antigen was detected in the serum of a 21-year-old Hispanic woman. The antibody detected an antigen resistant to treatment by papain, trypsin, alpha-chymotrypsin, or DTT. The antibody was strongly reactive by the IAT with all RBCs tested except those having the MkMk, GP.Hil/GP.Hil, or GP.JL/GP.JL phenotypes. The patient's RBCs typed M+N-S+/-s-U+, En(a+/-), Hut-, Mi(a-), Mur-, Vw-, Wr(a-b-), and were TSEN+, MINY+. Reactivity with Glycine soja suggested that her RBCs had a decreased level of sialic acid. Immunoblotting showed the presence of monomer and dimer forms of a GP(A-B) hybrid and an absence of GPA and GPB. Sequencing of DNA and PCR-RFLP using the restriction enzyme RsaI confirmed the presence of a hybrid GYP(AB). The patient's antibody was determined to be anti-EnaFR. She is the first person reported with the GP.JL phenotype associated with a deletion of GYPA and GYPB in trans to GYP.JL.

Loss of enzyme-sensitive antigens due to the presence of leukocytes, neomycin sulfate, and LISS.

Previous studies have shown that RBCs with residual WBCs stored in LISS and neomycin sulfate develop characteristics associated with enzyme-treated RBCs. During a mass screening program to antigen type donor RBCs, we observed that the Fya antigens on a RBC sample from an in-house panel became non-detectable with anti-Fya after incubation overnight in Diluent 2 from Micro Typing Systems, Inc. (MTS, Pompano Beach, FL). In response to this observation, we initiated an investigation to determine the cause. Tests were performed according to the manfacturer's instructions in MTS neutral gel cards or gel cards containing anti-IgG. We found that a reduction or loss of the Fya, Fyb, and M antigens occurs when RBCs were prepared from samples containing residual WBCs (as a source of enzymes) and subsequently incubated in media containing neomycin sulfate and LISS. We showed that the effect did not occur in the absence of neomycin sulfate. RBC antigens can be altered in LISS if they have first been exposed to neomycin. We recommend restricting the use of RBCs suspended in MTS Diluent 2 to the day of dilution (as indicated in the package insert) if preparing reagent RBCs from sources that were not leukoreduced and were stored in the presence of neomycin.