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[The effect of Rh-D immunoprophylaxis with hyperimmune anti-D-immunoglobulin on the occurrence of RhD immunization in pregnancy].

During prenatal immunohaematological examination in the period from January 1, 1991 to December 31, 1995, in the Croatian Institute of Transfusion Medicine we tested sera of 5107 RhD negative women. All of them had pregnancies in their medical history. The frequency of Rh immunization was 4.6% in 1991; 4.1% in 1992; 2.5% in 1993; 2.5% in 1994 and 2.4% in 1995. Rh immunization during the first pregnancy was observed in 0.46% of women, in 1.8% during the second, in 9.4% during the third, in 22.4% during the fourth pregnancy, and 33.8% in women with more than five pregnancies. In women that have no abortions in their medical history, anti-D alloantibodies were found with the frequency of 0.46% at the end of the first pregnancy, 1.2% at the end of the second pregnancy, 5.9% at the end of the third pregnancy, 14.3% at the end of the fourth pregnancy, and 15.3% in women with more than five pregnancies. The frequency of anti-D alloantibodies in women who in their medical history have only abortions is 3.4% after the first abortion, 10.5% after the second, 17.8% after the third and 20.8% after the fourth or more abortions. The frequency of antibodies of anti-D specificities in women who had abortions and births is 17.1% at the end of the third pregnancy, 26.2% at the end of the fourth pregnancy, and 42.7% after more than five pregnancies. The frequency of anti-D alloantibodies in women who were protected from Rh immunization by hyperimmune anti-D globulin is 1%. The obtained results demonstrate that prevention of Rh immunization by hyperimmune anti-D globulin does not comprise all the Rh negative women, and is especially inadequate after abortions and multiple pregnancies.

Female↗

Three molecular structures cause rhesus D category VI phenotypes with distinct immunohematologic features.

Rhesus D category VI (DVI) is the clinically most important partial D. DVI red blood cells were assumed to possess very low RhD antigen density and to be caused by two RHD-CE-D hybrid alleles. Because there was no population-based work-up, we screened three populations in central Europe for DVI. Twenty-six DVI samples were detected and examined by exon-specific RHD polymerase chain reaction with sequence-specific primers (PCR-SSP). A new genotype, hereby designated D category VI type III, was characterized as a RHD-Ce(3-6)-D hybrid allele by sequencing of the cDNA, parts of intron 1, and by PCR-restriction fragment length polymorphism (PCR-RFLP) of intron 2. Rhesus introns 5 and 6 were sequenced and the 3' breakpoints of all known DVI types shown to be distinct. We differentiated the 5' breakpoints of DVI type I and DVI type II by a newly devised RHD-PCR. Thus, the DVI phenotype originated in at least three independent molecular events. Each DVI type showed distinct immunohematologic features in flow cytometry. The number of RhD proteins accessible on the red blood cells' surface of DVI type III was normal (about 12,000 antigens/cell; DVI type I, 500; DVI type II, 2,400) based on the determination of an RhD epitope density profile. DVI type II and DVI type III occurred as CDe haplotypes, and DVI type I as a cDE haplotype. The distribution of the DVI types varied significantly in three German-speaking populations. Genotyping strategies should take account of allelic variations in partial RhD. The reconsideration of previous serologic and clinical data for partial D in view of the underlying molecular structures may be worthwhile.

Alleles↗

[Rhesus factor determination of the genome of stereomicroscopy selected amniocytes].

This study intended to develop a molecular biological method for the determination of the rhesus blood type through the genome of amniocytes. Using micropipettes amniocytes were isolated under a stereomicroscope from amniotic fluid samples. Single or several (> or = 10) amniocytes, respectively, were pipetted into 0.2 ml vials each, and were lysated at 65 degrees C for 15 min using an alkaline solution making the DNA accessible to polymerase chain reaction. Thereafter, the rhesus D gene as well as the CE gene were amplified with nested primers in two subsequent PCR. Finally, the amplification product was electrophoretically analyzed using agarose gel. When single aminocytes were analysed the amplification rate was 64% for the rhesus D gene, and 80% for the rhesus CE gene; "allele specific non amplification" was observed in 24% of cases. With 10 or more amniocytes per vial the amplification rate was 100%. This method permits to reliably predict the fetal rh-type at low costs within about 4 hours.

Amniocentesis↗

Perinatal mortality in Rh alloimmunized patients.

OBJECTIVE: Evaluate and compare the perinatal mortality of Rh-negative pregnancies managed at São Paulo Federal University during a 9-year period, using either amniocentesis or middle cerebral artery peak systolic velocity. METHOD: Descriptive observational study involving 291 consecutive Rh-negative pregnancies managed between January 1995 and January 2004. The perinatal mortality of 99 alloimmunized patients was compared with 192 Rh-negative unimmunized patients (control group). The perinatal mortality of patients managed with amniocenteses was compared to those managed with Doppler studies. RESULTS: There were 74 patients managed with amniocenteses and 25 managed with Doppler studies. Perinatal mortality was significantly higher in the 99 Rh-negative isoimmunized patients than in the 192 unimmunized patients (12.1% versus 1%, p=0.0001) and did not differ according to the management protocol used (amniocentesis 13.5% versus cerebral Doppler 8.0%, p=0.725). Mean gestational age and mean weight at birth in pregnancies managed with amniocenteses (35.7 weeks and 2586 g) did not differ significantly from those managed with Doppler (36.3 weeks and 2647 g). CONCLUSIONS: Perinatal mortality in Rh-negative alloimmunized patients remains high and does not differ whether pregnancies are managed through amniocentesis or cerebral Doppler evaluation.

Adult↗

Carboxyhemoglobin levels in neonatal immune hemolytic jaundice treated with intravenous gammaglobulin.

In order to examine the effect of intravenous immunoglobulin (IVIG) on the rate of hemolysis in immune hemolytic hyperbilirubinemia, we measured the carboxyhemoglobin levels of 5 newborn infants who were subjected to IVIG treatment. The pretreatment rate of hemolysis, in the 5 patients with isoimmune hemolytic jaundice (3 patients with Rh hemolytic disease of the newborn and 2 patients with ABO hemolytic disease of the newborn), as reflected by carboxyhemoglobin levels was higher than the rate of hemolysis in normal newborn infants. In 4 out of the 5 patients treated with IVIG, there was a rapid decline ( > 30%) of carboxyhemoglobin levels, a pattern which was different from that observed in normal newborn infants with no hemolytic jaundice and in 3 untreated patients with ABO hemolytic disease of the newborn. None of the treated patients required an exchange transfusion. Our preliminary results support the theory that the attenuation of jaundice observed following IVIG treatment in patients with immune hemolytic hyperbilirubinemia is caused, at least in part, by the reduction in hemolysis.

ABO Blood-Group System↗

Severe hemolysis due to a donor anti-D antibody after heart-lung transplantation. Association with lung and blood chimerism.

Hemolysis due to donor-derived red cell antibodies is a potential complication after minor ABO-mismatched solid organ transplantation. It has also been described in Rh-positive recipients receiving organs from previously isoimmunized Rh-negative donors. We report on a group O, Rh(D)-positive patient who received a heart-lung transplant from a group O, Rh(D)-negative donor presenting an anti-D antibody. Severe hemolytic anemia developed in the early postoperative period and lasted for 3 mo. Anti-D antibodies that were of donor origin were found in bronchoalveolar lavage fluid and serum. In addition, using the polymerase chain reaction for human leukocyte antigen class II genotyping, we were able to demonstrate the presence of a mixed population of donor and recipient cells in the lungs and the peripheral blood of the recipient. The chimeric state in the blood persisted until at least the fifty-seventh postoperative day. This first case report of hemolysis due to Rh(D) antibody after heart-lung transplantation emphasizes the need for routine red cell antibody screening of donors and for close monitoring of signs of red cell destruction in the recipient in cases of pretransplant donor isoimmunization.

Adult↗

Comparison of quantitative acid-elution technique and flow cytometry for detecting fetomaternal hemorrhage.

A study of the Kleihauer-Betke acid-elution technique for quantitating fetomaternal hemorrhage was performed to assess intra- and inter-technologist accuracy and precision as well as to delineate the statistically valid domain of the test as usually performed. The results were then compared to a parallel study quantitating fetomaternal hemorrhage by flow cytometry. Additionally, a statistical model for estimating efficacy of treatment with Rh immune globulin in the prevention of pregnancy-associated Rh(D) isoimmunization was developed. The results indicate that the acid-elution technique can be performed in a reproducible manner with acceptable accuracy and precision with whole blood fetomaternal hemorrhages 25 ml and higher if a background correction for false positive identification of fetal cells is included. Flow cytometric determination reveals significantly increased accuracy in comparison to the corresponding Kleihauer-Betke results.

Adult↗