Study Of Anti-D Antibodies In RhD-negative Recipients Transfused With DEL Red Blood Cells

Objective: The DEL variants are detectable only by sensitive absorbtion-elution test, because there were a small number of D antigen molecules in membrane of DEL red blood cell. Recently, the safety problom of DEL clinical transfusion has attracted high international attention. The Japanese and South Korean researchers reported successively that the"Asia type"DEL could induce anti-D immunization in D-negative recipients (primary and second immunization), and in 2001 in China, we found one report of anti-D alloimmunization in a D-negative recipient after the transfusion of DEL red blood cells. But in most reports there were no transfution reactions and anti-D antibody in Rh-negative individuals who received DEL red blood cells. In this paper, we tracked 20 Rh-negative recipients transfused with DEL red blood cells with the aim to determine whether D antigens in DEL individuals express alloimmunization by observing their clinical transfusion reactions and screening whether to produce anti-D antibodies in their serum.Methods:1 Screening DEL red blood cell in donorsRhD-DEL is defined on the serological aspect at present. The D antigen could be detected among individuals determined to be Rh-negative by the indirect antiglobulin test (IAT), using a more sensitive technique called the"absorption-elution"tests. We screened DEL red blood cells of donors by serological methods, including microplate saline test, indirect antiglobulin test and absorbtion-elution tests. In Rh blood group system, there are five common antigens: D, C, E, c and e, which can be identified using corresponding monoclonal antisera.2 Identification of RhD blood group and anti-D antibodies in the RhD-negative recipients transfused with DEL red blood cellsTo determine whether D antigens in DEL individuals express immunogenicity, we tracked 20 Rh-negative recipients transfused with DEL variants. Their blood was collectted at the stage from 5 days to six months after transfusion. All the people were Han Chinese aged 14 to 78 years old (average 38.75) and 10 individuals were female. We tested the red blood cells of patients by serological methods (microplate saline test, indirect antiglobulin test and absorbtion-elution tests) and RhD gene analysis. For RhD gene analysis, DNA was extracted using Tiangeng blood DNA kits (Beijing), the PCR-SSP was performed in accordance with Ready gene Drag Kits (inno-Trin Blood Group SSP). Irregular antibodies were screened by manual polybrene (MP), indirect antiglobulin test (IAT) and microtubes gel test (MGT) for the serum of samples. Antibody titer and Ig-types were identified by O-panel cells for those positive anti-Rh samples.Results:1. 421 RhD negative donors from Anhui Blood Center, from January 2010 to March 2011, were comfirmed by IAT. The results of adsorption-elution tests showed that 108 of 421 (25.6%) were DEL and 313 of 421 (74.4%) were true Rh-negative. The RhC, c, E and e antigen phenotypes were typed in all DEL samples, and the results showed 85.2% of the DEL samples had C antigen. 2. The results of adsorption-elution tests for 20 patient samples we collected showed that 5 (25%) were DEL and 15 (75%) were true Rh-negative. PCR-SSP showed all 5 DEL samples carried the RhD(K409K)allele. None of the 5 DEL-positive individuals were detected to have allo-anti-D, while anti-D was detected only in true Rh-negative individuals.Two women of 20 patients were identified as positive with irregular antibody by MP, IAT and MGT, and their antibody titer were 32 and 512 respectively. Irregular antibodies were not detected among all male individuals. The two positive irregular antibodies were confirmed as allo-anti-D specificity by O-panel cells. One of the women, 22-year-old, had a history of pregnancy, and her son's blood was RhD positive. That suggested alloimmunization by pregnancy may be the major cause of anti-D antibodies. In other case, anti-D antibodies in a 36-year-old woman with five histories of gestations or parturitions were always positive before and after the DEL transfution, and the titers were 512. Her case report showed she gave birth to one health girl in 1997 and received 2 units of cross-match-compatible group B DEL red blood cells. There was no evidence of acute or delayed hemolytic transfusion reactions after those transfusions and no remarkable changes in levels of hemoglobin and indirect bilirubin. She discharged from hospital after two weeks.Conclusions:1. RhC antigen is usually positive in"Asia type"DEL phenotype, so it can be used as an important marker for preliminary screening of"Asia type"DEL.2. In 20 samples we collected, anti-D antibody is not detected among DEL phenotype individuals and DEL phenotype is not found in anti-D antibody producing individuals, which suggests that anti-D antibody can't be induced in DEL phenotype individuals. 3. None of the RhD-negative men transfused with DEL red blood cells developed allo-anti-D. And there was no transfusion reactions on a ture RhD-negative woman with anti-D antibody after transfused with DEL red blood cells. These facts indicate that the RhD-negative recipients are not at the risk of alloimmunization after transfused with DEL red blood cells.