ABSTRACT
【Objective】 To solve the difficulty of RhD blood group typing in a patient with double population(DP) of red blood cells for RhD antigen by serological and genotyping analysis. 【Methods】 Separation of the two populations of red blood cells of the patient was performed using capillary centrifugation method. ABO, RhD and RhCE typing, direct anti-human globulin test (DAT), irregular antibody screening, antibody identification and blood crossmatching of the patient were conducted using the standard serological methods. The hybrid Rhesus zygosity analysis of the RHD gene was performed by PCR-RFLP method. RHD and RHCE genotype of the patients were identified by PCR-SSP method. 【Results】 The patient was B type but with DP of red blood cells for RhD, Rhc and RhE antigens. DAT of the patient was positive and the alloanti-D was detected in serum. The RHD zygosity was D-/D- homozygote. PCR-SSP testing showed the RHD gene deletion (RHD * 01N. 01/01N.01 genotype) and Ccee of RHCE genotype in the patient, which was consistent with RHD zygosity analysis. 【Conclusion】 This is a special case with D-negative phenotype which was wrongly detected as D-positive type after D-positive red blood cells transfusion in emergency. When the DP of red cells for D antigen encountered like this case, the RhD typing can be accurately determined by using RHD genotyping analysis to provide strong evidence to the clinical blood transfusion.
ABSTRACT
【Objective】 To explore the application of matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) in the genotyping of difficult blood typing samples, and to provide evidence for clinical blood transfusion. 【Methods】 Three ambiguous blood group samples, submitted to Shanghai Blood Center by Shanghai regional hospitals, were studied, of which Sample1 included the proband and his parents. Serological methods were used to perform blood group typing, direct antibody test, unexpected antibody screening and identification test. Blood group genotyping was performed by using the MALDI-TOF MS detection systeme stablished in our laboratory. Sanger sequencing was used to confirm gene mutation sites, and serological or flow methods were used to verify specific samples′ phenotype. 【Results】 Serological results indicated the existence of antibodies against high frequency antigens in sample 1 (including proband and her mother), 2 and 3. The genotyping results of MALDI-TOF MS showed that the proband of sample 1 was Di(a+ b+ ), her father was Di(a-b+ ), her mother was Di(a+ b-), sample 2 was p, and sample 3 was Jr(a-). Sequencing results of three samples were consistent with mass spectrometry typing results. Serological results showed that sample 2 had a p phenotype. The flow cytometry results suggested that sample 3 had a Jr(a-) phenotype. 【Conclusion】 For the first time, we applied MALDI-TOF MS technology to blood type genotyping of ambiguous clinical samples in China. Compared with other genotyping methods such as PCR-SSP, MALDI-TOF MS has the advantages of rapid detection, high throughput and high specificity, which would contribute to identification of difficult blood typing samples in the future, as well as rare blood group screening.
ABSTRACT
Background & objectives: In multitransfused thalassaemic patients, haemagglutination fails to phenotype the patient's blood group antigens due to the presence of donor-derived erythrocytes. DNA-based methods can overcome the limitations of haemagglutination and can be used to determine the correct antigen profile of these patients. This will facilitate the procurement of antigen-matched blood for transfusion to multitransfused patients. Thus, the aim of this study was to compare the serological phenotyping of common and clinically important antigens of Rh, Duffy, Kell, Kidd and MNS blood group systems with molecular genotyping amongst multitransfused thalassaemic patients. Methods: Blood samples from 200 patients with thalassaemia and 100 'O' group regular blood donors were tested using standard serological techniques and polymerase chain reaction-based methods for common antigens/alleles (C, c, D, E, e, Fya, Fyb, Jka, Jkb, K, k, M, N, S, s). Results: Genotyping and phenotyping results were discordant in 77 per cent of thalassaemic patients for five pairs of antithetical antigens of Rh, Duffy, Kell and Kidd blood group systems. In the MNS blood group system, 59.1 per cent of patients showed discrepancy. The rate of alloimmunization among thalassaemics was 7.5 per cent. Interpretation & conclusions: Molecular genotyping enabled the determination of the actual antigen profile in multitransfused thalassaemia patients. This would help reduce the problem of alloimmunization in such patients and would also aid in the better management of transfusion therapy.