Introduction: Determination of RhD variants in blood donors, pregnant women, and newborns is important for transfusion strategies, in order to prevent RhD alloimmunisation and hemolytic disease of fetuses and newborns. Implementation of molecular RHD typing in two transfusion institutes is presented in this article, from Banja Luka (Bosnia and Herzegovina) and Belgrade (Serbia). Study Design and Methods: Blood donors’ RhD was checked by direct agglutination assays (tube) and indirect antiglobulin test (gel). Molecular RHD typing was performed by PCR-SSP with fluorometric signal detection in both centres. Donors were selected by weak RhD serological reactivity (Banja Luka, 85 samples; Belgrade, 62 samples) or serologically RhD-negative C/E-positive results (Banja Luka, 92 samples; Belgrade, 61 samples). Results: Among serologically determined weak D donors from the institute from Banja Luka, weak D type 3 was the most frequent (58.8%), followed by type 1 (35.3%) and DNB (1.2%), whereas results obtained at the Belgrade institute were distributed between weak D type 1 (41.9%), type 3 (30.7%), type 14 (6.5%), type 15 (1.6%), and DNB with anti-D (1.6%). In 17.7% of serologically typed weak D samples from the Belgrade institute, the molecular typing result was standard D. Additionally, RHD presence was detected in 9.8% of serologically RhD-negative, C/E-positive samples from both institutes. Conclusion: Rh molecular testing was successfully implemented in both blood transfusion institutes in Banja Luka and Belgrade. This study proved the efficiency of serological algorithms for weak D, as well as the presence of the RHD gene among serologically tested RhD-negative, C/E-positive samples.
Introduction: The aim of this study was to investigate the allele and genotype frequencies of 8 human platelet antigen (HPA) systems among blood donors from the Blood Transfusion Institute of Serbia and to compare them with published studies. These data would be useful to establish the basis for a platelet apheresis donor registry. Material and Methods: Seventy-two unrelated male platelet apheresis/blood donors from Serbia were typed for 8 HPA systems (HPA-1 to HPA-6, HPA-9, and HPA-15) via the FluoGene method, based on polymerase chain reaction-sequence-specific amplification (PCR-SSP; PCR using sequence-specific primers) with fluorometric signal detection. Allele and genotype frequencies were estimated by direct counting and compared to the expected genotype frequencies according to the Hardy-Weinberg principle. The transfusion mismatch probability was calculated for every HPA specificity. Results: The allele frequencies were: HPA-1a, 0.868; HPA-1b, 0.132; HPA-2a, 0.917; HPA-2b, 0.083; HPA-3a, 0.611; HPA-3b, 0.389; HPA-5a, 0.903; HPA-5b, 0.097; HPA-9a, 0.993; HPA-9b, 0.007; HPA-15a, 0.472; and HPA-15b, 0.528. For HPA-4 and HPA-6 only allele a was detected. Discussion: The HPA allele frequencies of European populations showed no significant differences in comparison with our results. Statistically significant differences were revealed in comparison with some populations of non-European origin. In the tested donors no HPA-2 bb genotype was detected, but we found 1 donor with the rare HPA-9b allele. The biggest transfusion mismatch probability in the Serbian population is for systems HPA-15 (37.4%) and HPA-3 (36.2%), which means that more than a third of random transfusions could cause mismatch in these systems. This study was enabled by the introduction of molecular HPA typing, and it provides initial results of the HPA allele and genotype frequencies in the population of blood donors in Serbia. They will be used to provide a compatible blood supply on demand for treating patients with alloimmune thrombocytopenic disorders. The successful implementation of PCR-SSP with fluorometric signal detection could be further complemented in the future by the introduction of high-throughput methods, which will largely depend on the available financial resources.
Background: Most people are either RhD positive or RhD negative, but there is also a number of persons with D antigen variants. The aim of this study was to prove, by using molecular diagnostic tests, whether the RHD gene and D antigen on the red cell membrane of the blood donors serologically typed as RhD-negative with RhD phenotype Ccddee and ccddEe, are so weak that they cannot be proven by serology techniques or the available anti-D test serums. Methods: Samples used are those of regular voluntary donors who were serotyped as RhD-negative, C and/or E positive. Samples were collected from voluntary donors at the Institute for Transfusion Medicine of the Republic of Srpska during the period from April 2016 to December 2018. Results: Among the serologically proven RhD-negative donors, 346 had C and/or E in their phenotype and those were subjected to molecular screening test. Conclusion: The first results of molecular typing match those published in literature, i.e. the RHD gene is present in some serologically RhD-negative forms, which was proven by molecular testing.
Introduction: The Rh system is very complex, polymorphous and the most significant for clinical practice, along with the ABO blood group system. The D antigen is the most important antigen in the Rh system and the most immunogenic one, following the ABO antigens. The D antigen, which consists of a mosaic of epitopes, is determined in all the blood donors and patients. Most people are either RhD positive or RhD negative, but there is a certain number of people who have a variation of the D antigen, which are called weak D, partial D and DEL phenotypes. Aim of the Study: The objective is to use molecular methods to determine whether blood donors in the Republic of Srpska (with whom a serological weak D antigen has been detected) really have the weak D antigen, partial D, a combination of these two variants or if their D antigen is normally present, but the used anti-D serum tests did not have the avidity needed to prove the presence of this antigen in blood donors. Patients and Methods: Blood samples were used from regular blood donors, who had been determined as persons with a weaker D antigen (based on the agglutination strength) using serological techniques, the test tube method, the microplate method and the gel method. To determine the blood groups and red blood cell/erythrocyte antigen typing, the following methods were applied: a) test tube method or agglutination in an aqueous environment, b) gel method, c) microplate method and d) molecular determination of blood groups. Results: Blood group samples were collected from April 2016 to February 2017 in the Institute for Transfusion Medicine of Republika Srpska. During this period, blood was collected from 8153 voluntary donors. It was serologically proved that 40 donors (0.49%) had the weak D antigen. All results where the weak D antigen was determined serologically were confirmed by molecular testing. 23 respondents were proved to have weak D type 3 (0.28%), while 17 had weak D type 1 (0.20%). Conclusion: The results from the first molecular testing of our population is in accordance with the results of frequency of weak D antigen in the populations of other European countries, though it did show a small advantage of weak D type 3 over weak D type 1.
From multi-photon to single molecule, the past several decades have witnessed a revolution in fluorescent microscopy. These techniques have revealed the inner working of cells and tissue and have relied on symbiotic advances in advanced molecular probes, light emitting molecules and particles, and novel instrumentation. Following on these developments, researchers began to develop functional nanomaterials or materials that can response to their environment. One of the first such molecules reported electric fields, allowing neuron signaling to be observed. However, the optical signal generated by voltage reporters is often low, placing limitations on the measurements that can be performed. Thus, material scientists and chemists began to pursue the development of alternative systems. In parallel, the fields of organic solar cells and integrated photonics were actively pursuing the design of materials with similar active properties, thus forming a foundation for improved functional organic imaging agents. In this talk, I will discuss some of our recent work in developing functional imaging agents for multi-wavelength and multi-photon live-cell imaging, focusing on recent molecular designs performed using density functional theory as well as in vitro studies.
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