BACKGROUND: Blood group single nucleotide polymorphism genotyping probes for a limited range of polymorphisms. This study investigated whether massively parallel sequencing (also known as next-generation sequencing), with a targeted exome strategy, provides an extended blood group genotype and the extent to which massively parallel sequencing correctly genotypes in homologous gene systems, such as RH and MNS. STUDY DESIGN AND METHODS: Donor samples(n 5 28) that were extensively phenotyped and genotyped using single nucleotide polymorphism typing, were analyzed using the TruSight One Sequencing Panel and MiSeq platform. Genes for 28 protein-based blood group systems, GATA1, and KLF1 were analyzed. Copy number variation analysis was used to characterize complex structural variants in the GYPC and RH systems. RESULTS:The average sequencing depth per target region was 66.2 6 39.8. Each sample harbored on average 43 6 9 variants, of which 10 6 3 were used for genotyping. For the 28 samples, massively parallel sequencing variant sequences correctly matched expected sequences based on single nucleotide polymorphism genotyping data. Copy number variation analysis defined the Rh C/c alleles and complex RHD hybrids. Hybrid RHD*D-CE-D variants were correctly identified, but copy number variation analysis did not confidently distinguish between D and CE exon deletion versus rearrangement. CONCLUSION:The targeted exome sequencing strategy employed extended the range of blood group genotypes detected compared with single nucleotide polymorphism typing. This single-test format included detection of complex MNS hybrid cases and, with copy number variation analysis, defined RH hybrid genes along with the RHCE*C allele hitherto difficult to resolve by variant detection. The approach is economical compared with whole-genome sequencing and is suitable for a red blood cell reference laboratory setting.H uman blood group antigens are of significance in transfusion medicine, because patients who have made antibodies to red blood cell antigens are at risk of being affected by hemolytic transfusion reactions after the transfusion of incompatible blood. The International Society of Blood Transfusion has defined 36 blood group systems and over 350 blood group antigens.1 Different blood group systems exhibit varying degrees of antigen polymorphism, and the clinical significance of red blood cell antibodies also varies. [2][3][4] As a minimum requirement in blood transfusion safety, all blood donors are screened for ABO and the D antigen as well as for blood group antibodies known to be clinically significant. 5The majority of antigens are missense mutations and a consequence of single nucleotide variants (SNVs); however, genetic variations, such as insertions/deletions and splice-site variants, have a qualitative and/or quantitative impact on antigen expression. Blood group systems, such as RH and MNS, exhibit an additional layer of genetic variation. These arise because each system comprises homologous genes in which gene crossover or g...

Targeted exome sequencing resolved complex serology problems and defined both novel blood group alleles (CD55:c.203G>A, ABCB6:c.1118_1124delCGGATCG, ABCB6:c.1656-1G>A, and RHD:c.452G>A) and rare variants on blood group alleles associated with altered phenotypes. This study illustrates the utility of exome sequencing, in conjunction with serology, as an alternative approach to resolve complex cases.

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"Fluothane" is a new volatile anaesthetic agent developed and manufactured by Imperial Chemical Industries (Raventos, 1956; Raventos and Suckling, 1956). It is the most promising member of a series of agents with non-flammable and non-explosive properties which are being investigated independently by the Medical Research Council. It is a clear, colourless, heavy liquid with a specific gravity of 1860 (cf. chloroform 1500), a boiling-point of 510 C., and a sweetish odour not unlike that of chloroform. Chemically, it is a halogenated ethane with the structural formula F BrFluothane has been administered as the sole or the principal anaesthetic agent to a number of volunteers and to 310 patients (varying in age from 7 months to 91 years) for a variety of surgical procedures (lasting from 3 to 150 minutes). There were no deaths under anaesthesia in the series, although many of the patients came to operation in a poor physical state. The pre-operative conditions included diabetes, obstructive jaundice, cardiac failure, thyrotoxicosis, and asthma, but none were affected adversely by the anaesthetic.One patient died during the post-operative period, but this was probably not attributable to the anaesthetic.An 84-year-old man received open drop fluothane for the amputation of a gangrenous toe. The administration was uneventful and lasted for 15 minutes. The patient recovered consciousness before leaving the operating theatre and was quite well for the next four days. He then became increasingly irrational, developed bronchopneumonia, and died on the seventh post-operative day.Tlhe object of this investigation was to confirm the promising results obtained in animal trials ; to ascertain in humans the effect of known concentrations of fluothane vapour; and to exploit the administration of the drug by simple methods. Methods of AdmuinistrationSince it was known that the administration of fluothanein conjunction with nitrous oxide and oxygen from a standard Boyle apparatus was being investigated elsewhere (Johnstone, 1956) Macintosh, 1956) modified and calibrated to deliver known concentrations of fluothane vapour in air. In this way it has been possible to observe the action of fluothane without the complication of other anaesthetic agents, and to obtain an im-*The trade name given by Imperial Chemical Industries to 2-bromo-2-chloro-1,1,1-trifluoro-ethane. The drug is not yet freely available. pression of the effec:s produced by known concentrations of the agent.Open Drop Induction.-Fluothane has been administered by the open drop method to 63 patients and to a further 29 after the preliminary injection of a " sleep " dose of thiopentone (rarely more than 0.2 g.).. In most instances, induction was smooth and pleasant, producing unconsciousness in about one minute. Swallowing during induction was not uncommon, but this did not interfere with the administration and was not followed by vomiting. Coughing was rare and was always relieved by raising the mask. Struggling and breath-holding were unusual. Surgical intervention cou...

BACKGROUND Blood donors whose red blood cells (RBCs) exhibit a partial RhD phenotype, lacking some D epitopes, present as D+ in routine screening. Such phenotypes can exhibit low‐frequency antigens (LFAs) of clinical significance. The aim of this study was to describe the serologic and genetic profile for a blood donor with an apparent D+ phenotype carrying a variant RHD gene where D Exons 5 and 6 are replaced by RHCE Exon (5‐6). STUDY DESIGN AND METHODS Anti‐D monoclonal antibodies were used to characterize the presentation of RhD epitopes on the RBCs. RHD exon scanning and DNA sequencing of short‐ and long‐range polymerase chain reaction amplicons were used to determine the RHD structure and sequence. Extended phenotyping for LFAs RH23 (DW) and Rh32 was performed. RESULTS The donor serology profile was consistent with partial RhD epitope presentation. The donor was hemizygous for an RHD variant allele described as RHD*D‐CE(5‐6)‐D hybrid. The RHCE gene insert is at least 3.868 kb with 5′ and 3′ breakpoints between IVS4 + 132–c.667 and IVS6 + 1960–IVS6 + 2099, respectively. The sequence for this hybrid was assigned GenBank Accession Number KT099190.2. The RBCs were RH23 (DW)+ and Rh32–. CONCLUSION A novel RHD*D‐CE(5‐6)‐D hybrid allele encodes a partial RhD epitope and carries the LFA RH23 (DW). This and the epitope profile resemble the partial DVa phenotype. Given that RBCs from this individual lack some RhD epitopes, there is an alloimmunization risk if the donor is exposed to D+ RBCs. Conversely, transfusions of RH23 (DW)+ cells to RH23 (DW)– recipients also pose an alloimmunization risk.