PubMed | Bristol Institute for Transfusion science, International Blood Group Reference Laboratory, Statistics and Clinical Studies, University College London and 3 more.
Type: | Journal: Cytometry. Part B, Clinical cytometry | Year: 2016
Bone marrow examination has been the confirmatory test for congenital dyserythropoietic anemia type II (CDAII). Occasional spherocytes on peripheral blood smear can confound the diagnosis. Since a screening test is still unavailable, we explored the feasibility of using flow cytometry as a preliminary screening method.Thirteen monoclonal antibodies with specificities for eight erythrocyte membrane proteins were used in FACS analysis to probe the cellular features of red cells from CDAII, normal adults, hereditary spherocytosis (HS), and cord red cells. Confocal microscopy was performed on normal and CDAII to determine the overall distribution of CD44 and CD47. Their expression levels on cultured erythroblasts were also analyzed.The densely stained band 3 as seen in CDAII in gel electrophoresis was also obtained for Dantu phenotype. Likewise analysis of CDAII cases (n=26) using the eosin-5maleimide (EMA) binding test found 57% of patients giving results either positive or in the grey area for HS. Enhanced fluorescence of CD44 was detected in 96% of the CDAII patients, and anti-CD47 binding was also elevated to a lesser degree. Although RNA expressions of CD44 and CD47 in the cultured erythroblasts of normal controls and CDAII were similar, confocal microscopy revealed more CDAII red cells giving elevated fluorescence than normal red cells.A distinction between CDAII and HS can be made using the EMA Binding test and anti-CD44 binding. Confirmation of CDAII can subsequently be made based on clinical presentation together with either bone marrow examination or DNA sequencing of SEC23B. 2016 International Clinical Cytometry Society.
PubMed | Southmead Hospital, University of Bristol and International Blood Group Reference Laboratory
Type: Journal Article | Journal: BJOG : an international journal of obstetrics and gynaecology | Year: 2015
To determine whether a policy of offering cffDNA testing to all RhD-negative women at about 16 weeks gestation to avoid anti-D administration when the fetus is RhD-negative could be implemented successfully in the NHS without additional funding.Prospectively planned observational service implementation pilot and notes audit.Three maternity services in the South West of England.All RhD-negative women in a 6-month period.Prospective, intervention, cross-sectional observational study, using pre-intervention data as controls.Proportion of suitable women who offered and accepted the test. Accuracy of the cffDNA result as assessed by cord blood group result. Fall in anti-D doses administered.529 samples were received; three were unsuitable. The results were reported as RhD-positive (n = 278), RhD-negative (n = 185) or inconclusive, treat as positive (n = 63). Cord blood results were available in 502 (95%) and the only incorrect result was one case of a false positive (cffDNA reported as positive, cord blood negative - and so given anti-D unnecessarily). The notes audit showed that women who declined this service were correctly managed and that anti-D was not given when the fetus was predicted to be RhD-negative. The total use of anti-D doses fell by about 29% which equated to about 35% of RhD-negative women not receiving anti-D in their pregnancy unnecessarily.We recommend this service is extended to all UK NHS services.
Seltsam A.,Red Cross |
Wagner F.,Red Cross |
Lambert M.,National Blood Center |
Bullock T.,International Blood Group Reference Laboratory |
And 5 more authors.
Transfusion | Year: 2014
Alloantibodies to high-prevalence red blood cell (RBC) antigens are not easily identified by routine serologic techniques. This multicenter study was conducted to test the effectiveness of recombinant blood group proteins (rBGPs) at regional and international RBC reference laboratories. Study Design and Methods Single or mixed soluble rBGPs (Lu, Yt, Kn, JMH, Sc, Rg, Ch, Do, and Cr) were assessed for their ability to inhibit the reactivity of antibodies to specific antigens. Initially, the effect of rBGPs was validated by testing panels of well-characterized patient serum samples containing antibodies to high-prevalence antigens in the hemagglutination inhibition assay. Subsequently, the rBGPs were prospectively used for routine antibody identification and the results were compared to those obtained with RBC-based diagnostics. Results Panels of predefined antibodies to high-prevalence antigens were completely and specifically neutralized by the corresponding rBGP specificities. For prospective identification, antibodies to high-prevalence antigens (n=62) were specifically inhibited by the corresponding rBGP specificities except for some Complement Receptor 1-related antibodies, which may be directed to epitopes not expressed on the truncated recombinant Kn. In 14 cases, additional clinically relevant alloantibodies were identified. In cross-matching, the rBGPs were successfully used to inhibit the reactivity of clinically irrelevant antibodies to high-prevalence antigens to determine compatibility between donor and recipient. Conclusion rBGPs enable the identification of antibodies to high-prevalence antigens without the need for rare RBC reagents, which are often unavailable. Underlying antibodies can be reliably detected and cross-matching results validated, resulting in a more efficient blood supply for immunized patients. © 2014 AABB.
Daniels G.,International Blood Group Reference Laboratory |
Daniels G.,Bristol Institute for Transfusion science
Human Blood Groups: 3rd edition | Year: 2013
Human Blood Groups is a comprehensive and fully referenced text covering both the scientific and clinical aspects of red cell surface antigens, including: serology, inheritance, biochemistry, molecular genetics, biological functions and clinical significance in transfusion medicine. Since the last edition, seven new blood group systems and over 60 new blood group antigens have been identified. All of the genes representing those systems have now been cloned and sequenced. This essential new information has made the launch of a third edition of Human Blood Groups, now in four colour, particularly timely. This book continues to be an essential reference source for all those who require clinical information on blood groups and antibodies in transfusion medicine and blood banking. © 2013 Geoff Daniels.
Hill M.,University College London |
Hill M.,Great Ormond Street Hospital for Children NHS Trust |
Finning K.,International Blood Group Reference Laboratory |
Martin P.,International Blood Group Reference Laboratory |
And 6 more authors.
Clinical Genetics | Year: 2011
The effectiveness and clinical utility of non-invasive prenatal diagnosis (NIPD) for fetal sex determination using cell-free fetal DNA (cffDNA) was assessed by undertaking a prospective national audit of UK testing. NIPD was performed using real-time polymerase chain reaction analysis of the DYS14 or SRY gene in cffDNA extracted from maternal plasma. All cases referred for fetal sex determination from 1 April 2006 to 31 March 2009 were ascertained from two laboratories offering the test. Fetal gender determined by NIPD was compared with that based on ultrasound, invasive test or phenotype at birth. Indication and rate of invasive testing was ascertained. In the first year, results were issued in 150/161 pregnancies tested. Of the 135 with outcome data, results were concordant in 130/135 [96.3% (95% CI 91.6-98.8%)]. Reporting criteria were changed and in the subsequent 511 pregnancies the concordancy rate increased to 401/403 [99.5% (95% CI 98.2-99.9%)]. Over the 3 years only 32.9% (174/528) underwent invasive testing. NIPD for fetal sex determination using cffDNA is highly accurate when performed in National Health Service laboratories if stringent reporting criteria are applied. Parents should be advised of the small risk of discordant results and possible need for repeat testing to resolve inconclusive results. © 2010 John Wiley & Sons A/S.
Kumpel B.M.,International Blood Group Reference Laboratory |
Macdonald A.P.,Raigmore Hospital |
Bishop D.R.,Red Cell Immunohaematology |
Yates A.F.,Cheltenham General Hospital |
Lee E.,Red Cell Immunohaematology
Transfusion Medicine | Year: 2013
SUMMARY: Background: Fetomaternal haemorrhage (FMH) assessment by the Kleihauer-Betke test (KBT) is rapid but semi-quantitative and liable to false positive results. Objectives: To compare FMH estimated by KBT with flow cytometry (FC) quantitation for 37 patients with massive FMH, obstetric risk factors or technical problems. Methods: Maternal blood was sent for analysis by FC after KBT. A variety of reagents including anti-haemoglobin F (HbF), anti-D and combined anti-HbF/anti-carbonic anhydrase (CA) were used. Results: Eight cases of massive FMH (>100mL fetal cells) causing fetal death or severe neonatal anaemia in late gestation were confirmed by FC. Anti-HbF FC identified maternal F cells and fetal cells. In some cases these red cell populations merged but they could be differentiated by anti-CA, labelling F cells only. Using KBT, false positive FMH results were obtained for 12 patients, who had strongly stained cells that were then shown by FC to be maternal F cells. All these patients had increased F cells (>5% of total red cells) whereas only 16% of patients with FMH and 22% of donors had elevated F cells. In contrast, anti-D FC was simple and rapid, quantitating D-positive FMH in all 15 D-negative patients except one with massive FMH of weak D fetal cells. Leucocytes in four samples bound anti-D, variably, giving erroneously high FMH, but they could be eliminated from FC analysis using combined anti-D/anti-CD45. Conclusion: FMH quantitation using anti-D by FC is suitable for the majority of maternal samples and could enable accurate targeted dosing of anti-D prophylaxis. © 2013 British Blood Transfusion Society.
Tilley L.,International Blood Group Reference Laboratory |
Grimsley S.,International Blood Group Reference Laboratory
Transfusion and Apheresis Science | Year: 2014
Blood group genotyping has many advantages over conventional phenotyping for both blood donors and patients, and a number of high-throughput methods have now been developed. However, these are limited by a requirement for existing knowledge of the relevant blood group gene polymorphisms, and rare or novel mutations will not be detected. These mutations could be successfully identified by DNA sequencing of the blood group genes, and such an approach has been made feasible by the introduction of Next Generation Sequencing (NGS) technology. NGS enables many genes from multiple samples to be sequenced in parallel, resulting in sequencing information that could be used to obtain accurate blood group phenotype predictions in both blood donors and patients. © 2014 Elsevier Ltd.
Daniels G.,International Blood Group Reference Laboratory
British Journal of Haematology | Year: 2013
Anti-D (-RH1) of the Rh blood group system is clinically important as it causes haemolytic transfusion reactions and haemolytic disease of the fetus and newborn. Although most people are either D+ or D-, there is a plethora of D variants, often categorized as either weak D or partial D. These two types are inadequately defined and the dichotomy is potentially misleading. DVI is the D variant most commonly associated with anti-D production and UK guidelines recommend that patients are tested with anti-D reagents that do not react with DVI. Weak D types 1, 2, and 3 are seldom, if ever, associated with alloanti-D production, so a policy recommendation would be to treat patients with those D variants as D+, to preserve D- stocks, whereas patients with all other D variants would be treated as D-. All donors with D variant red cells, including DVI, should be treated as D+. © 2013 Crown copyright.
Soothill P.W.,University of Bristol |
Finning K.,International Blood Group Reference Laboratory |
Latham T.,International Blood Group Reference Laboratory |
Wreford-Bush T.,Haematology Southmead Hospital |
And 2 more authors.
BJOG: An International Journal of Obstetrics and Gynaecology | Year: 2015
Objective To determine whether a policy of offering cffDNA testing to all RhD-negative women at about 16 weeks' gestation to avoid anti-D administration when the fetus is RhD-negative could be implemented successfully in the NHS without additional funding. Design Prospectively planned observational service implementation pilot and notes audit. Setting Three maternity services in the South West of England. Population All RhD-negative women in a 6-month period. Methods Prospective, intervention, cross-sectional observational study, using pre-intervention data as controls. Main outcome measures Proportion of suitable women who offered and accepted the test. Accuracy of the cffDNA result as assessed by cord blood group result. Fall in anti-D doses administered. Results 529 samples were received; three were unsuitable. The results were reported as RhD-positive (n = 278), RhD-negative (n = 185) or inconclusive, treat as positive (n = 63). Cord blood results were available in 502 (95%) and the only incorrect result was one case of a false positive (cffDNA reported as positive, cord blood negative - and so given anti-D unnecessarily). The notes audit showed that women who declined this service were correctly managed and that anti-D was not given when the fetus was predicted to be RhD-negative. The total use of anti-D doses fell by about 29% which equated to about 35% of RhD-negative women not receiving anti-D in their pregnancy unnecessarily. Conclusions We recommend this service is extended to all UK NHS services. © 2015 Royal College of Obstetricians and Gynaecologists.
King M.-J.,International Blood Group Reference Laboratory |
Jepson M.A.,University of Bristol |
Guest A.,International Blood Group Reference Laboratory |
Mushens R.,International Blood Group Reference Laboratory
International Journal of Laboratory Hematology | Year: 2011
Introduction: Hereditary spherocytosis (HS) and hereditary pyropoikilocytosis (HPP, severe form of hereditary elliptocytosis) are unrelated red cell disorders caused by defects in distinct regions of the red cell cytoskeleton. The high predictive value of the eosin-5-maleimide (EMA)-binding test for the diagnosis of HS is because of its interaction with transmembrane proteins band 3, Rh protein, Rh glycoprotein and CD47, which are reduced on HS red cells. Our study was undertaken to determine why EMA-labelled HPP red cells were previously found to give much lower fluorescence readings than HS. Methods: Flow cytometry was used to determine the relative amounts of monoclonal antibodies bound to red cells from normal adults, HS and HPP groups. Confocal microscopy was used to visualise the overall staining pattern of the red cells with selected antibodies. Results: In flow cytometry, HPP red cells gave lower antibody binding to the four EMA-reactive membrane proteins than HS red cells and bound less antibody to glycophorins A and C, and CD59. Confocal images of Rh protein and band 3 immunostaining revealed a greater number of HPP red cells having partial or no fluorescence than in HS and normal controls. Conclusion: Lesser amounts of EMA-reactive membrane proteins were detected in HPP than HS red cells, thus confirming their lower fluorescence readings in the EMA-binding test. The concomitant reduction in glycophorins A and C, and CD59 in HPP could have caused cellular contraction, resulting in poikilocytosis. © 2010 Blackwell Publishing Ltd.