A flexible array format for large‐scale, rapid blood group DNA typing

Hashmi, Ghazala; Shariff, Tasmia; Seul, Michael; Vissavajjhala, Prabhakar; Hue‐Roye, Kim; Charles-Pierre, Dalisay; Lomas‐Francis, Christine; Chaudhuri, Asok; Reid, Marion E.

doi:10.1111/j.1537-2995.2005.04362.x

Cited by 146 publications

(151 citation statements)

References 20 publications

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“…This automated platform genotypes for 32 antigens in 12 different blood group systems, all encoded by simple SNPs. 34,35 The complexities and numerous alleles in the Rh blood group system have precluded testing for RH other than for the common non-variant forms of C/c and E/e antigens but automated testing is being developed. An automated test platform developed in Europe (BLOODChip, Progenika Biopharma S.A.) is now available in the United States; in addition to minor blood group antigens it includes an extensive number of variant RHD alleles.…”

Section: Rh Genotypingmentioning

confidence: 99%

Molecular biology of the Rh system: clinical considerations for transfusion in sickle cell disease

Chou

Westhoff²

2009

Hematology

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The last decade has witnessed an abundance of information detailing the genetic diversity of the RH locus which has exceeded all estimates predicted by serology. Well over 120 RHD and over 60 different RHCE alleles have been documented, and new alleles are still being discovered. For clinical transfusion medicine, RH genetic testing can now be used to determine RHD zygosity, resolve D antigen status, and detect altered RHD and RHCE genes in individuals at risk for producing antibodies to high-incidence Rh antigens, particularly patients with sickle cell disease (SCD).T ransfusion of patients with sickle cell disease (SCD) represents a significant challenge in clinical transfusion medicine with red blood cell (RBC) alloimmunization a primary and serious complication of transfusions. A major cause of alloimmunization in patients with SCD is the disparate distribution of red cell antigens between donors, who are primarily of European ancestry, and patients with SCD, who are primarily of African ancestry. Management of alloimmunization in SCD has been the subject of much debate, 1,2 and currently there is no standard approach. Over two thirds of alloantibodies formed by patients with SCD have Rh blood group specificities. Many programs transfuse patients with SCD with RBCs that are phenotype-matched for D, C/c, E/e, and K, and some also supply RBCs from African-American donors when possible. Although these approaches reduce the incidence of alloantibody production, patients still become alloimmunized. Genetic analysis of patients who develop antibodies in the face of conventional antigen matching has revealed that these patients carry altered RH alleles. RH Genes and Rh ProteinsTwo genes, RHD and RHCE, lie in close proximity on chromosome 1 and encode 416-amino acid Rh proteins: one encodes the D antigen, and the other encodes CE antigens in various combinations (ce, cE, Ce, or CE) (Figure 1). Each gene has ten exons; they are 97% identical and encode proteins that differ by 32 to 35 amino acids (Figure 1). This contrasts with most blood group antigens, which are encoded by single genes with alleles that differ TRANSFUSION MEDICINE ___________________________________________________________________________________ by only one or a few amino acids. For comprehensive reviews, see Westhoff 3 and Avent. The conventional RH genes shown in Figure 1 are found in all population groups, although with different frequencies. Commercial antibody reagents detect expression of the five principal Rh antigens: D, C, c, E, and e. Variant RHD and RHCE alleles encode Rh proteins with amino acid changes that cannot be distinguished serologically, but can be recognized by the immune system as foreign. Of relevance for transfusion in patients with SCD, RH alleles encoding altered C and e antigens are frequent in African ethnic groups. 5,6 Some patients with SCD are at risk for production of antibodies to both the RhCE and RhD proteins, a serious and potentially life-threatening complication. Variations of RhD

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Section: Rh Genotypingmentioning

confidence: 99%

Molecular biology of the Rh system: clinical considerations for transfusion in sickle cell disease

Chou

Westhoff²

2009

Hematology

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“…Oligonucleotide probes specific for the 16S rRNA genes of each of the 17 pathogens, as well as universal probes for Gram-positive, Gram-negative, and all bacteria, were designed (4, 12; N. Mezokh, K. Podual, and M. Seul, presented at the Association of Molecular Pathology AMP Annual Meeting, Orlando, FL, 16 November 2006), coupled to encoded beads stained with spectrally distinguishable combinations of fluorescent dyes, and embedded onto a bead array chip (2,6,10). After array assembly, the color code of each bead within the array was recorded.…”

mentioning

confidence: 99%

“…After array assembly, the color code of each bead within the array was recorded. Labeled PCR amplicon was hybridized to the bead array, followed by analysis of the fluorescence pattern by an image-reading microscope with fluorescence optics and a charge-coupled-device camera (2,6,10). Decoded image data were converted into normalized results and displayed as bar graphs.…”

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confidence: 99%

Bacterial Identification by 16S rRNA Gene PCR-Hybridization as a Supplement to Negative Culture Results

et al. 2011

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PCR-hybridization was compared to culture methods for evaluating suspected blood infections. A total of 231 clinical samples from blood culture bottles that were flagged positive by the BacT/Alert system or were negative 1 week after inoculation were tested. When the PCR-hybridization and culture method results were compared, the positive and negative concordance rates were 99.2% (122/123) and 89.5% (94/105), respectively. Of the negative blood cultures, 10.5% (11/105) were positive by PCR-hybridization. Supplemental testing of negative blood cultures may identify bacterial pathogens that are undetectable by culture methods.Blood culture is currently the gold standard method for identification of bacterial pathogens causing blood infections. Accurate and reliable identification of pathogens is critical so that proper and timely treatment can be initiated. Current culture procedures typically require from several days to a week for final results (1, 14), and incorrect results can occur, with false-positive and false-negative rates generally estimated at 2 to 3% (5, 8). Factors suspected of contributing to falsenegative culture results include insufficient blood sample inoculum, empirical or long-term antibiotic use prior to diagnosis, and infection due to fastidious organisms (3). This study compared identification of bacterial pathogens by traditional culture methods with PCR amplification of 16S rRNA genes from cultured blood samples in conjunction with hybridization to species-specific probes on a bead array chip.Clinical samples from aerobic blood culture bottles (BacT/ Alert SA standard aerobic culture media bottle, bioMérieux) were obtained from the clinical microbiology laboratory at the Children's Hospital Los Angeles. All of the clinical samples were obtained from pediatric patients. Generally, pediatric samples were used to inoculate one blood culture bottle, not a "set" of two bottles as is standard protocol for adult patients. Positive cultures were those with a positive result in the BacT/ Alert system within 1 week of inoculation, while negative cultures were those with a negative result 1 week after inoculation. A total of 231 blood cultures were selected for the study after they were flagged positive by BacT/Alert or upon a final negative result. After culture results were known, DNA was extracted from 500 l of blood culture broth in a class II cabinet using an alkaline lysis method (9, 13) and resuspended in a final volume of 500 l.

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“…Still we found 2 samples that we classified as indeterminate, as had normal phenotyping, but in molecular biology using multiplex PCR, where one did not present the exon 7 and the other did not submit the intron 4, but to accomplish the PCR by the method of micro arrangements RHD showed no changes. The genotyping were performed for the RHD gene, using the micro array technique "micro-array", which uses a multiplex PCR that does the simultaneous amplification of several DNA [23,24] fragments where polymorphisms occur, for a means of oligonucleotide probes deposited on a glass plate (glass, silica or other supports) which, when hybridized with target DNA fluoresce. For facilitate viewing of frequency of transfusions and presence of RhD variants, we conduct the following number of transfusions cuts, as shown in Table 3.…”

Section: Resultsmentioning

confidence: 99%

Study of Variations in RhD Holders of Sickle Cell Disease Patients of Amazon State, Brazil

Tezza¹,

Sanguino²,

Gomes³

et al. 2017

HTIJ

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Introduction: Transfusion therapy is a challenge for patients with sickle cell disease, it develops antibodies against erythrocyte antigens, especially against the Rh system antigens, more frequently against RhD antigen, as this presents allelic variations in the RHD gene, resulting in structural changes in the protein, contributing to alloimmunization these individuals. However the frequency of these variants is not known in patients with sickle cell disease in the Brazilian Amazon.Objective: In this study, our goal was to characterize the RhD variants and determine the relevance Transfusion in patients with Sickle Cell Disease in the State of Amazonas. Materials and Methods:We used 96 blood samples from patient's sickle, which were submitted to the RhD phenotype. Identification of RHD variant genotyping was performed by the microarray technique.Results: A total of 96 samples, 36 (37.5%) presented discrepant results in serology for the detection of RhD antigen, and 12 (33.3%) of these, characterized as variant RhD. Among the 12 samples molecularly characterized as variant RhD, 4 (33.33%) DF5, followed by 3 (25,00%) DIIIa, 2 (16.66%) DAU3, 1 (8.33%) DHMI, 1 (8.33%) DFV and 1 (8.33%) DAR. However, the total of 96 (100%) samples, two showed no exon 7 and the other did not show the intron 4 for multiplex PCR, however in serological tests showed normal results and the method of micro arrangements did not present RHD changes.Discussion: This study is of great importance because it is the first results of detection of RhD variants in patients with sickle cell disease state of Amazonas, since our population is differentiated from the rest of Brazil. Is important to mention, it was possible to demonstrate a higher frequency of DF5 variant in our sickle patients, this also contrasts with the world literature, since DF5 variant is more frequent in Caucasians. Conclusion:We emphasize the importance of characterization of RhD variants, in this population, as we can demonstrate significant results and that had not been described in the literature, making it clear techniques involving molecular biology increase transfusion safety and guarantee reliable results for the detection of erythrocyte antigens variants.

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A flexible array format for large‐scale, rapid blood group DNA typing

Abstract: It has been shown that microarray technology can be used to type DNA and detect new alleles in donor cohorts.

Cited by 146 publications

References 20 publications

Molecular biology of the Rh system: clinical considerations for transfusion in sickle cell disease

Molecular biology of the Rh system: clinical considerations for transfusion in sickle cell disease

Bacterial Identification by 16S rRNA Gene PCR-Hybridization as a Supplement to Negative Culture Results

Study of Variations in RhD Holders of Sickle Cell Disease Patients of Amazon State, Brazil

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