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Article | 09-November-2020

Clinical significance of an anti-Dib assessed by flow cytometry

Although antibodies to the Dib antigen are generally considered to be of potential clinical significance, we know of no reports assessing the clinical significance of anti-Dib (in vivo or in vitro). We report on an 88-year-old Japanese male gastrectomy patient who had alloanti-Dib. After transfusion of two Di(b–) units, three Di(b+) units had to be transfused, and there were no clinical signs of acute hemolysis. Di(b+) RBC survival was followed retrospectively by flow cytometry. On days 1

Regina M. Leger, Patricia A. Arndt, Asuncion Co, Lauren O’Brien, George Garratty

Immunohematology, Volume 13 , ISSUE 3, 93–96

Report | 26-October-2019

Proposed criterion for distinguishing ABO mosaics from ABO chimeras using flow cytometric analysis

Differentiation of ABO mosaics from chimeras is performed using flow cytometry (FCM) analysis. Although mosaics and chimeras have been distinguished by presence or absence of clear resolution using FCM analysis, the lack of quantitative metrics and definitive criteria for this differentiation has made some cases difficult to differentiate. In this study, therefore, we attempted to establish a definitive and quantitative criterion for this differentiation. When FCM histogram gates for group

Akira Oda, Nobuki Matsuyama, Mizuko Hirashima, Hiroyuki Ishii, Keiko Kimura, Harumichi Matsukura, Fumiya Hirayama, Keisei Kawa, Yasuo Fukumori

Immunohematology, Volume 31 , ISSUE 1, 24–28

Article | 14-October-2020

Detection of granulocyte antibodies by flow cytometry without the use of pure granulocyte isolates

Karen M. Kiekhaefer, Karen M. Cipolone, Jo L. Procter, Kazuhiko Matsuo, David F. Stroncek

Immunohematology, Volume 17 , ISSUE 3, 70–75

Article | 20-April-2020

Expression of Duffy antigen receptor for chemokines during reticulocyte maturation:using a CD71 flow cytometric technique to identify reticulocytes

Flow cytometric methods commonly used to identify reticulocytes are of limited usefulness in malarious areas,since RNA staining also detects plasmodia. An important antigen expressed on reticulocytes is Duffy antigen receptor for chemokines (DARC,also known as Fy), the receptor for Plasmodium vivax. An early marker for reticulocytes is CD71 (transferrin receptor). We have been interested in CD71 as an alternative marker for reticulocytes in the context of Fy expression. Flow cytometry was used

Ian J. Woolley, Erica M. Wood, R. Michael Sramkoski, Peter A. Zimmerman, John P. Miller, James W. Kazura

Immunohematology, Volume 21 , ISSUE 1, 15–20

Report | 26-October-2019

First example of an FY*01 allele associated with weakened expression of Fya on red blood cells

from the proband’s parents indicated that the father had the same FY genotype as the fetus. Flow cytometry, which has been previously demonstrated as a useful method to study antigen strength on cells, was used to determine if this new FY*01 allele was associated with reduced Fya expression on the father’s RBCs. Median fluorescence intensity of the father’s RBCs (after incubation with anti-Fya and fluorescein-labeled anti-IgG) was similar to known FY*01 heterozygotes and

Patricia A. Arndt, Trina Horn, Jessica A Keller, Rochelle Young, Suzanne M. Heri, Margaret A. Keller

Immunohematology, Volume 31 , ISSUE 3, 103–107

Review | 14-March-2020

Laboratory methods for Rh immunoprophylaxis: a review

for detecting a fetomaternal hemorrhage in D+ mothers or when the D type of the fetus or newborn is D– or unknown. The acid-elution (Kleihauer- Betke) assay is a sensitive laboratory method for quantifying a fetomaternal hemorrhage, but it is tedious, often inaccurate, and difficult to reproduce. Flow cytometry, using anti-D or anti- hemoglobin F reagents, offers a more precise quantification of fetal RBCs in maternal blood. However, flow cytometry services for this function are available in

S. Gerald Sandler, Srividya Sathiyamoorthy

Immunohematology, Volume 26 , ISSUE 3, 92–103

Case report | 09-November-2020

Quantitating fetomaternal hemorrhages of D+ red cells using an FITC-conjugated IgG monoclonal anti-D by flow cytometry: a case report

Several methods for quantitating fetomaternal hemorrhages (FMHs) have been described; these include the Kleihauer-Betke and red cell rosetting tests, and flow cytometry that uses an indirect antiglobulin technique, employing either FITC-conjugated IgG/unlabeled anti-D or streptavidin conjugates with biotinylated anti-D to enumerate D+ red cells in maternal blood. We have used a recently described directly conjugated FITC anti-D for direct flow cytometric (direct FC) quantitation of FMH in a

Anatole Lubenko, John Raymond Collier, Mark Williams, Damien Hindmarch, Sally Rosemary Wilson, Julie Pluck

Immunohematology, Volume 13 , ISSUE 1, 12–14

Article | 14-October-2020

A gel microtyping system for diagnosis of paroxysmal nocturnal hemoglobinuria

lysis (MIRL [CD59]). We evaluated the diagnostic value of a simple hemagglutination test using the gel microtyping system by comparing it with lytic tests (the Ham test and the sucrose lysis test) and with flow cytometry (FC) assessment of expression of GPI-anchored proteins (CD59 and CD55). Examining 51 blood samples from 48 patients, we found that the gel test is useful as a screening test for PNH diagnosis and can replace the Ham test and the sucrose lysis test. The threshold of the gel test is

Barbara Zupanska, Irena Bogdanik, Hanna Pyl

Immunohematology, Volume 18 , ISSUE 1, 9–12

Report | 20-March-2020

Characterization of three novel monoclonal anti-Oka

response, and the spleen B lymphocytes were fused with mouse myeloma X63-Ag8.653 cells to form antibodysecreting hybridomas. The resulting Mabs were tested serologically, by flow cytometry, and by immunoblotting. The specificity of each antibody was determined after excluding specificities to common antigens in the Rh, Kell, Duffy, Kidd, MNS, Lewis, Lutheran, P1, Colton, Diego, Xga, and Dombrock blood group systems. In each case only the Ok(a–) RBC sample was nonreactive. The Mabs and the

Mary H. Tian, Gregory R. Halverson

Immunohematology, Volume 25 , ISSUE 4, 174–178

Review | 29-December-2020

A review: applications of flow cytometry in immunohematology

Sandra J. Nance

Immunohematology, Volume 4 , ISSUE 3, 49–53

Article | 16-November-2020

A method to detect McLeod phenotype red blood cells

carrier females (obligate heterozygotes) is even more difficult because only a minor subpopulation of RBCs may express the weakened Kell phenotype. RBCs from 12 sets of mother/son or father/daughter pairs were tested by standard hemagglutination tube tests and by flow cytometry using both monoclonal and polyclonal Kell system antibodies. Monoclonal anti-K14 (G10) in tests with RBCs from McLeod males reacted ± by hemagglutination (control cells 2+) and had a median fluorecence of 6–11 by

Ragnhild Øyen, Marion E. Reid, Pablo Rubinstein, Harold Ralph

Immunohematology, Volume 12 , ISSUE 4, 160–163

Article | 03-November-2020

Warm autoimmune hemolytic anemia associated with an IgM autoanti-Ge

A 28-year-old male with a prior history of Hodgkin’s disease and a recent upper respiratory tract infection presented with autoimmune hemolytic anemia (AIHA). The patient’s red blood cells (RBCs) were spontaneously agglutinated after room temperature and 37°C washes. Dithiothreitol-treated RBCs reacted strongly with anti-C3 and were nonreactive with anti-IgG, -IgM, and -IgA; they reacted with anti-IgM (κ light chains only) by flow cytometry. The patient’s serum was

Thom S. Sererat, Douglas W. Veidt, Patricia A. Arndt, George Garratty

Immunohematology, Volume 14 , ISSUE 1, 26–29

Article | 17-November-2020

Quantifying the loss of ABO antigenicity in a patient with acute myeloid leukemia by flow cytometric analysis

The loss of B antigenicity from the red blood cells of a patient with acute myeloid leukemia is reported. The patient had normal B transferase levels, but had reduced levels of H transferase. Flow cytometry was used to quantify the loss of B antigenicity and monitor the expression of the B antigen throughout the progression of the disease.

Hazel J. Popp, Margaret Nelson, Cecily Forsyth, Mark Falson, Harry Kronenberg

Immunohematology, Volume 11 , ISSUE 1, 5–7

Article | 16-October-2019

Assessment of common red blood cell pretreatments to yield an accurate serologic antigen phenotype compared with genotype-predicted phenotype

)-approved PreciseType Molecular BeadChip9 (Immucor, Norcross, GA) can predict antigen status for many of the major clinically significant blood group systems (Table 1). Especially in patients with hemoglobinopathies, genotyping is a routine approach to obtaining an extended RBC phenotype.3,5,10 It has been previously documented that genotyping can provide more accurate RBC phenotype results than routine serology.11 A small study was performed to measure, by tube and flow cytometry, the effectiveness of

T. Horn, J. Hamilton, J. Kosanke, V.W. Hare, W. Kluver, W. Beres, S. Nance, M.A. Keller

Immunohematology, Volume 33 , ISSUE 4, 147–151

Article | 09-November-2020

A clinically significant anti-HLA-A2 detectable by extended incubation cytotoxicity and flow cytometric techniques but not by a standard NIH lymphocytotoxicity test

National Institutes of Health (NIH) lymphocytotoxicity test, but antibodies were not detected. However, an extended incubation cytotoxicity test demonstrated the presence of an anti-HLA-A2, and indirect immunofluorescence flow cytometry showed the presence of an IgG1 antibody reacting with 50 percent of cells in a random pool of lymphocytes. One week later, multispecific HLA antibodies were detectable by both NIH and extended incubation cytotoxicity tests. Flow cytometry showed a 16-fold increase in

Stephen F. Garner, John Petrochilos, Colin J. Brown, Suzette Cavanna, I. Chanarin, Cristina Navarrete

Immunohematology, Volume 13 , ISSUE 2, 49–53

Article | 17-November-2020

Loss of the Knops blood group system antigens from stored blood

immunoassay and by flow cytometry measurements. The CR1 expression polymorphism was ascertained by a Hind III restriction fragment length polymorphism analysis. RBC membrane proteins were separated by SDS-PAGE and analyzed for CR1 size by immunoblotting. During the 35-day study interval, no notable decrease was found in RBC-CR1 by flow cytometry. However, CR1 protein was shown to be lost by proteolytic cleavage, as well as by vesiculation. This CR1 protein loss may contribute to the variability of

Joann M. Moulds, L. Lee Brown, Elizabeth Brukheimer

Immunohematology, Volume 11 , ISSUE 2, 46–50

research-article | 30-November-2019

Lovastatin alters neurotrophin expression in rat hippocampus-derived neural stem cells in vitro

number three. For identification of these cells, immunocytochemical evaluation, and flow cytometry analysis were performed using anti-nestin monoclonal antibody (ab6142; 1:300; Abcam), followed by incubation with a fluorescein isothiocyanate (FITC)-conjugated rabbit anti-mouse antibody 1/300 (Millipore, Billerica, MA, USA, AP307F). Cells were cultured on cover slides and fixed in 3% paraformaldehyde for 20 minutes, followed by a permeabilization step in 100% methanol for 30 min at room temperature

Farzaneh Fakheri, Alireza Abdanipour, Kazem Parivar, Iraj Jafari Anarkooli, Hossein Rastegar

Acta Neurobiologiae Experimentalis, Volume 79 , ISSUE 4, 413–420

Article | 15-February-2021

An update on the CD59 blood group system

. In 2018, another CD59 allele was found14 (Table 2) and was carried in heterozygosity together with the wild-type allele. When testing the propositus’ RBCs by flow cytometry using fluorescence-labeled monoclonal anti-CD59, the authors found the density of CD59 comparable to that of controls and concluded that both wild-type CD59 and the variant CD59 were expressed on RBCs. If this finding holds true, the binding site for the reagent anti-CD59 was conserved in this variant. Any further

C. Weinstock, M. Anliker, I. von Zabern

Immunohematology, Volume 35 , ISSUE 1, 7–8

Review | 20-March-2020

Detection and identification of platelet antibodies and antigens in the clinical laboratory

As a result of the unique functional properties of platelets, morerobust methods were required for detection of antibodies raised against them. Immunofluorescence detection by flow cytometry, solid-phase red cell adherence, and antigen capture ELISAs are some of the current tests that have been developed to meet the challenges of platelet antibody detection and identification and antigen phenotyping. Recently developed protein liquid bead arrays are becoming the next-generation platelet

Brian R. Curtis, Janice G. McFarland

Immunohematology, Volume 25 , ISSUE 3, 125–135

Article | 01-June-2016


This paper presents an optimization of a single cell electrical model, based on Maxwell’s mixture Theory, applied to flow cytometry coupled to impedance spectroscopy. It is based on the discretization of the measurement area into a square reference volume, centered between micro-electrodes, and fixed impedance areas. The first one represents the sensing area, the one impacted by cell presence during measurement, and the second one, all other areas that contribute to global measured impedance

J. Claudel, M. Nadi, O. Elmazria, D. Kourtiche

International Journal on Smart Sensing and Intelligent Systems, Volume 9 , ISSUE 2, 526–536

Report | 09-October-2019

A detailed flow cytometric method for detection of low-level in vivo red blood  cell–bound IgG, IgA, and IgM

Wendy Beres, Geralyn M. Meny, Sandra Nance

Immunohematology, Volume 32 , ISSUE 4, 161–169

Article | 09-November-2020

Frequency of neutrophil-specific antigens among Koreans using the granulocyte indirect immunofluorescence test (GIFT)

NA1, NA2, NB1, NB2, NC1, and NE1 are a group of antigens specifically expressed on neutrophils. Antibodies against neutrophil-specific antigens are involved in alloimmune neonatal neutropenia (ANN), autoimmune neutropenia (AIN), and transfusion-related acute lung injury (TRALI). We investigated the frequencies of NA1, NA2, NB1, and Mart antigens in 105 healthy Korean blood donors (65 males, 40 females) by the granulocyte indirect immunofluorescence test (GIFT) employing flow cytometry. Antigen

Kyou S. Han, Tae H. Um

Immunohematology, Volume 13 , ISSUE 1, 15–16

Article | 01-April-2020

A single base insertion of the 4-α-galactosyltransferase gene led to the deficiency of Gb3 biosynthesis

of Gb3/CD77 antigen on the cell surface was evaluated by flow cytometry and by immunochemical techniques. All individuals with the p phenotype were found to have a single base insertion (A4GALT/insC) at the same nucleotide position. Neither the transfectant cells with a mutant gene (A4GALT/insC) of donor origin or those with a synthesized mutant gene (A4GALT/insC-Mu) expressed Gb3 antigen indicating that the presence of A4GALT/insC diminished the A4GALT enzyme activity. In addition, an allele

Mitsunobu Tanaka, Naoko Yamashita, Junko Takahashi, Fumiya Hirayama, Yoshihiko Tani, Hirotoshi Shibata

Immunohematology, Volume 22 , ISSUE 1, 23–29

Article | 17-November-2020

Application of the proteolytic enzyme papain in routine platelet serology

confirmed by flow cytometry. The reactivity of HLA antibodies with PP was generally enhanced. Inactivation by papain of platelet alloantigens in the HPA-2 and HPA-3 systems, but not in other systems, may assist in resolving mixtures of platelet alloantibodies. Also, detection of weak antibodies of other specificities may be enhanced. The use of PP may be a simple and useful serologic tool for investigating platelet alloantibodies.

John A.G. Lown, Brian J. Dale

Immunohematology, Volume 11 , ISSUE 4, 140–142

Report | 17-March-2020

Southeast Asian ovalocytosis is associated with increased expression of Duffy antigen receptor for chemokines (DARC)

reduced susceptibility to vivax malaria. Blood samples were collected from individuals living in the Madang Province of Papua New Guinea. Samples were assayed using a flow cytometry assay for expression of Fy on the surface of RBC and reticulocytes by measuring the attachment of a phycoerythrin-labeled Fy6 antibody. Reticulocytes were detected using thiazole orange. The presence of the SAO mutation was confirmed by PCR. There was a small (approximately 10%) but statistically significant (p=0.049, Mann

Ian J. Woolley, Paul Hutchinson, John C. Reeder, James W. Kazura, Alfred Cortés

Immunohematology, Volume 25 , ISSUE 2, 63–66

Report | 06-November-2019

Drug-induced immune neutropenia/ agranulocytosis

, sulfamethoxazoletrimethoprim, β-lactam antibiotics, clozapine, levamisole, and vancomycin. Assays used for detection of neutrophil drug-dependent antibodies (DDAbs) include flow cytometry, monoclonal antibody immobilization of granulocyte antigens, enzyme-linked immunosorbent assay, immunoblotting, granulocyte agglutination, and granulocytotoxicity. However, testing for neutrophil DDAbs is rarely performed owing to its complexity and lack of availability. Mechanisms proposed for DIIN have not been rigorously studied

Brian R. Curtis

Immunohematology, Volume 30 , ISSUE 2, 95–101

Article | 22-November-2020

Development of a flow cytometric test for the detection of D-positive fetal cells after fetomaternal hemorrhage and a survey of the prevalence in D-negative women

fetomaternal hemorrhage (FMH) of this volume is detectable in the maternal circulation as approximately 0.25 percent of the total RBCs. Our test utilizes a commercially available human monoclonal IgG anti-D that has been biotinylated and used with a dye-conjugated streptavidin. Flow cytometry is used to quantitate fluorescing D-positive RBCs. To date, 2,288 tests have been performed on blood samples from D-negative women attending local antenatal clinics or at the time of delivery. Evidence for an FMH has

Margaret Nelson, Hazel Popp, Kathy Horky, Cecily Forsyth, John Gibson

Immunohematology, Volume 10 , ISSUE 2, 55–59

original-paper | 22-October-2019

Salmonella-Infected Aortic Aneurysm: Investigating Pathogenesis Using Salmonella Serotypes

CD36 expression. To detect cell surface expression of CD36, flow cytometric analysis was performed using monoclonal FITC-conjugated anti-CD36 antibody (Abcam, ab82443). The THP-1-derived macrophages were incubated with the aforementioned antibody for 40 min in a dark room and washed three times with chilled phosphate-buffered saline (PBS) containing 0.02% NaN3. The cells were analyzed using flow cytometry. Salmonella infection. Each single Salmonella colony was inoculated in 5 ml of LB broth at 37


Polish Journal of Microbiology, Volume 68 , ISSUE 4, 439–447

original-paper | 31-October-2019

Cytokine Levels in the In Vitro Response of T Cells to Planktonic and Biofilm Corynebacterium amycolatum

three times in the wells of 24-well culture plates (Nunc, USA) and incubated with the bacterial conditioned media (PCM, BCM, PCMmix or BCMmix) for 24 h at 37°C and 5% CO2 at a ratio of 1:10 (total volume – 1000 µl). After incubation, the supernatants of the cell culture were collected and analyzed for cytokine concentrations. Detection of cytokines by flow cytometry. The level of cytokines in the supernatants of Jurkat T cell culture after their exposure to the PCM, BCM, PCM-mix, and BCMmix was


Polish Journal of Microbiology, Volume 68 , ISSUE 4, 457–464

Report | 06-November-2019

Drug-induced immune thrombocytopenia: incidence, clinical features, laboratory testing, and pathogenic mechanisms

candidate drug resulted in recurrent thrombocytopenia. Flow cytometry testing for DDAbs can be useful in confirmation of a clinical diagnosis, and monoclonal antibody enzyme-linked immunosorbent assay testing can be used to determine the platelet glycoprotein target(s), usually GPIIb/IIIa or GPIb/IX/V, but testing is not widely available. Several pathogenic mechanisms for DIIT have been proposed, including hapten, autoantibody, neoepitope, drug-specific, and quinine-type drug mechanisms. A recent

Brian R. Curtis

Immunohematology, Volume 30 , ISSUE 2, 55–65

Report | 01-December-2019

Comparison of estimation of volume of fetomaternal hemorrhage using KleihauerBetke test and microcolumn gel method in D-negative nonisoimmunized mothers

7.9–10.4 mL, p < 0.001). Microcolumn gel method is an effective screening test. Technologies like KB and flow cytometry are better options for detecting a large volume of FMH. Antepartum hemorrhage and cesarean delivery are risk factors for FMH. The 300-µg dose appears to be excessive immunoprophylaxis in the majority of cases. We need to analyze the relative cost-effectiveness of universal administration of 300 µg of Rh immune globulin vs. FMH quantitation with subsequent

Kshitija Mittal, Neelam Marwaha, Praveen Kumar, Subhash C. Saha, Beenu Thakral

Immunohematology, Volume 29 , ISSUE 3, 105–109

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