A Critical Review of Published Methods for Analysis of Red Cell Antigen-Antibody Reactions by Flow Cytometry, and Approaches for Resolving Problems with Red Cell Agglutination

Arndt, Patricia A.; Garratty, George

doi:10.1016/j.tmrv.2010.03.001

Cited by 20 publications

(24 citation statements)

References 23 publications

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“…In the absence of alloantibodies, post-transfusion RCS is reliably determined using mismatched RBC antigens 19, 20 . Thus, tracking of RCS based on mismatched Kidd RBC Jka and Jkb in the present study reinforces the conclusion that low density BioRBC determinations accurately reflect RCS, i.e., that ex vivo biotinylation does not artifactually shorten RCS 9, 10 .…”

Section: Discussionmentioning

confidence: 99%

Autologous Infant and Allogeneic Adult Red Cells Demonstrate Similar Concurrent Post-Transfusion Survival in Very Low Birth Weight Neonates

Widness

Kuruvilla

Mock

et al. 2015

The Journal of Pediatrics

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Objective Based on the hypothesis that neonatal autologous red blood cell (RBC) survival (RCS) is substantially shorter than adult RBC, we concurrently tracked the survival of transfused biotin-labeled autologous neonatal and allogeneic adult RBC into ventilated, very low birth weight (VLBW) infants. Study design RBC aliquots from the first clinically ordered, allogeneic adult RBC transfusion and from autologous infant blood were labeled at separate biotin densities (BioRBC) and transfused. Survival of these BioRBCs populations were concurrently followed over weeks by flow cytometric enumeration using leftover blood. Relative tracking of infant autologous and adult allogeneic BioRBC was analyzed by linear mixed modeling of batched weekly data. When possible, Kidd antigen (Jka and Jkb) mismatches between infant and donor RBCs were also used to track these two populations. Results Contrary to our hypothesis, concurrent tracking curves of RCS of neonatal and adult BioRBC in 15 study infants did not differ until week 7, after which neonatal RBC survival became shortened to 59% to 79% of adult enumeration values for uncertain reasons. Analysis of mismatched Kidd antigen RBC showed similar results, thus confirming that BioRBC tracking is not perturbed by biotin RBC labeling. Conclusions This study illustrates the utility of multi-density BioRBC labeling for concurrent measurement of RCS of multiple RBC populations in vivo. The similar RCS results observed for neonatal and adult BioRBCs transfused into VLBW infants provides strong evidence that the circulatory environment of the newborn infant—not intrinsic infant-adult RBC differences—is the primary determinant of erythrocyte survival. Trial registration Clinicaltrials.gov: NCT 00731588.

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Section: Discussionmentioning

confidence: 99%

Autologous Infant and Allogeneic Adult Red Cells Demonstrate Similar Concurrent Post-Transfusion Survival in Very Low Birth Weight Neonates

Widness

Kuruvilla

Mock

et al. 2015

The Journal of Pediatrics

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“…Based on this we used clones that greatly reduce charge masking and reduce aggregate formation. In addition, we used methods that aggregate formation, particularly when staining RBC for flow cytometry, such as: titering down antibodies, using directly conjugated antibodies, staining at room temperature, logarithmic scaling for forward scatter, side scatter, thorough vortexing and mixing of samples, and fixing samples with formaldehyde to reduce agglutination [51]. Together, these procedures significantly reduced aggregate formation for the phenotyping of RBC and PLT blood product.…”

Section: Methodsmentioning

confidence: 99%

Platelets and Erythrocyte-Bound Platelets Bind Infectious HIV-1 in Plasma of Chronically Infected Patients

et al. 2013

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Chronic HIV-1 infection is associated with persistent viremia in most patients, but it remains unclear how free virus may survive the potential hostile effects of plasma. We investigated whether sites might exist on the surfaces of circulating blood cells for protection of infectious HIV-1 particles. Red blood cells (RBC) either from blood of uninfected normal individuals, or from blood obtained without EDTA from chronically infected HIV-1 patients, invariably contained a small number of RBC having attached platelets as determined by flow cytometry, light microscopy, and immunofluorescence microscopy. After mixing normal RBC with platelet-rich plasma, discrete populations of RBC, platelets, and complexes of platelets attached to RBC were purified by fluorescence-activated cell sorting. Upon incubation of purified cells or platelets with HIV-1 followed by washing and co-incubation with CD4-positive peripheral blood mononuclear cells (PBMC), platelets, and platelet-RBC complexes, but not platelet-free RBC, caused infection of PBMC. Infection was prevented by pre-treating the platelet-RBC complexes with EDTA. Plasma and RBC (comprising a RBC/platelet-RBC mixture) from chronically infected patients with low viral loads were also co-incubated with PBMC ex vivo to determine the presence of infectious HIV-1. All freshly isolated plasmas from the HIV-1-infected donors, obtained in the absence of anticoagulant, were noninfectious. Interestingly, the RBC from most of the patients caused cell-cell infection of PBMC that was prevented by stripping the RBC with EDTA. A monoclonal antibody to DC-SIGN partially inhibited cell-cell HIV-1 infection of PBMC by normal RBC pre-incubated with platelets and HIV-1. We conclude: (a) platelet-free EDTA-free plasma from chronically infected HIV-1 patients, although containing viral RNA, is an environment that lacks detectable infectious HIV-1; (b) platelets and platelet-RBC complexes, but not purified RBC, bind infectious HIV-1; (c) DC-SIGN, and possibly other C-type lectins, may represent binding sites for infectious HIV-1 on platelets and platelet-RBC complexes.

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“…In this test, we opted to choose a cutoff with a high sensitivity to ensure the test will correctly identify 100% of all patients, as there are few testing options available to immune‐mediated DAT(–) patients. A few groups have used flow cytometry to diagnose AIHA in both DAT(–) and DAT‐positive patients . A recent study by Fayek and colleagues established a fluorescence cutoff with a sensitivity and specificity of 100%.…”

Section: Discussionmentioning

confidence: 99%

Western immunoblotting as a new tool for investigating direct antiglobulin test–negative autoimmune hemolytic anemias

et al. 2015

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WB analysis is more sensitive than conventional test tube DAT or elution analysis. Our assay confirms: 1) previous studies showing normal RBCs are sensitized with IgG, perhaps due to natural autoantibody to senescence; 2) that some normal RBCs have increased levels of IgG without signs of disease; and 3) that WB distinguishes between non-immune- and immune-mediated hemolytic anemia in DAT(-) patients and may be useful for clinical diagnosis.

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A Critical Review of Published Methods for Analysis of Red Cell Antigen-Antibody Reactions by Flow Cytometry, and Approaches for Resolving Problems with Red Cell Agglutination

Cited by 20 publications

References 23 publications

Autologous Infant and Allogeneic Adult Red Cells Demonstrate Similar Concurrent Post-Transfusion Survival in Very Low Birth Weight Neonates

Autologous Infant and Allogeneic Adult Red Cells Demonstrate Similar Concurrent Post-Transfusion Survival in Very Low Birth Weight Neonates

Platelets and Erythrocyte-Bound Platelets Bind Infectious HIV-1 in Plasma of Chronically Infected Patients

Western immunoblotting as a new tool for investigating direct antiglobulin test–negative autoimmune hemolytic anemias

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