Semiautomated data analysis of flow cytometric estimation of fetomaternal hemorrhage in D− women

Greiss, M. Ayoub; Armstrong-Fisher, S.S.; Perera, W.S.; Brown, Peter; Urbaniak, S. J.

doi:10.1046/j.1537-2995.2002.00159.x

Cited by 8 publications

(4 citation statements)

References 23 publications

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“…The evaluation of the performances of the Fetal Cell Count kit demonstrates a good linearity and precision of the method for artificial mixtures ranging from 0.01 to 5.00 percent fetal cells representative of more than 99.9 percent of the clinical situation with FMH 1 . These data are very close to those obtained by other investigators performing flow cytometry with a single marker 17,38,40,41 . They confirm that flow cytometry is more precise than the KBT known to have a poor reproducibility 4,42 despite several attempts to standardize the method 43 and the introduction of external quality assurance scheme for Kleihauer testing 44 .…”

Section: Discussionsupporting

confidence: 75%

“…1 These data are very close to those obtained by other investigators performing flow cytometry with a single marker. 17,38,40,41 They confirm that flow cytometry is more precise than the KBT known to have a poor reproducibility 4,42 despite several attempts to standardize the method 43 and the introduction of external quality assurance scheme for Kleihauer testing. 44 Two major reasons may explain these findings.…”

Section: Discussionmentioning

confidence: 87%

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Identification and quantification of fetal red blood cells in maternal blood by a dual‐color flow cytometric method: evaluation of the Fetal Cell Count kit

Porra¹,

Bernaud²,

Guéret³

et al. 2007

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Section: Discussionsupporting

confidence: 75%

Section: Discussionmentioning

confidence: 87%

Identification and quantification of fetal red blood cells in maternal blood by a dual‐color flow cytometric method: evaluation of the Fetal Cell Count kit

Porra¹,

Bernaud²,

Guéret³

et al. 2007

Transfusion

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“…After repeating this washing step three times, 50 μL monoclonal FITC‐conjugated anti‐D (LDG76) antibody (Quant‐Rho, Quotient Biodiagnostics, Newtown, PA) was added, and the solution was mixed thoroughly by vortexing. This antibody is an IgG1κ heterohybrid (human‐murine) monoclonal antibody specific for epD3 in the nine epitope model and epD5 in the thirty‐six epitope model of D antigen and has been shown to have a high affinity for D antigen, achieving sufficiently high antibody‐to‐cell ratio [20]. The volume of antibody was determined in pilot experiments to ensure antibody saturation (data not shown).…”

Section: Methodsmentioning

confidence: 99%

A novel laboratory technique demonstrating the influences of RHD zygosity and the RhCcEe phenotype on erythrocyte D antigen expression

et al. 2011

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D antigen is the most immunogenic and clinically relevant antigen within the complex Rh blood group system. Variability of D antigen expression was first described decades ago but has rarely been investigated quantitatively, particularly in the context of RHD zygosity along with RhCcEe serological phenotype. With IRB approval, 107 deidentified blood samples were analyzed. Rh phenotypes were determined serologically by saline technique using monoclonal antibodies against D, C, c, E, and e antigens. RHD zygosity was determined using both PCR-restriction fragment length polymorphisms and quantitative real-time PCR techniques. A novel and robust method was developed for quantitation of erythrocyte D antigen sites using calibrated microspheres and flow cytometry, allowing correlation of D antigen density with RHD zygosity and expression of Rh CcEe antigens. Subjects homozygous for RHD expressed nearly twice the number of D antigen sites compared with RHD hemizygotes (33,560 ± 8,222 for DD versus 17,720 ± 4,471 for Dd, P < 0.0001). Expression of c or E antigens was associated with significantly increased erythrocyte D antigen expression, whereas presence of C or e antigens reduced expression. These data and this novel quantitation method will be important for future studies investigating the clinical relevance of D antigen variability.

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“…Initial FC assays targeted the D antigen on fetal RBCs, distinguishing D+ fetal RBCs from D-maternal RBCs using polyclonal or monoclonal anti-D reagents. 46,[67][68][69][70][71][72][73][74] Nance and colleagues 67 compared the results of testing mixtures of D+ cord RBCs and D-adult RBCs by the rosette screen, K-B acid elution assay, and an FC (anti-D) assay. They reported that results by FC (anti-D) were more accurate, reproducible, and sensitive.…”

Section: Flow Cytometry Anti-d Methodsmentioning

confidence: 99%

Laboratory methods for Rh immunoprophylaxis: a review

Sandler¹,

Sathiyamoorthy

2010

Immunohematology

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The recommended dose of Rh immune globulin for postpartum Rh immunoprophylaxis is based on an estimation of the volume of the fetomaternal hemorrhage, if any, measured as the percent of fetal RBCs in a sample of the D-mother's blood. Laboratory methods for distinguishing fetal from maternal RBCs have been based on their different blood types (D+ versus D-) or predominant hemoglobin content (hemoglobin F versus hemoglobin A). We conducted a review of the medical literature describing laboratory methods for detecting and quantifying fetal RBCs in maternal blood samples. We also used data collected for the College of American Pathologists Fetal RBC Detection Surveys to determine which laboratory methods are used currently in hospitals in the United States. The rosette screen is used widely for identifying D-mothers who may require additional doses of Rh immune globulin for postpartum immunoprophylaxis. As the rosette screen targets the D antigen, it is not suitable 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 antihemoglobin F reagents, offers a more precise quantification of fetal RBCs in maternal blood. However, flow cytometry services for this function are available in relatively few hospital laboratories in the United States because of logistic and fiscal impediments.

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Semiautomated data analysis of flow cytometric estimation of fetomaternal hemorrhage in D− women

Abstract: Semiautomated data acquisition and calibration curve analysis represents a further step toward standardization of flow cytometry for accurate FMH quantification and facilitates evaluation and control of day-to-day variations between laboratories, flow cytometers, and operators.

Cited by 8 publications

References 23 publications

Identification and quantification of fetal red blood cells in maternal blood by a dual‐color flow cytometric method: evaluation of the Fetal Cell Count kit

Identification and quantification of fetal red blood cells in maternal blood by a dual‐color flow cytometric method: evaluation of the Fetal Cell Count kit

A novel laboratory technique demonstrating the influences of RHD zygosity and the RhCcEe phenotype on erythrocyte D antigen expression

Laboratory methods for Rh immunoprophylaxis: a review

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