bluuebunny/crossref_metadata_2025_split huggingface dataset:
Evaluation of a flow cytometric fluorescence quenching assay of phagocytosis of sensitized sheep erythrocytes by polymorphonuclear leukocytes
Authors: Van Amersfoort Edwin S. , Van Strijp Jos A. G. 
Published: June 2005

AbstractA number of reports have been published describing phagocytosis assays for flow cytometric analysis. In some of these, the fluorescence quenching technique has been used to discriminate between adherent and ingested particles. In this report, we have evaluated the efficacy of a quantitative fluorescence quenching technique with crystal violet and trypan blue for application in a phagocytosis assay with polymorphonuclear leukocytes and sensitized sheep red blood cells. We set the requirements to a high quenching efficiency of the fluorescence of extracellularly bound particles and no intracellular quenching. The later was determined using polymorphonuclear leukocytes stained with the fluorescent nuclear dye hydroethidine. We observed that both trypan blue and crystal violet efficiently quench the fluorescence of PKH26 (a red fluorescent membrane‐associated dye) erythrocytes but that only crystal violet quenches intracellular fluorescence. In testing trypan blue and crystal violet from different manufacturers, there was no real difference between different brands of crystal violet, but only the trypan blue from Merck turned out to be an efficient quencher, whereas the other brands of trypan blue showed low quenching efficiency. Trypan blue at a concentration of 25–50 μg/ml proved to be a good quencher of the fluorescent erythrocytes and exerted minimal side effects: over 90% quenching of the erythrocytes, no intracellular quenching, moderate increase in autofluorescence of the polymorphonuclear leukocytes, and no cell loss. © 1994 Wiley‐Liss, Inc.
Detection of bromodeoxyuridine incorporation by alteration of the fluorescence emission from nucleic acid binding dyes using only an argon ion laser
Authors: Frey Tom 
Published: June 2005

AbstractA method was developed that uses paired DNA dyes to detect the incorporation of bromodeoxyuridine (BrdUrd) into cellular DNA and requires only 488 nm excitation. The flurescence of thiazole blue, TO‐PRO‐3 and LDS‐751 was found to be enhanced by the presence of Brd Urd in DNA. Pairing LDS‐751, thiazole blue, or TO‐PRO‐3 with propidium iodide (PI) for flow cytometry allowed the differentiation of cells containing BrdUrd from BrdUrd unlabeled cells. LDS‐751 can be excited directly at 488 nm, and TO‐PRO‐3 or thiazole blue can be excited indirectly by resonance energy transfer from PI. The enhancement of fluorescence from these dyes is correlated with a decrease in PI fluorescence, suggesting an increased energy transfer from PI to the red emitting dye. Flurescence from other dyes, including thiazole orange, TO‐PRO‐1, rhodamine 800, oxazine 750, and 7‐aminoactinomycin D, was not altered by the presence of BrdUrd in DNA. Results also were obtained showing that BrdUrd detection using PI and TO‐PRO‐3 is compatible with immunofluorescence staining with FITC‐labeled antibodies. © 1994 Wiley‐Liss, Inc.
Carcinoma in situ of the testis detected by DNA flow cytometry of testicular fine‐needle aspirates
Authors: Hittmair Anton , Rogtsch Hermann , Feichtinger Hans , Hobisch Alfred , Mikuz Gregor 
Published: June 2005

AbstractTesticular carcinoma in situ (CIS) is usually diagnosed histologically on surgical biopsies. The present study was performed to test an alternative approch—DNA flow cytometry (FCM) of testicular fine‐needle aspirates (FNA)—for the detection of this lesion. FNAs from 18 cases of testicular germ cell tumors (TGCT) and tumor adjacent parenchyma were analysed by DNA FCM. DNA histograms of all cell nuclei and histograms representing selectively the hyperdiploid compatment were analysed. The presence and extention of CIS were determined by histology and immunohistochemistry. In 16 to 18 cases, CIS was histologically present, whereas aneuploid peaks were detected in only 11 cases in DNA histograms of all cell nuclei. In the analysis of the histograms of the hyperdiploid region, 4 additional cases of CIS could be identified increasing the sensitivity to 15/16 cases (93.8%). In all but one case, the DNA indices of CIS and invasive tumors were identical. The study demonstrates that DNA FCM of testicular FNAs using the described data acquisition and analysis could also be applicable for detection of CIS in a clinical situation. © 1994 Wiley‐Liss, Inc.
An optimized method for routine HLA‐B27 screening using flow cytometry
Authors: Hulstaert F. , Albrecht J. , Hannet I. , Lancaster P. , Buchner L. , Kunz J. , Falkenrodt A. , Tongio M. , de Keyser F. , Veys E. M. , Noens L. , Mir N. , Costello C. , Becker R. , Strauss K. 
Published: June 2005

AbstractFlow cytometry and monoclonal antibodies are promising tools for HLA‐antigen detection. Previous approaches have been hampered by the lack of a carefully standardized system for calibration and sample analysis. A new system for HLA‐B27 screening was developed using a FACScan flow cytometer, software for automated calibration and analysis, calibration beads, and the anti‐HLA‐B27‐FITC/anti‐Leu4‐PE (CD3) monoclonal antibodies. The median fluorescence channel result for the HLA‐B27‐FITC signal of CD3+ T lymphocytes is compared to a decision marker. Values lower than this threshold are read as HLA‐B27 negative and those above are recommended for retesting with the classic microcytotoxicity assay on the presumption of HLA‐B27 positivity. The anti‐HLA‐B27 antibody reacts with all six HLA‐B27 subtypes and shows a weaker binding to HLA‐B7. The screening test results were compared with those from the microcytotoxicity assay for HLA‐typing in studies involving several European centers. The observed sensitivity was 100% (95% Cl: 98.6–100) and the specificity was 97.4% (95% Cl: 96.4–98.3). Other performance studies verified the reproducibility and reliability of results obtained with the screening system. © 1994 Wiley‐Liss, Inc.
Two color flow cytometric analysis of concomitant acute myeloid leukemia and chronic lymphocytic leukemia
Authors: Tamul Karen R. , Meyers Dawn C. , Bentley Stuart A. , Folds James D. 
Published: June 2005

AbstractDuring evaluation for macrocytic anemia and thrombocytopenia, a 74‐year‐old female was found to have a leukocytosis with two apparent populations of malignant cells identified in her peripheral blood smear and bone marrow aspirate. Morphologic characteristics of the two cell types were unusual, and cytochemical assays yielded ambiguous results. Two‐color flow cytometric analysis demonstrated two distinct cell populations with immunophenotyping patterns consistent with chronic lymphocytic leukemia (CD5 + /CD20 +) and acute myelocytic leukemia (CD33 + /CD34 +), detected concurrently. The concomitant appearance of CLL and AML has been reported only rarely. The use of two‐color flow cytometry to differentiate the populations demonstrates the utility of this technology in resolving unusual hematological malignancies. © 1994 Wiley‐Liss, Inc.
Phenotypic and functional abnormalities in the peripheral blood T‐cells of patients with primary Sjogren's syndrome
Authors: Aziz Karim Elias , McCluskey Peter J. , Wakefield Denis 
Published: June 2005

AbstractChanges in regulatory T‐cell subset (including the recently described CD4 helper inducers or suppressor inducers) balance in the peripheral blood may play a role in the pathogenesis of primary Sjogren's syndrome (SS). Direct immunofluorescence and flow cytometry were used to quantitate and analyse peripheral blood lymphocytes in 15 patients with primary SS and 15 control subjects. A reduction in the percentage of circulating CD4 lymphocytes was observed in patients with SS. There was no quantitative abnormality in the percentage of circulating CD4+ 2H4+ (suppressor inducer), CD4+ 4B4+ (helper inducer), CD2, CD3, CD8, CD8+ 2H4+, CD8+ 4B4+, CD25 (IL‐2R), CD19, CD16, CD57 lymphocytes in the patients. Circulating CD8 lymphocytes expressing the activation marker HLA‐DR were increased in the patients. The functional status of peripheral blood lymphocytes was assessed by PHA (phytohaemagglutinin) stimulation followed by monitoring their proliferative response by radiolabelled thymidine uptake and expression of CD25 (Inter‐leukin‐2 receptor). A reduction in the proliferative response of total, CD4‐depleted, and CD8‐depleted lymphocytes suspensions to PHA was demonstrated. The level of expression of CD25 (IL‐2 receptor) was similar in patients and controls before and after 24 h stimulation with PHA. We conclude that there is a disturbance in the functional properties of peripheral blood T cells that can contribute to the immunopathogenesis of SS. Meanwhile, the quantitative reduction of suppressor/inducer lymphocytes as defined by the CD4 2H4 phenotype can be precluded from a role in the development of such an autoimmune condition. © 1994 Wiley‐Liss, Inc.
Simultaneous expression of CD2 on a B‐cell, small lymphocytic neoplasm
Authors: Viciana Ana L. , Lancet Frederick C. , Chamizo Wilfredo , Voigt Walter , Ruiz Phillip 
Published: June 2005

AbstractAn unusual neoplasm, best classified as a B‐cell chronic lymphocytic leukemia on the basis of cytofluorographic, immunohistologic, and gene rearrangement analysis also co‐expressed the T‐cell associated marker CD2 (sheep erythrocyte receptor), but without other cell markers restricted to T cells. This case was associated with a more aggressive course than typically seen with B‐CLL or malignant lymphoma, small lymphocytic type, implying along with previous reports that CD2 expression may serve as a marker of small lymphocytic tumor cell behavior and the ultimate clinical course. Thus, surface phenotypic analysis of these particular hematopoietic neoplasms may serve not only for identification of the malignancy, but could also render prognostic information. © 1994 Wiley‐Liss, Inc.
Development and clinical evaluation of an amplified flow cytometric fluoroimmunoassay for Clostridium difficile toxin A
Authors: Renner E. D. 
Published: June 2005

AbstractA rapid (2 h) amplified flow cytometric fluoroimmunoassay (AFCF) for Clostridium difficile toxin A was developed and compared with the cytotoxin assay (CTA) and culture of the organism from stool specimens from patients with suspected C. difficile‐associated gastrointestinal disease (CAD). For this assay polyclonal antitoxin A was attached to 10‐μm diameter and monoclonal antitoxin A was attached to fluorescent 0.1 μm‐diameter polystyrene microspheres. The microspheres and sample were reacted together as in a conventional double‐antibody sandwich assay. However, laser flow cytometric measurement allowed the omission of separation and washing steps by gating on light scattered by the larger microspheres and measuring only the fluorescence associated with these particles. The amount of fluorescence from the attached 0.1 μm microspheres was dependent on the concentration of toxin A in the sample. The AFCF detected purified toxin A at levels of 1 pg/ml and was linear from 1 to 40 pg/ml. The AFCF was compared with the CTA and culture of C. difficile for clinical use by comparing results from 198 stool specimens from patients with suspected CAD. The AFCF was 85.7% sensitive and 95.8% specific relative to the CTA, and 85.2% sensitive and 98.3% specific compared to the culture assay. If the isolation of toxigenic C. difficile or the patients clinical course was considered indicative of CAD, the sensitivities of the AFCF, CTA, and culture assay were 77.4%, 67.7% and 96.8%, respectively. The AFCF demonstrated a specificity of 98.8%, while both CTA and culture had a specificity of 100%. The AFCF for toxin A is a sensitive and rapid test, and in laboratories with a flow cytometer offers an alternative method for the laboratory diagnosis of CAD. © 1994 Wiley‐Liss, Inc.
Retrospective evaluation of flow cytometry as a platelet crossmatching procedure
Authors: Gates Karen , MacPherson Bruce R. 
Published: June 2005

AbstractWe performed a retrospective analysis of flow cytometry as a platelet crossmatching procedure. Sera from 17 alloimmunized refractory patients were tested against 32 donor platelets, which had been stored as platelet‐rich plasma for up to 36 months. Overall, 14/32 (44%) crossmatches were positive. The mean 1 h posttransfusion corrected count increments (CCIs) were 9,195 and 2,269 for a negative and a positive crossmatch, respectively. The predictive value of a positive crossmatch was 86%, whereas the predictive value of a negative crossmatch was 56%. When samples with low background fluorescence or with high panel‐reactive antibody (PRA) levels were evaluated separately, the accuracy of the crossmatch improved from 69% to 80%. When compared to the platelet adhesion immunofluorescence test (PAIFT) and the standard and antiglobulin‐enhanced lymphocytotoxicity tests for the detection of HLA antibodies, flow cytometry appeared to be more sensitive. We conclude that flow cytometry is a useful technique for platelet crossmatching, particularly for alloimmunized patients for whom HLA compatible platelets may not be readily available. © 1994 Wiley‐Liss, Inc.
Artifactual staining of monoclonal antibodies in two‐color combinations is due to an immunoglobulin in the serum and plasma
Authors: Nicholson Janet K. A. , Rao Patricia E. , Calvelli Theresa , Stetler‐Stevenson Maryalice , Browning Sandra W. , Yeung Lily , Marti Gerald E. 
Published: June 2005

AbstractTwo‐color whole blood lysis is the assay of choice for lymphocyte immunophenotyping because of the additional information it provides. Recently, artifactual double‐staining of some specimens has been observed with this assay. In these cases, the samples appear to be uncompensated for spectral overlap or to inappropriately coexpress two antigens simultaneously. This artifact can result in the apparent coexpression of CD4 and CD8 (observed in lymphoblastic processes) or of CD5 and CD20 (characteristic of chronic lymphocytic leukemia) in normal persons, leading to an erroneous diagnosis. Using plasma, serum, or immunoglobulin preparations from donors who exhibit this artifact we sought to determine 1) the source of the artifact and 2) ways to overcome it. This staining is apparently due to an immunoglobulin in the donors' serum and plasma which does not have specific reactivity with mouse immunoglobulin. Washing whole blood samples or blocking with mouse immunoglobulin is a convenient way of avoiding this artifact. © 1994 Wiley‐Liss, Inc.
Leukemia‐associated changes identified by quantitative flow cytometry: I. CD10 expression
Authors: Lavabre‐Bertrand Thierry , Janossy George , Ivory Kamal , Peters Rowajda , Secker‐Walker Lorna , Porwit‐MacDonald Anna 
Published: June 2005

AbstractWe have compared CD10 antigen expression in normal fetal bone marrow with that of B‐lineage acute lymphoblastic leukemia (ALL). Both quantitative indirect immunofluoresence (QIFI) and direct immunofluorescence (IF) tests with Quantum beads were used to convert median fluorescence intensity (MFI) values into numbers of antigen molecules expressed per cell (AgE). Lymphoid precursors in the fetal marrow and liver expressed 3–12.5 × 103 CD10 molecules/cell with an upper limit of 5 × 104/cell (MaxAgE). The median CD10 AgE in the different cases of acute B‐lineage ALL were variable and ranged from undetectable to very high values (> 1.8 × 105). In 24 of the 72 cases (33%) tested with QIFI the median CD10 AgE was above the highest values seen in normal samples (> 5 × 104/cell). An additional 23.6% of cases had higher median values than the normal median CD10 AgE. Next, CD10 antigen was quantitated in 78 cases during the routine multiparameter analysis of B‐lineage leukemia using CD10/class II/CD34 3‐color IF test or CD10/TdT 2‐color IF test. The aberrant overexpression was confirmed in 43.6% of ALL cases. The CD10bright display suggested ALL diagnosis even when few cells were available for study, e.g., in early relapse and in ALL masquerading as aplastic anemia. The levels of CD10 expression were maintained in relapse. In addition, different CD10 levels were associated with the various chromosomal alterations: high CD10 levels (> 3 × 104/cell) with hyperdiploidy, low CD10 levels (1.8–4 × 103/cell) with the t(1;19), and undetectable levels (< 1.2 × 103/cell) with the t(4;11) translocations. These findings show that while all these diseases are part of the precursor B‐cell spectrum, the CD10 changes are linked to disease‐associated alterations instead of truly reflecting the features of subtypes of normal B‐cell precursors. The quantitative CD10 assessment should therefore be part of the routine flow cytometric assessment in ALL, and further careful studies are warranted during the regeneration phase following chemotherapy. High CD10 expression alone during strong bone marrow regeneration may not be leukemia specific and both the multiparameter analysis described above and parallel studies for gene rearrangements will be necessary to establish the relative values of these assays in detecting minimal disease amidst regenerating marrow. © 1994 Wiley‐Liss, Inc.
Computerized image analysis and flow cytometric evaluation of ovarian bordeline tumors: A study of 24 cases
Authors: Esposito Michael J. , Fuchs Alexander 
Published: June 2005

AbstractDNA content evaluation in the study of ovarian borderline tumors has been shown to be a useful adjunct to histopathologic diagnosis. This study compares flow cytometry and computerized image analysis (CIA) in evauating the DNA content of these lesions.Twenty‐four cases of ovarian borderline tumors (15 serous, 9 mucinous) were studied utilizing formalinfixed paraffin‐embedded tissue. Flow cytometry of cell suspensions and CIA of cell suspensions and paraffin sections were compared in the evaluation of DNA content.Twenty‐three tumors (96%) were diploid and only 1 (4%) was aneuploid. There was 100% correlation between flow cytometry and CIA of cell suspensions. Image analysis of Feulgen‐stained paraffin sections was found to be unreliable when compared with the use of cell suspensions in the evaluation of DNA content. Clinical follow‐up in 17 patients showed recurrent disease in two patients with diploid tumors and no evidence of disease in the patient with an aneuploid tumor.The reported rate of aneuploidy in borderline tumors varies depending on the diagnostic method employed in evaluating DNA content. This study shows good correlation between flow cytometry and image analysis of cell suspensions and supports the low rate of aneuploidy found in these lesions by other authors utilizing these method. The majority of borderline ovarian tumors have been shown to have a diploid DNA content; however, the finding of aneuploidy in histologically classified borderline tumors warrants close clinical follow‐up since aneuploidy in these tumors has been associated with an adverse prognosis. © 1994 Wiley‐Liss, Inc.
DNA image cytometric analysis of paraffin‐embedded sections of small renal cortical neoplasms
Authors: Farnsworth William V. , Derose Patricia B. , Cohen Cynthia 
Published: June 2005

AbstractDNA ploidy analysis of neoplasms is most frequently performed by flow cytometry of single nuclear suspensions or image analysis of Feulgen‐stained cytologic specimens. To assess the prognostic significance of ploidy in renal adenomas (less than 3 cm in diameter), nuclear DNA content of 21 renal cortical neoplasms was analyzed by image cytometry (CAS 200, Cell Analysis Systems, Inc., Elmhurst, IL) on Feulgen‐stained 5‐μM fixed embedded tissue sections. Five (36%) of 14 small neoplasms (so‐called renal adenomas; (mean diameter, 7.2 mm; range, 2–24 mm) were in the aneuploid range. Four (57%) of seven carcinomas (mean diameter, 6.3 cm; range, 3.5–10 cm) were aneuploid, a frequency not significantly diifferent from that in the adenomas (P = 0.4). Mean DNA index (1.38) for aneuploid adenomas and carcinomas was identical. Diploid renal adenomas (mean diameter, 5.9 mm) were smaller, although not significantly so (P > 0.05), than the aneuploid ones (mean, 9.7 mm). Size of diploid (mean, 6.7 cm) and aneeuploid (mean, 6.1 cm) renal carcinomas was similar. Clinical follow‐up studies to evaluate utility of DNA ploidy and proliferative activity in prognosis and management of small renal cortical neoplasms are indicated. © 1994 Wiley‐Liss, Inc.
Detection of terminal deoxynucleotidyl transferase by flow cytometry: A three color method
Authors: Horvatinovich Joseph M. , Sparks Sara D. , Borowitz Michael J. 
Published: June 2005

AbstractThis short communication describes a three‐color flow cytometric analysis for terminal deoxynucleotidyl transferase (TdT). The method combines CD45‐gating principles and a new permeabilization method based on a commercially prepared lysing reagent. While traditional permeabilizing fixatives are often cumbersome to use, distort cell light scatter, and fail to retain the fluorochrome peridin chlorophyll alpha protein (perCP), this simple method retains light scatter and does not affect perCP, making it ideal for three‐color analysis. The procedure simply involves staining whole blood or bone marrow with CD45 perCP and an appropriate phycoerythrin (PE) conjugated monoclonal antibody (MoAb) and lysing with FACS lysing solution (Becton Dickinson, San Jose, CA). After washing the cells, fluorescein isothiocyanate (FITC) conjugated anti‐TdT is added. The cells are washed, fixed with formaldehyde, and acquired on a flow cytometer compensated for three‐color analysis. Display of CD45 perCP vs. right angle light scatter (RALS) allows identification of blasts. Dual color expressionof anti‐TdT FITC and a PE conjugated MoAb identifies TdT useful for TdT analysis but may also prove to be a valuable tool for looking at expression of other cytoplasmic antigens in combination with surface antigens on CD45‐defined blast populations. © 1994 Wiley‐Liss, Inc.
CD3+, CD56+ non‐MHC restricted cytotoxic T lymphocytes in two fraternal aids patients: A case report
Authors: Warzynski Michael J. , Barbour Stephen D. , Winkler‐Pickett Robin T. , Ortaldo John R. 
Published: June 2005

AbstractWe describe two brothers with hemophilia and AIDS, with an unusually large percentage of CD3+, CD56+ lymphocytes. They experienced no major complications with opportunisitic infections (OI) and infrequent secondary infections, even though they had nearly 0% CD4 lymphocytes for 3 years. Both patients died in 1991 of progressive cardiomyopathy. The patients' lymphocytes were immunophenotyped by flow cytometry and analyzed for functional cytolytic activity against K562 and HIV infected HUT 78 cell lines. Single color CD4 counts were 2–9% for 4 years. Dual color CD4 counts at our laboratory demonstrated 0–1% CD4 for the last 6 months. When CD3+ lymphocytes were examined, both patient 1 and patient 2 demonstrated a significantly higher proportion and absolute number of CD3“bright”+, CD56+ double‐positive cells, 47% and 22%, respectively, compared to other HIV‐positive children with hemophilia (≤2%). Functional studies with could not be augmented by in vitro addition of IL‐2, whereas patient 2 had no NK activity unless IL‐2 was added. Functional studies with HIV‐infected HUT‐78 cells demonstrated patient 2 had cytolic activity against HIV‐infected cells and patient 1 had high nonspecific cytolytic activity even against uninefected HUT‐78 cells, whereas controls had minimal activity to HUT‐78 cells or HIV‐infected HUT‐78 cells. The case report raises a speculative question requiring a larger database, whether the anti‐HIV activity and/or unuasal clinical course without typical 0.1. of some AIDS patients may be related to the presence of CD3“bright” +, CD56+ lymphocytes of their immune system. © 1994 Wiley‐Liss, Inc.