SEARCH · PubMed Health

Results for “Flow Cytometry”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,297 records · Page 72Linked to original sources

PubMed2001

Allergen-induced basophil activation: CD63 cell expression detected by flow cytometry in patients allergic to Dermatophagoides pteronyssinus and Lolium perenne.

BACKGROUND: In this study, we determined by flow cytometry the percentage of basophils activated after in vitro stimulation by allergens and expressing the CD63 marker. The diagnostic reliability of the technique was assessed as well as its correlation with other in vitro diagnostic parameters. METHODS: Fifty-three patients suffering from asthma and/or allergic rhinitis following sensitization to Dermatophagoides pteronyssinus and 51 patients sensitized to Lolium perenne were investigated. Twenty-four atopic patients not sensitive to these allergens and 38 healthy subjects were also selected as controls. The basophil activation test determines the percentage of basophils which express CD63 as an activation marker, by means of flow cytometry, after in vitro stimulation with allergen, using double labelling with monoclonal antibody anti-CD63-PE and anti-IgE-FITC. RESULTS: No differences in basal values (non-activated control) were found between sensitized patients, atopic controls and healthy controls. On the other hand, sensitized patients showed a significantly higher percentage of activated basophils after stimulation by allergens in vitro than both control groups (P < 0.001). We found a significant correlation between skin tests and basophil activation tests (r = 0.72, P < 0.001). We also found a positive and significant correlation between basophil activation tests and histamine release tests (r = 0.80, P < 0.001), allergen-specific sulphidoleukotriene production (r = 0.7, P < 0.001) and the occurrence of serum allergen-specific IgE (r = 0.71, P < 0.001). CONCLUSION: The basophil activation test is a highly reliable technique in the diagnosis of allergy to inhalant allergens. The sensitivity of the basophil activation test was 93.3%, and its specificity 98.4%, when using a cut-off point of 15% activated basophils as positive result.

Adult↗

PubMed1989

[Significance of flow cytometry in the prognosis of tumors of the kidney in adults].

The DNA index was measured by flow cytometry on paraffin-embedded tissue blocks, from a series of 50 renal tumours studied retrospectively. These tumours consisted of 48 renal cell carcinomas and 1 oncocytoma. The mean follow-up was 3 years. TNM classification and Syrjanen's combined histological grading were used. There was a significant correlation between ploidy and nuclear grading (P = 0.02); between ploidy and TNM classification (72% of diploid tumours were T1 and T2, M0, N0, P = 0.01); between ploidy and development of metastases during the follow-up (71% of aneuploid tumours developed metastases, P = 0.04). In contrast, ploidy was not related with tumour differentiation (P = 0.052), and did not influence the overall long-term survival (P = 0.2). The small number of patients studied did not allow a significant survival rate to be established for each stage according to ploidy. The measurement of the DNA index by flow cytometry is prognostically useful for renal tumours, but must be compared with other prognostic parameters. Its therapeutic value will only be able to be assessed in the future; especially in conservative surgery indications, possibly in the choice of complementary therapeutic associations and even in the approach to treatment of metastases.

Adult↗

PubMed2002

The use of flow cytometry to evaluate temporal changes in inflammatory cells following focal cerebral ischemia in mice.

Recent studies indicate that inflammation following cerebral ischemia contributes to neuronal damage. The local activation of resident cells and efficient recruitment of leukocytes into the central nervous system are critical steps in this inflammatory process. Here we describe studies using flow cytometry to examine the temporal pattern of inflammatory cell activation and infiltration following transient middle cerebral artery occlusion (MCAO) in mice. We found an increase in activated microglia/macrophages as early as 18 h post occlusion, which peaked at 48 h and remained abundant at 96 h post occlusion. Neutrophils were significantly increased by 48 h and remained elevated at 96 h post occlusion. T lymphocytes were increased relatively late (72 and 96 h) post occlusion. The flow cytometry data correlate well both quantitatively and qualitatively with immunohistochemistry analysis performed on the same mice. The present study demonstrates the power of flow cytometry in analyzing the inflammatory process following cerebral ischemia and offers temporal information on the cellular changes in mice following transient MCAO.

Animals↗

PubMed1998

Quantification of somatic and spermatogenic cell proliferation in the testes of ruminants, using a proliferation marker and flow cytometry analysis.

We compared 2 methods for the quantification of proliferation in somatic and spermatogenic compartments of post mortem-collected testes in cattle and roe deer. Proliferation was evaluated by estimation of the tissue polypeptid specific antigen (TPS) using an ELISA. This proliferation-specific marker was detected in homogenized cells after selective enrichment of different cell types by density gradient centrifugation. The haploid, diploid and tetraploid cells were monitored by one-parameter flow cytometry and analyzed for mitotic cell cycle. Somatic and spermatogenic cells were discriminated by dual-parameter flow cytometry after DNA staining with propidium iodide and selective labelling of stromatic cells with a vimentin antibody. The TPS was related to the ploidy of cells and their somatic or spermatogenic type. High concentrations of TPS were found in both species. The TPS values varied with different contents of spermatogenic and somatic cells in the fractions of the density gradient. The TPS was positively correlated with spermatogenic cells in the G2/M phase of mitotic cycle (r = 0.474; P < 0.01) and negatively correlated with somatic cells (r = -0.676; P < 0.0001) in roe deer (n = 40). Discrimination of germinative and stromatic cells in the G2-M phase showed their varying proliferation during the annual cycle in roe deer. The quantification of tetraploid spermatogenic cells allowed the calculation of an exact meiotic transformation (ratio haploid:tetraploid cells). In conclusion, TPS indicates proliferation in the germinative compartment of the testes. However, this marker provides only relative values, without information on the number and type of proliferating cells. Dual-parameter flow cytometry using specific staining for vimentin proves to be a better method for studying changing mitotic and meiotic steps during the involution and recrudescence of testes in seasonally breeding ruminants, as it relates proliferative processes directly to both spermatogenic and somatic cells.

Animals↗

PubMed1994

DNA ploidy measurements in prostate cancer: differences between image analysis and flow cytometry and clinical implications.

The relationship between DNA ploidy and prognostic parameters has been established in prostate cancer. However, comparison of various techniques used for DNA ploidy analysis has not been sufficiently addressed in prostate carcinoma. In the present study, grossly identified discrete carcinoma foci from 48 consecutive radical prostatectomy specimens were analyzed by flow cytometry and image analysis using both imprints and tissue sections. Correlations with other prognostically important pathological findings such as grade, size, extracapsular extension, positive surgical margin, and seminal vesicle and lymph node involvement were done. Image analysis detected 26 (54.2%) nondiploid tumors compared to 15 (31.3%) nondiploid tumors by flow cytometry. No significant differences were demonstrated between imprints and tissue sections. DNA ploidy status showed a good correlation with most other pathological findings listed above. In conclusion, DNA ploidy status determination gives clinically useful information which correlates well with the other useful pathological parameters of prognostic importance. Image analysis may be more sensitive than flow cytometry in detecting nondiploid populations. Tissue sections appear to be as reliable as imprints when properly controlled.

Adenocarcinoma↗

PubMed1997

A comparison of conventional microscopy, immunofluorescence microscopy and flow cytometry in the detection of Giardia lamblia cysts in beaver fecal samples.

A variety of domestic and wild animals are considered to be potential sources of giardiasis in humans. As a result, numerous studies have been reported on the prevalence of Giardia lamblia infection in animals. The majority of these surveys have involved various floatation techniques followed by conventional microscopy in order to detect cysts in fecal samples. Immunofluorescence microscopy has become popular in recent years for the detection of G. lamblia cysts in both clinical and environmental samples. This technique can be automated by combining it with flow cytometry. The present study represents a direct comparison of conventional microscopy, immunofluorescence microscopy, and flow cytometry in terms of their relative efficiency in the detection of G. lamblia cysts in beaver fecal samples. As a result of viewer fatigue, or low cyst concentrations, false negatives were common with conventional microscopy, leading to low prevalence estimates. By specifically targeting the cysts, immunofluorescence microscopy provided more reliable results in a shorter time than conventional methods. When flow cytometry was used in combination with immunofluorescence, a larger number of samples could be examined in a relatively short period of time. The results obtained indicated that this technique allowed for more consistent recognition than either conventional or immunofluorescence microscopy of positive samples containing smaller numbers of cysts.

Animals↗

PubMed1992

Are DNA flow cytometry measurements providing useful information in the management of the node-negative breast cancer patient?

The appropriate management of the breast cancer patient with early stage disease is a controversial, frustrating issue. If laboratory tests could accurately predict tumor behavior, however, the clinician and patient would be greatly aided in their treatment decisions. Although imperfect, there are several new and significant factors that can be used to predict patient prognosis; the most promising and well studied of these factors are DNA flow cytometry measurements. There are at least two estimates of tumor aggressiveness that we can obtain from DNA flow cytometry: one is an estimate of the tumor DNA content or ploidy and the other is an estimate of the tumor proliferative capacity. These measurements have their greatest clinical impact in the node negative patient predicting for relapse-free survival and overall survival. Estimates of proliferative capacity are independent predictors of patient prognosis. Estimates of DNA content are at times controversial and yet still are helpful in distinguishing prognostic subgroups of proliferative activity and may have additional clinical relevance. This discussion will summarize the data obtained from DNA flow cytometry measurements supporting their use as clinically important markers of prognosis in the node-negative patient.

Breast Neoplasms↗

PubMed2005

Flow cytometry of Candida albicans for investigations of surface marker expression and phagocytosis.

Several cytoplasmic virulence factors of Candida albicans are altered in the presence of estrogen and this fact may imply the existence of a global virulence regulatory system in this organism. The response of virulence-associated surface markers to estrogen, however, has not been studied. We exploited flow cytometry methods for assessment of the iC3b receptor analog and mannoproteins on 2 clinical yeast strains selected for their different rates of growth in the presence of estradiol 17beta. Although, as expected, iC3b receptor analog expression increased in the presence of glucose, growth in the presence of estradiol did not increase the levels of iC3b receptor analog on either organism. Exposure to human serum caused massive conversion to mycelial growth, but cells examined by flow cytometry did not show increased levels of iC3b receptor analog expression, possibly due to inability of the flow cytometer to sample the mycelial forms of Candida. In contrast, estradiol increased expression of mannoproteins as evidenced by concanavalin A binding to yeast. This increase occurred in both yeast strains but was less pronounced with strain GT188, which also showed limited growth in estradiol compared to strain GT142. Effective phagocytosis by human neutrophils required exposure of yeast to human serum. Yeast grown in the presence of estradiol were ingested by human PMN but not at a significantly greater rate than yeast grown without estradiol. While flow cytometry appears to be useful in determining estrogen-enhanced concanavalin A binding to yeast, it probably does not reflect the surface markers on large mycelial masses. Consequently, the results of this study are applicable to Candida primarily in its yeast form.

Antibodies, Monoclonal↗

PubMed1991

Proliferative activity of human tumors: assessment using bromodeoxyuridine and flow cytometry.

Cell kinetics of human carcinoma xenografts and human solid tumors were evaluated by means of two-color flow cytometry using single sampling at appropriate time intervals after bromodeoxyuridine (BrdU) labeling. The tumors were resected several hours after the administration of BrdU, and flow cytometry was used to measure the DNA content and BrdU incorporation. Once the BrdU labeling index (LI) and DNA synthesis time (Ts) were obtained, the potential doubling time (Tpot) was calculated from these values. In 11 xenografts, the cell kinetic data were compared with the actual tumor doubling time (Td), and a good correlation between Tpot and Td was obtained (r = 0.91, P less than 0.005). In a clinical study, 33 patients with gastric cancer, colorectal cancer, lung cancer, or other solid tumors were analyzed. Aneuploid tumors had a significantly higher LI value (P less than 0.01) and a shorter Tpot than diploid tumors. The cell loss rate of human tumors ranged from 40% to 80%. These cell kinetic parameters therefore accurately indicated the level of proliferative activity of human tumors.

Animals↗

PubMed2005

Advantages of flow cytometry immunophenotyping for the diagnosis of central nervous system non-Hodgkin's lymphoma in AIDS patients.

BACKGROUND: Neurological disorders are common in HIV-infected patients. Central nervous system (CNS) lymphoma should always be considered because it is an important cause of morbidity and mortality. OBJECTIVES: To investigate the clinical utility of flow cytometry immunophenotyping (FCI) in diagnosing or discarding leptomeningeal involvement in HIV-infected patients and to compare its sensitivity with that of conventional cytological methods. METHODS: Fifty-six cerebrospinal fluid (CSF) samples from 29 HIV-infected patients were independently evaluated by flow cytometry and cytology. The description of an aberrant immunophenotype was the criterion used to define the malignant nature of any CSF cell population. RESULTS: FCI and cytology gave concordant results for 48 of the 56 CSF samples studied: 37 were negative for malignancy and 11 had evidence of CNS lymphoma. Discordant results were obtained for eight CSF samples, and the accuracy of the FCI findings could be demonstrated for four CSF samples described as positive for malignancy according to the FCI criteria. CONCLUSIONS: A high level of agreement was found between the results obtained using the two methods, but FCI gave at least 25% higher sensitivity than conventional cytomorphological methods for the detection of malignant cells. This advantage suggests that, in case of negative flow cytometry results, disorders other than non-Hodgkin's lymphoma should be strongly considered.

Adult↗

PubMed1983

Effects of methylprednisolone on concanavalin A-induced human lymphocyte blastogenesis: a comparative analysis by flow cytometry, volume determination and 3H-thymidine incorporation.

The inhibition of concanavalin A-induced human peripheral blood lymphocyte blastogenesis by methylprednisolone (MP) was studied by using flow cytometry and tritiated thymidine (3H-TdR) incorporation. Flow cytometric determinations of volume, low angle forward light scatter, and nucleic acid showed MP to be a potent inhibitor of blastogenesis. The effects were concentration-dependent and correlated with 3H-TdR uptake. By using the single cell analytic capability of flow cytometry, the target stages of the cell cycle where MP affects lymphocyte activation were determined. Evidence is presented that steroids can block both early and late phases of this process.

Adult↗

PubMed1988

Prenatal detection of 46,XY,rec(5),dup q, inv(5)(p13q33) using DNA analysis, flow cytometry, and in situ hybridization to supplement classical cytogenetic analysis.

Distinguishing between balanced and unbalanced chromosome complements segregating from parental rearrangements may be difficult using only classical cytogenetic techniques if banding morphology is similar under both expectations. In these situations, supplementing cytogenetic analysis with molecular genetic techniques and flow cytometry may provide increased diagnostic accuracy. To illustrate this, we present a case in which similar band pattern morphology would be expected for both the balanced carrier (heterozygote) and the recombinant dup q chromosome complements segregating from a mother with a balanced inversion [46,XX,inv(5)(p13q33)]. The parents came to Northwestern for consultation after receiving conflicting interpretations of their first amniotic fluid cultures. An ultrasound examination was said to be normal. They inquired whether there were ways to increase their confidence that the complement was unbalanced. Their reluctance to terminate the pregnancy was due to a 6-year history of infertility. After extensive counselling, the couple elected repeat amniocentesis. Further cytogenetic analysis of repeat amniotic fluid cultures by G-banding and R-banding, molecular genetic analysis with highly polymorphic DNA probes, and quantitative flow cytometry were performed. Results agreed that an unbalanced fetal complement was present. Southern blot analysis with a 5p marker definitively demonstrated a lack of maternal 5p material in the fetus, and in situ hybridization showed a 5q marker at either end of the recombinant chromosome. Flow cytometry was consistent with this interpretation. Because of the advanced gestational age, the parents elected to terminate based on cytogenic results of the second amniocentesis, rather than to wait another 1-2 weeks for results of other methods.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

PubMed1996

DNA ploidy and S-phase in recurrent astrocytomas: a retrospective study by flow cytometry of deparaffinized specimens.

Twenty-two patients with recurrent astrocytic tumors were treated surgically two or even three times. At the time of the first surgery 6 tumors were fibrillary astrocytomas (grade II), 9 anaplastic astrocytomas (grade III) and 7 glioblastomas (grade IV). Histopathological specimens from second surgery demonstrated in 12 cases signs of higher grades of malignancy. Flow cytometry (FCM) did not reveal any significant changes of S-phase fraction (p = 0.55). This study supports the theory that, given enough time, the histopathology of brain tumors change significantly from more benign forms to more malignant ones. The flow cytometry (FCM) could trace a weak tendency to higher S-phase fractions at the time of the second surgery. No apparent change of ploidy pattern was observed. In spite of the unequivocal histopathological changes of the recurrent astrocytomas the flow cytometry failed to indicate similar changes in terms of ploidy and S-phase fraction parameters.

Adult↗

PubMed2000

Viability of Escherichia coli O157:H7 in natural river water determined by the use of flow cytometry.

The enzymatic activity and viability of Escherichia coli O157:H7 in natural river water was determined by flow cytometry. River water was collected at two sites (an agricultural area and an industrial area) on the Aigawa River (Osaka, Japan). To facilitate estimation of the physiology of E. coli O157 in natural river water, bacterial cells in the water were stained with 6-carboxyfluorescein diacetate (6CFDA) and propidium iodide (PI). The cells were sorted into two populations, using a flow cytometer, based on their esterase activity. Each population was stained with E. coli O157:H7 fluorescent antibody (FA), and E. coli O157:H7 cells were observed in the esterase-active population. River water samples collected at the same points were incubated with yeast extract containing antibiotics to prevent cell division, and bacterial cells in the incubated samples were stained with PI and FA. Escherichia coli O157:H7 existed in both the viable (elongated and/or fattened) and inactive bacterial population determined by flow cytometry. These results indicate that E. coli O157:H7 may retain metabolic activity and growth potential in the natural aquatic environment.

Colony Count, Microbial↗

PubMed1992

Purification of human blood basophils using negative selection by flow cytometry.

Basophils were purified from peripheral blood of normal donors using Percoll discontinuous gradients and negative selection by flow cytometry. The mean purity of basophils obtained was 84.7 +/- 4.1 (s.d.)% (range 77.3-90.0%, n = 13). The overall yield of these procedures was 16.0 +/- 2.6% (range 11.0-19.9%, n = 13), and cell viability of purified basophils exceeded 90%. Properties of highly purified basophils obtained by flow cytometry did not differ from those of partially enriched basophil preparations from Percoll discontinuous gradients in respect of: (i) intracellular histamine content; (ii) percentage of spontaneous histamine release in buffer; and (iii) percentage of histamine release triggered by ionophore A 23187 or anti-IgE. Moreover, purified basophils responded chemotactically to complement C5a in a dose-dependent manner. These findings suggest that our procedure for purification of human basophils does not affect the functions of basophils and may be useful for in vitro studies on the role of basophils in hypersensitivity reactions such as bronchial asthma.

Basophils↗

PubMed1995

Measuring resistance to phagocytosis of group A and G streptococci: comparison of direct bactericidal assay and flow cytometry.

M protein is thought to contribute to the ability of non-opsonized group A and group G streptococci (GAS and GGS, respectively) to resist phagocytosis by polymorphonuclear leukocytes. In previous studies, correlation between M protein expression and phagocytosis was determined by incubating these pathogens in human blood and comparing colony-forming bacterial counts prior to and after exposure to blood (direct bactericidal assay; DBA). Here, we report the application of flow cytometry to measure GAS and GGS resistance to phagocytosis. The results of the assays were in complete agreement with those from DBAs. Nevertheless, flow cytometry was regarded as superior to DBA because of its speed and potential uses for quantitative studies. In addition, the use of anti-CD11b monoclonal antibody for granulocyte staining guaranteed a non-compromized granulocyte function. The optimized protocol for flow cytometry presented here could be utilized to directly measure the involvement of individual protein types in bacterial resistance to phagocytosis.

Blood Bactericidal Activity↗

PubMed2003

Single human leukocyte antigen flow cytometry beads for accurate identification of human leukocyte antigen antibody specificities.

BACKGROUND: It is difficult to assign antibody specificity for highly sensitized patients using a cell panel with multiple antigens per reaction. We describe here a single antigen bead panel for accurate identification of human leukocyte antigen (HLA) antibody specificities by flow cytometry. METHODS: A total of 110 single recombinant HLAs, including 34 A locus alleles, 57 B locus alleles, and 19 C locus alleles, were produced by a mammalian expression system. These single antigens were coated onto eight different colored microbeads, which were mixed together in one tube for simultaneous detection of HLA antibodies against eight different antigens per flow cytometry test. RESULTS: Single HLA reacted specifically with the serologically defined monoclonal antibodies. The single antigen panel provided higher resolution than the regular cell panel for antibody detection by uncovering the masked specificities. Single antigens also provided higher sensitivity than the multiple antigens coated onto beads for HLA antibody detection as demonstrated by serum dilution studies. In 10 sera from patients who had rejected a kidney transplant, single antigen beads identified antibodies to 31 of 35 antigens that were mismatched in the donor. Most important, none of the reactions were against antigens present in the recipient. CONCLUSION: An accurate and sensitive HLA antibody detection method is described using flow cytometry beads coated with single HLAs produced by recombinant technology. The single antigen beads should be useful in predicting negative crossmatch in highly sensitized organ recipients and highly sensitized patients requiring platelets.

Antibody Specificity↗

PubMed2000

A ruthenium probe for cell viability measurement using flow cytometry, confocal microscopy and time-resolved luminescence.

The capability of the new luminescent probe (dibenzo[h,j] dipyrido[3,2-a:2',3'-c]phenazine)bis(2,2'-bipyridine)ruthenium(II) dication, (RB2Z), to discriminate live and dead cells has been tested on rat hepatocytes and mouse lymphocytes. RB2Z-stained cells were analyzed using flow cytometry, fluorescence (confocal) microscopy and time-resolved luminescence measurements. The established viability probes propidium iodide (PI) and SYTOX green (SG) were used as controls. The intense luminescence of RB2Z at 606 nm is localized in the nucleus of nonviable cells. Viability measurements by flow cytometry and fluorescence microscopy using RB2Z as dead-cell marker yield the same results as PI and SG. The luminescence lifetime of RB2Z also displays sensitivity to cell viability (0.45 and 0.82 microsecond in presence of fully viable and dead cells, respectively). This ruthenium complex is photostable under laser sources and its 200 nm Stokes shift facilitates multicolor labeling experiments in flow cytometry and fluorescence microscopy. Unlike the currently available probes, the long-lived excited state of RB2Z also allows assays based on luminescence decay measurements.

Animals↗