CCL

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1.

001-es BibID:BIBFORM055851
035-os BibID:(scopus)84949232854
Első szerző:Doan-Xuan, Quang-Minh (biofizikus)
Cím:FRET Imaging by Laser Scanning Cytometry on Large Populations of Adherent Cells / Quang-Minh Doan-Xuan, Nikoletta Szalóki, Katalin Tóth, János Szöllősi, Zsolt Bacso, György Vámosi
Dátum:2014
ISSN:1934-9297 1934-9300
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Current Protocols in Cytometry. - Suppl. 70. (2014), p. [1-29]. -
További szerzők:Szalóki Nikoletta (1981-) (biológus) Tóth Katalin (biofizikus) Szöllősi János (1953-) (biofizikus) Bacsó Zsolt (1963-) (biofizikus) Vámosi György (1967-) (biofizikus)
Internet cím:DOI
Intézményi repozitóriumban (DEA) tárolt változat
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2.

001-es BibID:BIBFORM050042
035-os BibID:PMID:23504771
Első szerző:Fábián Ákos István (aneszteziológus)
Cím:TripleFRET measurements in flow cytometry / Ákos Fábián, Gábor Horváth, György Vámosi, György Vereb, János Szöllősi
Dátum:2013
Megjegyzések:A frequently used method for viewing protein interactions and conformation, Forster (fluorescence) resonance energy transfer (FRET), has traditionally been restricted to two fluorophores. Lately, several methods have been introduced to expand FRET methods to three species. We present a method that allows the determination of FRET efficiency in three-dye systems on a flow cytometer. TripleFRET accurately reproduces energy transfer efficiency values measured in two-dye systems, and it can indicate the presence of trimeric complexes, which is not possible with conventional FRET methods. We also discuss the interpretation of energy transfer values obtained with tripleFRET in relation to spatial distribution of labeled molecules, specifically addressing the limitations of using total energy transfer to determine molecular distance
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
analysis
Antibodies
Antibodies,Monoclonal,Humanized
article
Biophysics
cell biology
Cell Line
chemistry
cytometry
Dyes
Energy Transfer
ENERGY-TRANSFER
FLOW
Flow Cytometry
Fluorescence
Fluorescence Resonance Energy Transfer
fluorescent dye
Fluorescent Dyes
FRET
Humans
Hungary
Immunoconjugates
immunology
methods
Protein Binding
PROTEIN INTERACTIONS
Research
Research Support
resonance energy transfer
Staining and Labeling
statistics & numerical data
Support
Megjelenés:Cytometry Part A. - 83 : 4 (2013), p. 375-385. -
További szerzők:Horváth Gábor (1974-) (biofizikus) Vámosi György (1967-) (biofizikus) Vereb György (1965-) (biofizikus, orvos) Szöllősi János (1953-) (biofizikus)
Pályázati támogatás:NK 101337
OTKA
K75752
OTKA
TÁMOP-4.2.2-08/1-2008-0019
TÁMOP
TÁMOP-4.2.1/B-09/1/KONV-2010-0007
TÁMOP
Receptor tirozin kinázok mint terápiás célpontok : működésük szabályozásának, és a közöttük fellépő molekuláris kölcsönhatások vizsgálata
TÁMOP-4.2.2.A-11/1/KONV-2012-0025
TÁMOP
Baross Gábor Program REG_EA_09-1-2009-0010
Egyéb
K77600
OTKA
K103965
OTKA
Internet cím:DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

3.

001-es BibID:BIBFORM039514
Első szerző:Matkó János (biológus)
Cím:GPI-microdomains (membrane rafts) and signaling of the multi-chain interleukin-2 receptor in human lymphoma/leukemia T cell lines / Matko, J., Bodnar, A., Vereb, G., Bene, L., Vamosi, G., Szentesi, G., Szollosi, J., Gaspar, R., Horejsi, V., Waldmann, T. A., Damjanovich, S.
Dátum:2002
ISSN:0014-2956
Megjegyzések:Subunits (alpha, beta and gamma) of the interleukin-2 receptor complex (IL-2R) are involved in both proliferative and activation-induced cell death (AICD) signaling of T cells. In addition, the signaling beta and gamma chains are shared by other cytokines (e.g. IL-7, IL-9, IL-15). However, the molecular mechanisms responsible for recruiting/sorting the alpha chains to the signaling chains at the cell surface are not clear. Here we show, in four cell lines of human adult T cell lymphoma/leukemia origin, that the three IL-2R subunits are compartmented together with HLA glycoproteins and CD48 molecules in the plasma membrane, by means of fluorescence resonance energy transfer (FRET), confocal microscopy and immuno-biochemical techniques. In addition to the beta and gamma(c) chains constitutively expressed in detergent-resistant membrane fractions (DRMs) of T cells, IL-2Ralpha (CD25) was also found in DRMs, independently of its ligand-occupation. Association of CD25 with rafts was also confirmed by its colocalization with GM-1 ganglioside. Depletion of membrane cholesterol using methyl-beta-cyclodextrin substantially reduced co-clustering of CD25 with CD48 and HLA-DR, as well as the IL-2 stimulated tyrosine-phosphorylation of STATs (signal transducer and activator of transcription). These data indicate a GPI-microdomain (raft)-assisted recruitment of CD25 to the vicinity of the signaling beta and gamma(c) chains. Rafts may promote rapid formation of a high affinity IL-2R complex, even at low levels of IL-2 stimulus, and may also form a platform for the regulation of IL-2 induced signals by GPI-proteins (e.g. CD48). Based on these data, the integrity of these GPI-microdomains seems critical in signal transduction through the IL-2R complex.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
Megjelenés:European Journal Of Biochemistry. - 269 : 4 (2002), p. 1199-1208. -
További szerzők:Dóczy-Bodnár Andrea (1970-) (biofizikus) Vereb György (1965-) (biofizikus, orvos) Bene László (1963-) (biofizikus) Vámosi György (1967-) (biofizikus) Szentesi Gergely (1976-) (kémia-fizika tanár) Szöllősi János (1953-) (biofizikus) Gáspár Rezső (1944-) (biofizikus) Horejsi, Václav Waldmann, Thomas A. Damjanovich Sándor (1936-2017) (biofizikus)
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DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

4.

001-es BibID:BIBFORM065104
035-os BibID:(WoS)000380371400013 (Scopus)84993661764
Első szerző:Mocsár Gábor (biofizikus)
Cím:MHC I expression regulates co-clustering and mobility of interleukin-2 and -15 receptors in T cells / G. Mocsár, J. Volkó, D. Rönnlund, J. Widengren, P. Nagy, J. Szöllősi, K. Tóth, C. K. Goldman, S. Damjanovich, T. A. Waldmann, A. Bodnár, G. Vámosi
Dátum:2016
ISSN:0006-3495
Megjegyzések:MHC glycoproteins form supramolecular clusters with interleukin-2 and -15 receptors in lipid rafts of T cells.The role of highly expressed MHC I in maintaining these clusters is unknown. We knocked down MHC I inFT7.10 human T cells, and studied protein clustering at two hierarchic levels: molecular aggregations andmobility by FRET and fluorescence correlation spectroscopy, and segregation into larger domains orsuperclusters by superresolution STED microscopy. FCS based molecular brightness analysis revealed thatthe studied molecules diffused as tight aggregates of several proteins of a kind. Knockdown reduced thenumber of MHC I containing molecular aggregates and their average MHC I content, and decreased theheteroassociation of MHC I with IL-2R?/IL-15R?. The mobility of not only MHC I but also that of IL-2R?/IL-15R? increased, corroborating the general size decrease of tight aggregates. A multifaceted analysis of STEDimages revealed that the diameter of MHC I superclusters diminished from 400-600 to 200-300 nm, whereasthose of IL-2R?/IL-15R? hardly changed. MHC I and IL-2R?/IL-15R? colocalized with GM1 gangliosiderichlipid rafts, but MHC I clusters retracted to smaller subsets of GM1- and IL-2R?/IL-15R?-rich areas uponknockdown. Our results prove that changes in expression level may significantly alter the organization andmobility of interacting membrane proteins.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Biophysical Journal. - 111 : 1 (2016), p. 100-112. -
További szerzők:Volkó Julianna (1983-) (biotechnológus) Rönnlund, Daniel Widengren, Jerker Nagy Péter (1971-) (biofizikus) Szöllősi János (1953-) (biofizikus) Tóth Katalin (Heidelberg) Goldman, Caroline K. Damjanovich Sándor (1936-2017) (biofizikus) Waldmann, Thomas A. Dóczy-Bodnár Andrea (1970-) (biofizikus) Vámosi György (1967-) (biofizikus)
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DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

5.

001-es BibID:BIBFORM004633
035-os BibID:(scopus)0344447088 (wos)000074651400008
Első szerző:Nagy Péter (biofizikus)
Cím:Intensity-based energy transfer measurements in digital imaging microscopy / Nagy P., Vamosi G., Bodnar A., Lockett S. J., Szollosi J.
Dátum:1998
ISSN:175-7571
Megjegyzések:Investigation of protein-protein associations is important in understanding structure and function relationships in living cells. Using Forster-type resonance energy transfer between donor and acceptor labeled monoclonal antibodies we can assess the cell surface topology of membrane proteins against which the antibodies were raised. In our current work we elaborated a quantitative image microscopic technique based on the measurement of fluorescence intensities to calculate the energy transfer efficiency on a pixel-by-pixel basis. We made use of the broad excitation and emission spectrum of cellular autofluorescence for background correction of images. In addition to the reference autofluorescence images (UV background) we recorded three fluorescent images (donor, acceptor and energy transfer signal) of donor-acceptor double labeled samples, and corrected for spectral spillage of the directly excited donor and acceptor fluorescence into the energy transfer image. After careful image registration we were able to calculate the energy transfer efficiency on a pixel-by -pixel basis. In this paper, we also present a critical comparison between results obtained with this method and other approaches (photobleaching and flow cytometric energy transfer measurements)
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Antibodies
Antibodies, Monoclonal
Biophysics
Cell Line
chemistry
Comparative Study
Energy Transfer
Fluorescence
Human
Hungary
Image Processing, Computer-Assisted
immunology
Membrane Proteins
methods
Microscopy
Microscopy, Fluorescence
Models, Theoretical
Proteins
Signal Processing,Computer-Assisted
Spectrometry, Fluorescence
statistics and numerical data
Support, Non-U.S.Gov't
Megjelenés:European Biophysics Journal. - 27 : 4 (1998), p. 377-389. -
További szerzők:Vámosi György (1967-) (biofizikus) Dóczy-Bodnár Andrea (1970-) (biofizikus) Lockett, Steven J. Szöllősi János (1953-) (biofizikus)
Internet cím:elektronikus változat
Borító:

6.

001-es BibID:BIBFORM039658
035-os BibID:(scopus)0036628631 (wos)000176656500002
Első szerző:Sebestyén Zsolt
Cím:Long wavelength fluorophores and cell-by-cell correction for autofluorescence significantly improves the accuracy of flow cytometric energy transfer measurements on a dual-laser benchtop flow cytometer / Zsolt Sebestyén, Péter Nagy, Gábor Horváth, György Vámosi, Reno Debets, Jan W. Gratama, Denis R. Alexander, János Szöllősi
Dátum:2002
ISSN:0196-4763
Megjegyzések:Flow cytometric fluorescence resonance energy transfer (FCET) is an efficient method to map associations between biomolecules because of its high sensitivity to changes in molecular distances in the range of 1-10 nm. However, the requirement for a dual-laser instrument and the need for a relatively high signal-to-noise system (i.e., high expression level of the molecules) pose limitations to a wide application of the method.METHODS:Antibodies conjugated to cyanines 3 and 5 (Cy3 and Cy5) were used to label membrane proteins on the cell surface. FCET measurements were made on a widely used benchtop dual-laser flow cytometer, the FACSCalibur, by using cell-by-cell analysis of energy transfer efficiency.ResultsTo increase the accuracy of FCET measurements, we applied a long wavelength donor-acceptor pair, Cy3 and Cy5, which beneficially affected the signal-to-noise ratio in comparison with the classic pair of fluorescein and rhodamine. A new algorithm for cell-by-cell correction of autofluorescence further improved the sensitivity of the technique; cell subpopulations with only slightly different FCET efficiencies could be identified. The new FCET technique was tested on various direct and indirect immunofluorescent labeling strategies. The highest FCET values could be measured when applying direct labeling on both (donor and acceptor) sides. Upon increasing the complexity of the labeling scheme by introducing secondary antibodies, we detected a decrease in the energy transfer efficiency.CONCLUSIONS:We developed a new FCET protocol by applying long wavelength excitation and detection of fluorescence and by refining autofluorescence correction. The increased accuracy of the new method makes cells with low receptor expression amenable to FCET investigation, and the new approach can be implemented easily on a commercially available dual-laser flow cytometer, such as a FACSCalibur.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Algorithms
analysis
Antibodies
Biophysics
Calibration
Cell Line, Transformed
Cells
Comparative Study
Dyes
Energy Transfer
Flow Cytometry
Fluorescein
Fluorescence
Fluorescence Resonance Energy Transfer
Fluorescent Dyes
Humans
Hungary
instrumentation
Lasers
Membrane Proteins
methods
Proteins
Reproducibility of Results
Research
Spectrometry, Fluorescence
Subtraction Technique
Support
Tumor Cells, Cultured
egyetemen (Magyarországon) készült közlemény
Megjelenés:Cytometry. - 48 : 3 (2002), p. 124-135. -
További szerzők:Nagy Péter (1971-) (biofizikus) Horváth Gábor (1974-) (biofizikus) Vámosi György (1967-) (biofizikus) Debets, Reno Gratama, Jan Willem Alexander, Denis R. Szöllősi János (1953-) (biofizikus)
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DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

7.

001-es BibID:BIBFORM050053
Első szerző:Szalóki Nikoletta (biológus)
Cím:High throughput FRET analysis of protein-protein interactions by slide-based imaging laser scanning cytometry / Nikoletta Szalóki, Quang Minh Doan-Xuan, János Szöllősi, Katalin Tóth, György Vámosi, Zsolt Bacsó
Dátum:2013
Megjegyzések:Laser scanning cytometry (LSC) is a slide-based technique combining advantages of flow and image cytometry: automated, high-throughput detection of optical signals with subcellular resolution. Fluorescence resonance energy transfer (FRET) is a spectroscopic method often used for studying molecular interactions and molecular distances. FRET has been measured by various microscopic and flow cytometric techniques. We have developed a protocol for a commercial LSC instrument to measure FRET on a cell-by-cell or pixel-by-pixel basis on large cell populations, which adds a new modality to the use of LSC. As a reference sample for FRET, we used a fusion protein of a single donor and acceptor (ECFP-EYFP connected by a seven-amino acid linker) expressed in HeLa cells. The FRET efficiency of this sample was determined via acceptor photobleaching and used as a reference value for ratiometric FRET measurements. Using this standard allowed the precise determination of an important parameter (the alpha factor, characterizing the relative signal strengths from a single donor and acceptor molecule), which is indispensable for quantitative FRET calculations in real samples expressing donor and acceptor molecules at variable ratios. We worked out a protocol for the identification of adherent, healthy, double-positive cells based on light-loss and fluorescence parameters, and applied ratiometric FRET equations to calculate FRET efficiencies in a semi-automated fashion. To test our protocol, we measured the FRET efficiency between Fos-ECFP and Jun-EYFP transcription factors by LSC, as well as by confocal microscopy and flow cytometry, all yielding nearly identical results. Our procedure allows for accurate FRET measurements and can be applied to the fast screening of protein interactions. A pipeline exemplifying the gating and FRET analysis procedure using the CellProfiler software has been made accessible at our web site
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
Acceptor molecules
analysis
article
Biophysics
cell biology
Cells
Confocal
Confocal microscopy
cytometry
Energy Transfer
ENERGY-TRANSFER
FLOW
Flow Cytometry
Fluorescence
FLUORESCENCE RESONANCE
Fluorescence Resonance Energy Transfer
FRET
Hela Cells
Hungary
Image Cytometry
Laser Scanning Cytometry
Microscopy
molecular interaction
MOLECULAR-INTERACTIONS
Photobleaching
PROTEIN INTERACTIONS
protein protein interaction
Protein-protein interactions
Research
Research Support
resonance energy transfer
Software
Support
Megjelenés:Cytometry. Part A. - 83 : 9 (2013), p. 818-829. -
További szerzők:Doan-Xuan, Quang-Minh (1986-) (biofizikus) Szöllősi János (1953-) (biofizikus) Tóth Katalin (biofizikus) Vámosi György (1967-) (biofizikus) Bacsó Zsolt (1963-) (biofizikus)
Pályázati támogatás:K 77600
OTKA
K 103965
OTKA
NK 101337
OTKA
K 75752
OTKA
CK78179
OTKA
TÁMOP-4.2.1/B-09/1/KONV-2010-0007
TÁMOP
TÁMOP-4.2.2.A-11/1/KONV-2012-0023-"VÉD-ELEM"
TÁMOP
TÁMOP-4.2.2.A-11/1/KONV-2012-0025
TÁMOP
Internet cím:DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

8.

001-es BibID:BIBFORM040284
Első szerző:Szegedi István (hematológus, onkológus, nefrológus)
Cím:Differential regulation of umbilical cord blood and leukemic B cells by interferon-alpha (IFN-[alfa]) : observations in cultured cells / Szegedi I., Kiss C., Karászi E., Vámosi G., Szöllősi J., Kovács P., Benkő I.
Dátum:2006
ISSN:1219-4956
Megjegyzések:The exact mechanism of the beneficial therapeutic action of interferon-a (IFN-alpha) in B-cell-lineage malignancies has not been adequately explained. Here we report on the differential effect of IFN-alpha2b on non-malignant B cells of umbilical cord blood and leukemic B-cell lines JY, BL-41 and BCBL-1. Leukemic cell proliferation was characterized by colony assay, whereas apoptosis was investigated by flow cytometry of propidium iodide-stained cells. The degree of differentiation was evaluated by measuring the expression level of Fcgamma receptor-II (FcgammaRII) labeled with anti-CD32-FITC monoclonal antibody using flow cytometry. IFN-alpha protected umbilical cord blood CD19-positive B lymphocytes from apoptotic cell death in vitro. IFN-alpha significantly decreased colony formation of all three cell lines, and in contrast to normal cells, induced apoptosis in JY and BL-41 and excessive necrosis in HHV-8 infected BCBL-1 cells. FcgammaRII was upregulated both in normal and in leukemic B cells as indicated by an increase both in the proportion of CD32-positive cells and the mean fluorescence intensity. From our results it seems that antiproliferative, apoptotic and differentiative effects of IFN-alpha are interrelated but distinct cellular events, which are differentially regulated in normal, leukemic and virus-infected cells of the B-cell lineage.
Tárgyszavak:Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Megjelenés:Pathology & Oncology Research. - 12 : 3 (2006), p. 159-163. -
További szerzők:Kiss Csongor (1956-) (hematológus, onkológus) Karászi Éva Vámosi György (1967-) (biofizikus) Szöllősi János (1953-) (biofizikus) Kovács Péter (1939-) (farmakológus) Benkő Ilona (1954-) (orvos, farmakológus)
Internet cím:Szerző által megadott URL
DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

9.

001-es BibID:BIBFORM004846
035-os BibID:(scopus)3242784810 (wos)000223353700003
Első szerző:Szentesi Gergely (kémia-fizika tanár)
Cím:Computer program for determining fluorescence resonance energy transfer efficiency from flow cytometric data on a cell-by-cell basis / Szentesi, G., Horvath, G., Bori, I., Vamosi, G., Szollosi, J., Gaspar, R., Damjanovich, S., Jenei, A., Matyus, L.
Dátum:2004
Megjegyzések:The determination of fluorescence resonance energy transfer (FRET) with flow cytometry (FCET) is one of the most efficient tools to study the proximity relationships of cell membrane components in cell populations on a cell-by-cell basis. Because of the high amount of data and the relatively tedious calculations, this procedure should be assisted by powerful data processing software. The currently available programs are not able to fulfill this requirement. We developed a Windows-based program to calculate fluorescence resonance energy transfer efficiency values from list mode flow cytometry standard (FCS) files. This program displays the measured data in standard plots by generating one- and two-parameter histograms on linear or logarithmic scales. A graphical gating tool allows the user to select the desired cell population according to any combination of the parameter values. The program performs several statistical calculations, including mean, S.D., percent of the gated data. We have implemented two types of data sheet for FRET calculations to aid and guide the user during the analysis: one with population-mean-based autofluorescence correction and the other with spectrum-based cell-by-cell autofluorescence correction. In this paper, we describe the gating algorithms, the file opening procedure and the rules of gating. The structure of the program and a short description of the graphical user-interface (GUI) are also presented in this article.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Algorithms
analysis
Biophysics
Cell Membrane
Computer Simulation
Energy Transfer
Flow Cytometry
Fluorescence
Fluorescence Resonance Energy Transfer
Humans
Hungary
methods
Research
Software
Support
egyetemen (Magyarországon) készült közlemény
Megjelenés:Computer Methods and Programs in Biomedicine. - 75 : 3 (2004), p. 201-211. -
További szerzők:Horváth Gábor (1974-) (biofizikus) Bori Imre (1929-2004) Vámosi György (1967-) (biofizikus) Szöllősi János (1953-) (biofizikus) Gáspár Rezső (1944-) (biofizikus) Damjanovich Sándor (1936-2017) (biofizikus) Jenei Attila (1966-) (biofizikus) Mátyus László (1956-) (biofizikus)
Internet cím:elektronikus változat
DOI
Borító:

10.

001-es BibID:BIBFORM042722
Első szerző:Szűcs Sándor (biokémikus, vegyész)
Cím:Single-cell measurement of granulocyte respiratory burst with digital imaging fluorescence microscopy / S. Szűcs, G. Vámosi, R. Póka, A. Sárváry, H. Bárdos, M. Balázs, L. Tóth, J. Kappelmayer, J. Szőllősi, R. Ádány
Dátum:1998
ISSN:0007-1048
Tárgyszavak:Orvostudományok Elméleti orvostudományok idézhető absztrakt
Megjelenés:British Journal of Haematology. - 102 : 1 (1998), p. 2-139. -
További szerzők:Vámosi György (1967-) (biofizikus) Póka Róbert (1960-) (szülész-nőgyógyász, klinikai onkológus) Sárváry Attila (1971-) (népegészségtan szakorvos) Bárdos Helga (1969-) (megelőző orvostan és népegészségtan szakorvos) Balázs Margit (1952-) (sejtbiológus, molekuláris genetikus) Tóth L. Kappelmayer János (1960-) (laboratóriumi szakorvos) Szöllősi János (1953-) (biofizikus) Ádány Róza (1952-) (megelőző orvostan és népegészségtan szakorvos)
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DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

11.

001-es BibID:BIBFORM004640
Első szerző:Szűcs Sándor (biokémikus, vegyész)
Cím:Single-cell measurement of superoxide anion and hydrogen peroxide production by human neutrophils with digital imaging fluorescence microscopy / Sándor Szűcs, György Vámosi, Róbert Póka, Attila Sárváry, Helga Bárdos, Margit Balázs, János Kappelmayer, László Tóth, János Szöllősi, Róza Ádány
Dátum:1998
ISSN:0196-4763
Megjegyzések:Besides flow cytometry, fluorescence microscopy combined with computerized image analysis offers an alternative tool for assessing phagocyte oxidant generation at the single-cell level. This technique provides an opportunity for the direct visualization of cells and simultaneous measurement of cellular fluorescence intensity. Thus, we developed a simple method for the quantitative evaluation of intracellular superoxide anion and hydrogen peroxide production with image cytometry by using hydroethidine and dihydrorhodamine 123 dyes, respectively. Human neutrophils stimulated with phorbol dibutyrate and labeled by these fluorogenic substrates showed intense, well recognizable red or green fluorescence. The intensity of signals from individual granulocytes of cytospin preparations were quantitatively measured in digitized images. There was a great heterogeneity in response to the stimulus within the granulocyte population as shown by the integrated fluorescence intensity values. In agreement with the results of parallel flow cytometric experiments, this simple image analysis performed on cells of cytospin preparations was able to detect the defects in the oxidative metabolism of neutrophils from patients with cervix carcinoma. We demonstrated that even minor alterations in superoxide anion/hydrogen peroxide generation can be detected by image cytometry as efficiently as by flow cytometry. This result validates imaging microscopy as an alternative to flow cytometry in such experiments. In addition, the image cytometric technique allows the observation of the kinetics of free radical production in individual cell under adherent conditions. Therefore, we carried out image analysis of the oxidative burst of neutrophils adherent to uncoated glass and fibronectin- and type IV collagen-coated surfaces in response to stimulation with phorbol dibutyrate or N-formyl-methionyl-leucyl-phenylalanine. We elaborated a calibration technique for the quantitative measurement of the ethidium bromide generation mediated by superoxide anion within individual adherent granulocytes. The ethidium bromide production varied between 0.48 and 1.17 amol/cell/min.
Tárgyszavak:Orvostudományok Elméleti orvostudományok idegen nyelvű folyóiratközlemény külföldi lapban
analysis
Anions
Carcinoma
Dyes
epidemiology
Ethidium
Flow Cytometry
Fluorescence
Human
Hungary
Hydrogen Peroxide
Image Cytometry
Image Processing
Computer-Assisted
Kinetics
metabolism
methods
Microscopy
Microscopy
Fluorescence
Neutrophils
Superoxides
Support
Non-U.S.Gov't
egyetemen (Magyarországon) készült közlemény
Megjelenés:Cytometry. - 33 : 1 (1998), p. 19-31. -
További szerzők:Vámosi György (1967-) (biofizikus) Póka Róbert (1960-) (szülész-nőgyógyász, klinikai onkológus) Sárváry Attila (1971-) (népegészségtan szakorvos) Bárdos Helga (1969-) (megelőző orvostan és népegészségtan szakorvos) Balázs Margit (1952-) (sejtbiológus, molekuláris genetikus) Kappelmayer János (1960-) (laboratóriumi szakorvos) Tóth László (orvos) Szöllősi János (1953-) (biofizikus) Ádány Róza (1952-) (megelőző orvostan és népegészségtan szakorvos)
Internet cím:DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:

12.

001-es BibID:BIBFORM079931
035-os BibID:(cikkazonosító)3370 (scopus)85071327332 (wos)000477041100254
Első szerző:Vámosi György (biofizikus)
Cím:EGF Receptor Stalls upon Activation as Evidenced by Complementary Fluorescence Correlation Spectroscopy and Fluorescence Recovery after Photobleaching Measurements / Vámosi György, Friedländer-Brock Elza, Ibrahim Shehu M., Brock Roland, Szöllősi János, Vereb György
Dátum:2019
ISSN:1661-6596 1422-0067
Megjegyzések:To elucidate the molecular details of the activation-associated clustering of epidermal growth factor receptors (EGFRs), the time course of the mobility and aggregation states of eGFP tagged EGFR in the membranes of Chinese hamster ovary (CHO) cells was assessed by in situ mobility assays. Fluorescence correlation spectroscopy (FCS) was used to probe molecular movements of small ensembles of molecules over short distances and time scales, and to report on the state of aggregation. The diffusion of larger ensembles of molecules over longer distances (and time scales) was investigated by fluorescence recovery after photobleaching (FRAP). Autocorrelation functions could be best fitted by a two-component diffusion model corrected for triplet formation and blinking. The slow, 100?1000 ms component was attributed to membrane localized receptors moving with free Brownian diffusion, whereas the fast, ms component was assigned to cytosolic receptors or their fragments. Upon stimulation with 50 nM EGF, a significant decrease from 0.11 to 0.07 ?m2/s in the diffusion coefficient of membrane-localized receptors was observed, followed by recovery to the original value in ~20 min. In contrast, the apparent brightness of diffusing species remained the same. Stripe FRAP experiments yielded a decrease in long-range molecular mobility directly after stimulation, evidenced by an increase in the recovery time of the slow component from 13 to 21.9 s. Our observations are best explained by the transient attachment of ligand-bound EGFRs to immobile or slowly moving structures such as the cytoskeleton or large, previously photobleached receptor aggregates.
Tárgyszavak:Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Fluorescence correlation spectroscopy
FCS
fluorescence recovery after photobleaching
FRAP
epidermal growth factor receptor
translational diffusion
EGFR?eGFP fusion protein
Megjelenés:International Journal Of Molecular Sciences. - 20 : 13 (2019), p. 1-22. -
További szerzők:Friedländer Elza (1980-) (biofizikus) Ibrahim, Shehu M. Brock Roland Szöllősi János (1953-) (biofizikus) Vereb György (1965-) (biofizikus, orvos)
Pályázati támogatás:K119690
OTKA
NN129371
OTKA
GINOP-2.3.2-15-2016-00050
GINOP
GINOP-2.3.3-15-2016-00003
GINOP
Internet cím:Szerző által megadott URL
DOI
Intézményi repozitóriumban (DEA) tárolt változat
Borító:
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