Imaging Membrane Curvature inside a FcεRI-Centric Synapse in RBL-2H3 Cells Using TIRF Microscopy with Polarized Excitation

Total internal reflection fluorescence microscopy with polarized excitation (P-TIRF) can be used to image nanoscale curvature phenomena in live cells. We used P-TIRF to visualize rat basophilic leukemia cells (RBL-2H3 cells) primed with fluorescent anti-dinitrophenyl (anti-DNP) immunoglobulin E (IgE) coming into contact with a supported lipid bilayer containing mobile, monovalent DNP, modeling an immunological synapse. The spatial relationship of the IgE-bound high affinity IgE receptor (FcεRI) to the ratio image of P-polarized excitation and S-polarized excitation was analyzed. These studies help correlate the dynamics of cell surface molecules with the mechanical properties of the plasma membrane during synapse formation.

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Imaging Membrane Curvature inside a Model Immunological Synapse in RBL-2H3 Cells using TIRF Microscopy with Polarized Excitation

10.1101/608463 ◽

2019 ◽

Author(s):

Rosa Machado ◽

Justin Bendesky ◽

Madison Brown ◽

Kathrin Spendier ◽

Guy M. Hagen

Keyword(s):

Total Internal Reflection ◽

Synapse Formation ◽

Immunological Synapse ◽

Membrane Curvature ◽

Leukemia Cells ◽

Live Cells ◽

Supported Lipid Bilayer ◽

Surface Molecules ◽

Rat Basophilic Leukemia Cells ◽

Basophilic Leukemia

AbstractTotal internal reflection fluorescence microscopy with polarized excitation (P-TIRF) can be used to image nanoscale curvature phenomena in live cells. We used P-TIRF to visualize rat basophilic leukemia cells (RBL-2H3 cells) coming into contact with a supported lipid bilayer, modeling an immunological synapse. These studies help correlate the dynamics of cell surface molecules with the mechanical properties of the plasma membrane during synapse formation.

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TIRF Microscope Image Sequences of Fluorescent IgE-FcεRI Receptor Complexes inside a FcεRI-Centric Synapse in RBL-2H3 Cells

Data ◽

10.3390/data4030111 ◽

2019 ◽

Vol 4 (3) ◽

pp. 111

Author(s):

Rachel Drawbond ◽

Kathrin Spendier

Keyword(s):

Immunological Synapse ◽

Immunoglobulin E ◽

Image Sequences ◽

Live Cells ◽

Background Intensity ◽

Pixel Intensity ◽

Microscope Image ◽

Signaling Complex ◽

Receptor Complexes ◽

Average Pixel Intensity

Total internal reflection fluorescence (TIRF) microscope image sequences are commonly used to study receptors in live cells. The dataset presented herein facilitates the study of the IgE-FcεRI receptor signaling complex (IgE-RC) in rat basophilic leukemia (RBL-2H3) cells coming into contact with a supported lipid bilayer with 25 mol% N-dinitrophenyl-aminocaproyl phosphatidylethanolamine, modeling an immunological synapse. TIRF microscopy was used to image IgE-RCs within this FcεRI-centric synapse by loading RBL-2H3 cells with fluorescent anti-dinitrophenyl (anti-DNP) immunoglobulin E (IgE) in suspension for 24 h. Fluorescent anti-DNP IgE (IgE488) concentrations of this suspension increased from 10% to 100% and corresponding non-fluorescent anti-DNP IgE concentrations decreased from 90% to 0%. After the removal of unbound anti-DNP IgE, multiple image sequences were taken for each of these ten conditions. Prior to imaging, anti-DNP IgE-primed RBL-2H3 cells were either kept for a few minutes, for about 30 min, or for about one hour in Hanks buffer. The dataset contains 482 RBL-2H3 model synapse image stacks, dark images to correct for background intensity, and TIRF illumination profile images to correct for non-uniform TIRF illumination. After background subtraction, non-uniform illumination correction, and conversion of pixel units from analog-to-digital units to photo electrons, the average pixel intensity was calculated. The average pixel intensity within FcεRI-centric synapses for all three Hanks buffer conditions increased linearly at a rate of 0.42 ± 0.02 photo electrons per pixel per % IgE488 in suspension. RBL-2H3 cell degranulation was tested by detecting β-hexosaminidase activity. Prolonged RBL-2H3 cell exposure to Hanks buffer inhibited exocytosis in RBL-2H3 cells.

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Ultrastructural evidence for mast cell-macrophage interrelationships in the dura mater of the rat: Infrequent mast cell-nerve associations

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100159308 ◽

1990 ◽

Vol 48 (3) ◽

pp. 352-353

Author(s):

Ruth V.W. Dimlich

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Dura Mater ◽

Spatial Relationship ◽

Potassium Phosphate ◽

Plasma Extravasation ◽

Close Spatial Relationship ◽

Male Wistar Rats ◽

Headache Pain ◽

Relationship Of

Mast cells in the dura mater of the rat may play a role in cerebral pathologies including neurogenic inflammation (vasodilation; plasma extravasation) and headache pain . As has been suggested for other tissues, dural mast cells may exhibit a close spatial relationship to nerves. There has been no detailed ultrastructural description of mast cells in this tissue; therefore, the goals of this study were to provide this analysis and to determine the spatial relationship of mast cells to nerves and other components of the dura mater in the rat.Four adult anesthetized male Wistar rats (290-400 g) were fixed by perfusion through the heart with 2% glutaraldehyde and 2.8% paraformaldehyde in a potassium phosphate buffer (pH 7.4) for 30 min. The head of each rat was removed and stored in fixative for a minimum of 24 h at which time the dural coverings were removed and dissected into samples that included the middle meningeal vasculature. Samples were routinely processed and flat embedded in LX 112. Thick (1 um) sections from a minimum of 3 blocks per rat were stained with toluidine blue (0.5% aqueous).

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Faculty Opinions recommendation of T cell receptor signaling precedes immunological synapse formation.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1004974.58104 ◽

2002 ◽

Author(s):

Andre Veillette

Keyword(s):

T Cell ◽

T Cell Receptor ◽

Synapse Formation ◽

Immunological Synapse ◽

Cell Receptor ◽

Receptor Signaling ◽

T Cell Receptor Signaling ◽

Cell Receptor Signaling

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Faculty Opinions recommendation of A network of PDZ-containing proteins regulates T cell polarity and morphology during migration and immunological synapse formation.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1027011.367938 ◽

2006 ◽

Author(s):

Sachdev Sidhu

Keyword(s):

T Cell ◽

Cell Polarity ◽

Synapse Formation ◽

Immunological Synapse

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Faculty Opinions recommendation of Mechanisms for segregating T cell receptor and adhesion molecules during immunological synapse formation in Jurkat T cells.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.1098808.554944 ◽

2008 ◽

Author(s):

Deborah Leckband

Keyword(s):

T Cells ◽

T Cell ◽

Adhesion Molecules ◽

T Cell Receptor ◽

Synapse Formation ◽

Immunological Synapse ◽

Cell Receptor ◽

Jurkat T Cells

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Geo-Spatial Analysis of Population Density and Annual Income to Identify Large-Scale Socio-Demographic Disparities

ISPRS International Journal of Geo-Information ◽

10.3390/ijgi10070432 ◽

2021 ◽

Vol 10 (7) ◽

pp. 432

Author(s):

Nicolai Moos ◽

Carsten Juergens ◽

Andreas P. Redecker

Keyword(s):

Population Density ◽

Large Scale ◽

Methodological Approach ◽

Spatial Relationship ◽

Postal Code ◽

Annual Income ◽

Choropleth Maps ◽

Clustering Approach ◽

Relationship Of ◽

Insight Into

This paper describes a methodological approach that is able to analyse socio-demographic and -economic data in large-scale spatial detail. Based on the two variables, population density and annual income, one investigates the spatial relationship of these variables to identify locations of imbalance or disparities assisted by bivariate choropleth maps. The aim is to gain a deeper insight into spatial components of socioeconomic nexuses, such as the relationships between the two variables, especially for high-resolution spatial units. The used methodology is able to assist political decision-making, target group advertising in the field of geo-marketing and for the site searches of new shop locations, as well as further socioeconomic research and urban planning. The developed methodology was tested in a national case study in Germany and is easily transferrable to other countries with comparable datasets. The analysis was carried out utilising data about population density and average annual income linked to spatially referenced polygons of postal codes. These were disaggregated initially via a readapted three-class dasymetric mapping approach and allocated to large-scale city block polygons. Univariate and bivariate choropleth maps generated from the resulting datasets were then used to identify and compare spatial economic disparities for a study area in North Rhine-Westphalia (NRW), Germany. Subsequently, based on these variables, a multivariate clustering approach was conducted for a demonstration area in Dortmund. In the result, it was obvious that the spatially disaggregated data allow more detailed insight into spatial patterns of socioeconomic attributes than the coarser data related to postal code polygons.

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Intrasellar Intercarotid Communicating Artery Associated with Agenesis of the Right Internal Carotid Artery: Case Report

Neurosurgery ◽

10.1227/00006123-198812000-00019 ◽

1988 ◽

Vol 23 (6) ◽

pp. 770-773 ◽

Cited By ~ 6

Author(s):

Masahiko Udzura ◽

Hiroo Kobayashi ◽

Yoshio Taguchi ◽

Hiroaki Sekino

Keyword(s):

Magnetic Resonance Imaging ◽

Carotid Artery ◽

Internal Carotid Artery ◽

Internal Carotid ◽

Spatial Relationship ◽

Resonance Imaging ◽

Anomalous Artery ◽

Magnetic Resonance Imaging Mri ◽

The Right ◽

Relationship Of

Abstract A 54-year-old man with a right hemiparesis was found to have an intrasellar intercarotid communicating artery associated with agenesis of the right internal carotid artery. Magnetic resonance imaging (MRI) studies demonstrated the spatial relationship of the anomalous artery to the surrounding structures, thus suggesting an embryonic enlargement of the capsular artery as a source of this anomalous artery.

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Immunoglobulin E mediated membrane conductance changes in rat basophilic leukemia cells

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y88-056 ◽

1988 ◽

Vol 66 (3) ◽

pp. 328-331 ◽

Cited By ~ 3

Author(s):

Carlos Barajas-López ◽

Jan D. Huizinga

Keyword(s):

Immunoglobulin E ◽

Calcium Influx ◽

Membrane Conductance ◽

Membrane Resistance ◽

Leukemia Cells ◽

Rat Basophilic Leukemia Cells ◽

Conductance Changes ◽

Electrophysiological Effects ◽

Basophilic Leukemia ◽

Stimulation Of

Electrophysiological effects of anaphylactic stimulation of rat basophilic leukemia cells (RBL-2H3) were studied using conventional microelectrodes. Stimulation of passively sensitized cells by anti-immunoglobulin E resulted in hyperpolarization followed by depolarization. These changes in membrane polarization were associated with a decrease in input membrane resistance. No effect of anaphylactic stimulation was seen in Ca2+-free solution or when Ca2+ influx was blocked by Co2+, but it was mimicked by the Ca2+ ionophore A-23187. This suggests that the changes in ionic conductances were associated with calcium influx. These results support the hypothesis that membrane conductance changes are involved in the stimulus-secretion process of the RBL-2H3 cells.

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