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

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  • Rachel Drawbond

    (UCCS Center of the Biofrontiers Institute, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA
    Department of Mathematics, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA)

  • Kathrin Spendier

    (Department of Mathematics, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA
    Department of Physics and Energy Science, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA)

Abstract

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 (IgE 488 ) 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 % IgE 488 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.

Suggested Citation

  • Rachel Drawbond & Kathrin Spendier, 2019. "TIRF Microscope Image Sequences of Fluorescent IgE-FcεRI Receptor Complexes inside a FcεRI-Centric Synapse in RBL-2H3 Cells," Data, MDPI, vol. 4(3), pages 1-11, July.
    • Handle: RePEc:gam:jdataj:v:4:y:2019:i:3:p:111-:d:252489
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    1. Kenkre, V.M. & Spendier, K., 2022. "A theory of coalescence of signaling receptor clusters in immune cells," Physica A: Statistical Mechanics and its Applications, Elsevier, vol. 602(C).

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