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

1
UCCS Center of the Biofrontiers Institute, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA
2
Department of Mathematics, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA
3
Department of Physics and Energy Science, University of Colorado at Colorado Springs, Colorado Springs, CO 80918, USA
*
Author to whom correspondence should be addressed.
Data 2019, 4(3), 111; https://doi.org/10.3390/data4030111
Received: 23 May 2019 / Revised: 23 July 2019 / Accepted: 24 July 2019 / Published: 28 July 2019
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. View Full-Text
Keywords: total internal reflection fluorescence microscopy; TIRF; rat basophilic leukemia cells; RBL-2H3; IgE receptor; FcεRI; plasma membrane; supported lipid bilayer total internal reflection fluorescence microscopy; TIRF; rat basophilic leukemia cells; RBL-2H3; IgE receptor; FcεRI; plasma membrane; supported lipid bilayer
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Drawbond, R.; Spendier, K. TIRF Microscope Image Sequences of Fluorescent IgE-FcεRI Receptor Complexes inside a FcεRI-Centric Synapse in RBL-2H3 Cells. Data 2019, 4, 111.

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