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. 2018 Oct 17;3:84. Originally published 2018 Jul 17. [Version 2] doi: 10.12688/wellcomeopenres.14493.2

Figure 2. IL-15 mediated trans-signaling in CD4+ T cells requires TCR engagement.

Figure 2.

( A) Representative TIRFM images of CD4+ T cells incubated on glass-supported planar bilayers containing ICAM-1, CD80, OVA 323-339/I-A b and IL-15/IL-15Rα, as indicated. After 30 min incubation at 37°C in HBS/HSA buffer, cells were fixed and stained for TCR (red, Alexa Fluor 568) and phospho-STAT5 (pSTAT5) (green, Alexa Fluor 488) in PBS buffer. White arrows in the image panels indicate central accumulation of TCR at the T cells IS. ( B) Quantification of pSTAT5 fluorescence in A. Data are presented as percentage increase in pSTAT5 fluorescence, relative to unstimulated controls. Data are from 2 independent experiments (N=32-57) (mean ± S.E.M). P values, one-way ANOVA. Imaging was performed on a Nikon Ti microscope with a 100x TIRF objective, N.A. 1.49, controlled by Nikon Elements software. Fluorescence images were captured using an Ixon cooled EMCCD camera (512 x 512 pixels, Andor Technology). Mean fluorescence intensity at contact interfaces was quantified from 14 bit images using Metamorph software. Brightness and contrast are adjusted uniformly across image groups for clarity.