Fig 1: Foxp3 expressing T cells have elevated lipid droplet content. (A) Quantitation using Imagestream, of LDs with nile red staining, and palmitate uptake using BODIPY-labeled palmitate on resting and activated Foxp3+ and Foxp3- T cells from the same activation cultures from C57BL/6.Foxp3 hCD2/CD52 knock in mice. The bar chart shows the composite data from 8 separate experiments. *p < 0.05 by ANOVA. (B) Measurement using Imagestream, of LDs and fluorescent palmitate uptake in resting, and activated CD4+Foxp3+ICOS+/- cells from the same cultures. (C) Top panels; Measurement of LD number in CD4+Foxp3+ and Foxp3- cells using the Imagestream “spot count” feature. Bottom panels; Measurement of LD number in CD4+Foxp3+ICOS+ and ICOS- cells. (D) Representative images of Foxp3- and Foxp3+ cells showing bright field, Hoescht, Foxp3, and nile red staining analyzed with Imagestream. (E) Measurement of proximity (“Similarity” feature in Imagestream) of nile red staining and perilipin, histogram and bottom image panel, compared to nile red and BODIPY-palmitate, histogram and upper image panel, in NIH3T3 cells. (F) Measurement of LD by nile red staining in a tamoxifen-inducible Foxp3 T cell line EL4.cFoxp3 or the parental cell line EL4 in the presence and absence of 4'OH tamoxifen. Error bars represent standard error of the mean, *p < 0.05. Representative of three separate experiments. (G) Expression of DGAT1 and DGAT2 protein (top panels) and mRNA transcripts in Foxp3+ T cells and Tconv from B6.KI mice. For mRNA analysis cells were analyzed straight after isolation or after 24 h activation with anti-CD3/28 beads. Error bars represent standard error of the mean, *p < 0.05. Representative of three separate experiments.
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