Fig. 5.

Sera from Prdm1^fl/flFoxp3^YFP-Cre tumor-bearing mice increase the anti-tumor activity of macrophages. a-b) B16-OVA model was established as in Fig. 2c. IF staining of IgE and CD68 in the tumor and quantitation of IgE⁺CD68⁺ and IgE⁺CD68⁻ cells (a); or IF staining of FcεR1α and CD68 in the tumor (b). Each dot represents the counted numbers within a field of view (160 ×). WT: n = 15 views from 6 mice; KO: n = 14 views from 6 mice. c-d) BMDMs were co-cultured with CTV-labelled B16-OVA cells treated with or without a same volume of sera (2.5 μl, 1.25% of the total culture) collected from WT or Prdm1^fl/flFoxp3^YFP-Cre (KO) tumor-bearing mice for 2 h (c) or overnight (d) in quadruplicates. In a group, sera were pre-treated with anti-IgE (α-IgE). c) BMDMs with phagocytosed B16-OVA cells were indicated as CTV⁺F4/80⁺ cells (upper left, representative F4/80⁺ plot; lower left, representative histogram of CTV expression gated on F4/80⁺ cells; right, quantitation). d) Percent tumor killing is shown after quantifying dead tumor cells (CTV⁺ cells positive for viability-dye). Tumor cells alone treated with sera were included as controls. Triangles, tumor cells and BMDMs with no sera added. ns, no significance, * P < 0.05, ** P < 0.01, *** P < 0.001 and **** P < 0.0001 (a,c,d, unpaired two-tailed Student’s t-test). Bars, mean ± SEM. e) The correlation of CD86 and FCER1A expression in top 25% CD68^hi SKCM patients from the TCGA dataset (n = 110) was analyzed by Pearson correlation (two-tailed, no adjustment for multiple comparisons because of one correlation test for a gene pair). The values of the coefficients (r) and significance (p) are indicated. f) Kaplan-Meier analysis of OS of patient cohorts expressing differential FceM1 signature (top 33% vs bottom 33%) based on combined log-averaging of CD68, CD86 and FCER1A transcript levels from the TCGA-SKCM dataset. P value is generated using two-tailed LogRank test. Median, median survival time