Fig 1: Human malignant ascites supernatant suppresses CD206 BiTE activity but not FRβ BiTE activity. a CFSE-stained MDMs were co-cultured with T cells (10:1 E:T ratio) and the indicated BiTEs, in the presence or absence of 50% ascitic supernatant from three patients (Patients 1, 2 and 5). 96 h later, percentage live MDMs was determined with propidium iodide staining and a Celigo image cytometer. b T cell activation was assessed by flow cytometric analysis of CD25 expression after 96 h of co-culture with MDMs and the indicated BiTEs, in the presence or absence of 50% ascitic supernatant from three patients (Patients 1, 2 and 5). c Quantities of IL-6, IL-10 and total (active and latent) TGF-β in malignant ascites fluid from six different patients, as determined by enzyme-linked immunosorbent assay. Normal serum (NS) pooled from three healthy donors was included as a control. d Quantities of soluble CD206 in malignant ascites fluid from nine different patients was determined by enzyme-linked immunosorbent assay. Pooled NS was used as a control. Each condition was measured in biological triplicate and represented as mean ± SD (a-d). Statistical significance was assessed by one-way ANOVA with Dunnett’s post-hoc analysis compared with “Pooled NS” (c, d), or two-way ANOVA followed by Bonferroni post-hoc analysis, with each treatment being compared to the “Mock” condition within the relevant group (a and b). (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, non-significant)