Figure 2.

Apparent loss of intermediate monocytes in response to LPS stimulation ex vivo. (A) Representative flow cytometry plots showing monocyte subsets in healthy controls incubated for 3 h in medium (B) or LPS. (C) Intermediate monocytes as a proportion of all monocytes sampled from healthy controls were reduced after 3 h incubation with LPS compared with medium alone (p < 0.0001). (D) Viability of healthy control PBMCs sampled following 24 h incubation with medium and LPS by mammalian LIVE/DEAD™ viability/cytotoxicity kit (Invitrogen™), showing no increase in cell death with LPS (ns = not statistically significant). (E) No difference in proportion of intermediate monocytes from healthy controls following treatment with LPS when using standard tissue culture or ultra-low bind plates. (F) Intracellular staining for CD14 and (G) CD16 in monocytes was not increased following stimulation with LPS for 3 h. (H) Stimulation of magnetically-isolated healthy control monocytes with LPS for 3 h resulted in loss of cells from the intermediate monocyte gate (p = 0.031). (I) Intermediate monocytes were reduced in whole blood sampled from healthy donors incubated with LPS-stimulated for 3 h compared to medium alone (p = 0.004).