As key elements and targets for various intracellular pathogens, macrophages play an essential role in host defense. Although ovine peripheral blood monocyte-derived cell lines have been established, their phenotypic characteristics and functional properties remain unknown. We have established several ovine macrophage cell lines from peripheral blood adherent cells that can proliferate spontaneously in long-term culture in vitro. Characteristics of macrophages were shaped by cell morphology, cell adhesion, expression of cell surface markers, phagocytic activity and inflammatory response. Furthermore, the differences of genes expression (such as membrane proteins, pro-inflammatory cytokines and chemokine) were compared between blood macrophages (BMs) and alveolar macrophages (AMs), and between BMs and splenic macrophages (SMs), respectively. The expression of membrane genes (CD11b and CD80), pro-inflammatory cytokines (IL-1β and TNFα) and chemokines (IL-8/CXCL8 and CCL-21) was lower than that in AMs or SMs, but not CD200. Moreover, BMs maintained lower expression level of M1 macrophage related genes (iNOS and IDO), but high expression levels of M2 macrophage related genes (ARG2 and TGFβ1). BMs showed lower phagocytic ability than AMs and SMs. Compared with AMs, BMs showed higher salmonella proliferation rate within cells. Collectively, BMs could suppress inflammatory responses and possessed partly phenotypic characteristics of M2 macrophages. Copyright © 2018 Elsevier B.V. All rights reserved.