Abstract Both pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs) activate antigen-presenting cells, often through the same pattern recognition receptors (PRR), such as Toll-like receptors (TLR). The TLR4–CD14–MD2 and TLR2–CD14 complexes have been shown to play a role in the recognition of lipopolysaccharide (LPS) and peptidoglycan (PG), respectively. Since many DAMPs have also been known to activate TLR2 or TLR4 pathways, we dissected the role of each molecule in the receptor complexes (TLR2–D14–MD2) responding to DAMP (necrotic cells) or PAMP (LPS and PG). CD14 played a significant role in the activation of NF-κB in response to necrotic cells in the presence or absence of TLR2. However, MD2 did not play a significant role in NF-κB activation by necrotic cells. Intriguingly, MD2 did play a significant role in activating NF-κB by PG in the presence of TLR2–CD14. Compared with CD14 pos B6 mice, CD14 neg B6 mice showed delayed production of IL12p40 in response to necrotic cells in vivo. Microarray analysis showed that various pro-inflammatory genes of peritoneal cells were regulated in response to necrotic cells, in a CD14-dependent manner. The CD14 appears to recognize necrotic cells in addition to LPS, PG, apoptotic cells, and lipids, suggesting that CD14 might be a universal adaptor for DAMP and PAMP. On the contrary, MD2 recognizes only exogenous PAMP, when complexed with TLR2–CD14 or TLR4–CD14. Taken together, MD2 appears to discriminate between DAMP and PAMP.