<p>(A) Immunodepletion of ∼38 kDa sTLR2 in breast milk (BM) described in materials and methods. Quantitative analysis using TLR2 ELISA indicated a significant (<i>P<0.05)</i> decrease in sTLR2-depleted (sTLR2-D) compared to mock-D breast milk (BM). Western blot analysis revealed that ∼38 kDa and ∼83 kDa sTLR2 were markedly reduced compared to mock-D BM. (B-D) 500 ng/mL Pam<sub>3</sub>CSK<sub>4</sub> was incubated with media or BM that was either mock-D or sTLR2-D for 1 hr at 37°C before being placed on cells. Supernatants were collected for IL-8 ELISA after 18 hours. Results represent (B) U937, (C) HEK293-TLR2 and (D) HEK293. (E) Commercial rsTLR2 was used at varying concentration with or without sCD14 and showed no inhibition of IL-8 or (F) IL-6 production in HEK293-TLR2 cells. Significant increases in pro-inflammatory cytokine, IL-8, was observed in sTLR2-D compared to mock-D BM. *<i>P<0.05,</i> **<i>P<0.01</i>, ***<i>P<0.001</i>. Errors bars, SEM. A representative data set from triplicate experiments is shown.</p
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