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Irini Bournazou, John D. Pound, Rodger Duffin, Stylianos Bournazos, Lynsey A. Melville, Simon B. Brown, Adriano G. Rossi, Christopher D. Gregory
Published in Volume 119, Issue 1
J Clin Invest. 2009; 119(1):20–32 doi:10.1172/JCI36226
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Figure 2
Biochemical characterization of the inhibitory factor.

Conditioned media from BL2 cells cultured for 24 hours were size fractionated using filters with 50 kDa (A) molecular weight cutoff sizes. Unfiltered medium was included as control. *P < 0.001 compared with the corresponding positive control. Error bars indicate SEM. Ion exchange analysis included the use of Q Sepharose beads (positively charged) in order to distinguish positively and negatively charged molecules in the <100 kDa fraction (B) of the BL medium. Unbound molecules (Q1 fraction) were collected, whereas bound molecules were eluted from the beads (Q2 fraction). Neutrophil migration toward these fractions in the presence of fMLP (100 nM) was assessed. Q1 and Q2 fractions (unbound and eluant fraction) of serum-free medium (no BL) were included as control. P < 0.05 compared with the corresponding control. Error bars indicate SEM. (C) Chemotaxis assay of neutrophils toward BL-conditioned medium that was heat inactivated (90°C for 10 minutes). (D) MALDI-TOF mass spectrum for the tryptic digest of the peptide band identified as lactoferrin.