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. 2007 Apr 26;404(Pt 1):31–43. doi: 10.1042/BJ20061483

Figure 2. Ng binds to PA in vitro.

Figure 2

(A) PIP strips were blocked with 3% fatty acid-free BSA and incubated overnight with recombinant Ng or GAP-43 at 0.5 μg/ml. After extensive washing with TBST, anti-GAP-43 (797) or anti-Ng (756) affinity-purified antibodies were used to analyse protein content. The experiments were performed independently at least three times with similar results. A representative blot is shown. LPA, lysophosphatidic acid; LPC, lysophosphatidylcholine; PE, phosphatidylethanolamine; S1P, sphingosine 1-phosphate. (B) Hybond-C nitrocellulose strips were spotted with different amounts of egg-yolk PA and incubated with different concentrations of Ng. Ng–PA binding was quantified by densitometry of films exposed for different lengths of time and is expressed in arbitrary units. (C) The effect of ionic strength on Ng binding to PA was analysed by changing the NaCl concentration during overnight incubations with Ng as indicated. (D) The effect of pH on Ng binding to PA was analysed using a Tris/glycine buffer during overnight incubations with Ng. Results are means±S.E.M. for at least three independent experiments.