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. 2011 Jan 6;286(11):8752–8758. doi: 10.1074/jbc.M110.186833

FIGURE 6.

FIGURE 6.

PrPC-G-lyso-PI reduced the co-precipitation of cPLA2 with PrPSc. A, membrane extracts from ScN2a cells that had been treated for 3 h with control medium (■) or 25 ng of PrPC-G-lyso-PI (□) were separated by centrifugation on a sucrose density gradient, and the amount of cPLA2 in each fraction was determined by ELISA. Values shown are the mean amount of cPLA2 (units) ± S.D. from an experiment run in triplicate. Cell extracts were prepared from control ScN2a cells (fraction 1) or ScN2a cells that had been pretreated with 25 ng of PrPC-G-lyso-PI (fraction 2). Membranes were isolated and incubated with either mAb ICSM35 (anti-PrP) or mAb CH-7 (anti-cPLA2) and protein G magnetic beads. Immunoprecipitates were prepared (± proteinase K digestion), separated by PAGE, and transferred to membranes. The presence of PrPSc was detected by mAb ICSM18, and cPLA2 was detected by mAb CH-7. Immunoblots show the amount of PrPSc (B) and cPLA2 (C) precipitated by mAb ICSM35 (anti-PrP) or the amount of PrPSc (D) and cPLA2 (E) precipitated by mAb CH-7 (anti-cPLA2).