Fig. 1. Monoclonal antibody 2C11 recognises PtdIns(4,5)P2 inserted in a membrane bilayer. (A) 2C11 recognises PtdIns(4,5)P2 but not DNA or RNA on dot-blot. Serial dilutions of PtdIns(4,5)P2 (0.0125-0.4 µg), DNA and total PC12 cell RNA (0.03-1 µg) were spotted on membranes and probed with 2C11 antibody. (B) 2C11 specifically interacts with PtdIns(4,5)P2-containing liposomes. Liposomes containing radioactive PC as a tracer and either PtdIns (empty bars) or PtdIns(4,5)P2 (filled bars) were incubated with immobilised anti-IgM or 2C11. Radioactivity associated with beads was measured and results expressed as a percentage of the total. Error bars represent s.d. based on three experiments. (C,D) Cryosections of extruded liposomes containing PtdIns(4,5)P2 (C) and PtdIns (D) were probed with 2C11 followed by gold-conjugated secondary antibodies. 34% of the total PtdIns(4,5)P2-containing liposomes (n=170) were labelled with one or more gold particles compared with 0% (n=96) in the case of PtdIns-containing liposomes. Bars, 50 nm.
