Construction and surface expression of the F and G gene of HMPV isolate SIN06-NTU271. (A) pCAGGS/F-cmyc and pCAGGS/G-FLAG were surface-biotinylated and examined by immunoprecipitation with either anti-cmyc or anti-FLAG respectively. Each surface-labelled protein was treated with PNGasF (P), EndoH (E) or non-treated (N). Mock transfected pCAGGS (pc) vector was used to determine specificity of the two antibodies. (B) pCAGGS/F-cmyc transfected 293T cells were either non-treated or treated with 0.5 μg/ml TPCK-trypsin for 1 hr at 37°C. The cells were surface-biotinylated and immunoprecipitated (IP) using anti-cmyc (C) Cells transfected with pCAGGS/M and immunoblotted using anti-M. (D and E) Cells singly transfected with pCAGGS/F-cmyc (F), pCAGGS/G-FLAG (G) or pCAGGS (Mock) transfected 293T cells were immunoprecipitated (IP) with either anti-cmyc (cmyc) or anti-FLAG (FLAG) antibodies and Immunoblotted (IB) with (D) anti-cmyc or (E) anti-FLAG.