The HMPV G protein facilitates virus-like particle (VLP) formation by interaction with the F and M proteins. Cells were transfected with (A) pCAGGS/G-FLAG, (B) pCAGGS/F-cmyc, (C) pCAGGS/M, (D) pCAGGS/F-cmyc and pCAGGS/G-FLAG, (E) pCAGGS/M and pCAGGS/G-FLAG, and (F) pCAGGS/F-cmyc and pCAGGS/M. HMPV protein expression in each transfection was confirmed by analysing the cell lysates by immunoblotting using the appropriate antibody (Total). The transfected cells were processed for VLP formation as described in methods and applied to the discontinuous gradient analysis (Gradient analysis). The material at the (1) 0-20% (w/v) sucrose, (2) 20-50% (w/v) sucrose and (3) 50-60% (w/v) sucrose interfaces was harvested and immunblotted using anti-FLAG, anti-cmyc or anti-M as indicated. High molecular mass G proteins are highlighted by “*”.