Figure 4

Intracellular and secreted forms of LASV GP2 variants and co-transfections with sGP1 in HEK-293T/17 cells. Cell extracts and supernatants from HEK-293T/17 transfected with LASV GP2 variants or from co-transfections with sGP1 were analyzed for the expression and secretion of GP1 and GP2. Ten micrograms of transfected cell protein were resolved on SDS-PAGE gels, blotted onto nitrocellulose membranes and probed with anti-LASV GP2 (panel A) or GP1 mAbs (panel B). Similarly, twenty μL of supernatant from each transfection were resolved on SDS-PAGE gels, blotted, and probed with the same anti-GP2 (panel C) and anti-GP1 mAbs (panel D). Control plasmid pcDNA3.1(+):intA (lane 1) was transfected alongside constructs sGP2-h HC (lanes 2), sGP2-hλLC (lanes 3), and sGP2-GPC SP (lane 4). The sGP2 constructs were co-transfected in the same order in lanes 5, 6, and 7 with equimolar amounts sGP1 expression plasmid. A transfection with sGP1 construct is shown in lane 8. Molecular weight markers with sizes (kDa) are shown to the left of the panel. Panels showing expression profile from cell extracts (C) and supernatants (S) are demarcated by vertical lines, along with the respective anti-LASV mAb probes. The co-transfection profiles with GP1 and GP2 constructs are indicated by (-) and (+) symbols at the bottom of the figure.