Fig. 2

JSRV Env Mabs bind to human 293 cells expressing the JSRV Env protein. Human 293 cells were transiently transfected with the JSRV Env-expression plasmids pCIneoJenv, which contains the JSRV Env coding region cloned into the pCIneo expression plasmid (Promega, Madison, WI), or pSX2Jenv [2]. JSRV Env expression is driven by a CMV promoter in pCIneoJenv, and by a Moloney murine leukemia virus (MoMLV) promoter in pSX2Jenv. The 208F/LJenvSN cells are 208F rat fibroblasts transduced with the LJenvSN retroviral vector, which contains the JSRV Env coding region cloned into the LXSN retroviral vector [41], and Env is expressed from the MoMLV promoter in LXSN. For analysis, the adherent cells were suspended using EDTA (no trypsin), centrifuged onto slides, fixed with formaldehyde, and subjected to immunostaining. The secondary (2°) antibody was a goat anti-mouse IgG (H + L) conjugated to Alexa Fluor 488 (a green-fluorescent dye) obtained from Invitrogen. Env expression is detected in all of these cell types by the JSRV Env Mabs