FIG. 2.

Immunoprecipitation of viral proteins from chimeric YF/JE viruses. (A) Parental or chimeric viruses were grown in LLC-MK₂ cells, labelled with [³⁵S]methionine, and harvested from the media at 48 to 60 h postinfection. Viral E proteins were immunoprecipitated from the media as described in Materials and Methods, and proteins were analyzed on an SDS–9% polyacrylamide gel. The E protein of YF5.2iv virus was immunoprecipitated with rabbit polyclonal antiserum against YF. The E proteins of the JE-S (JE SA₁₄-14-2), YF/JE-S, YF/JE-N, and JE-N viruses were immunoprecipitated with mouse hyperimmune ascitic fluid against JE virus. (B) Viral proteins produced in Vero cells. Proteins were labelled with [³⁵S]methionine for 6 h, and lysates were prepared at 48 h postinfection as described in Materials and Methods. Virus in the media was harvested and immunoprecipitated with hyperimmune ascitic fluid to either YF or JE, as described for panel A. Mock-infected lysates were immunoprecipitated with a mixture of YF and JE hyperimmune ascitic fluids. Proteins were analyzed on 13% SDS–polyacrylamide gels. Tunicamycin was added (+) or not added (−) at the time of labelling at a concentration of 7.5 μg/μl. Molecular mass markers, indicated by small lines in the right margin, represent 220, 97.4, 66, 46, 30, and 14.3 kDa, respectively.