Figure 3

The glycosphingolipid GM1 was detected in purified mature virions. Virions were purified by a two-step nycodenz gradient method. (A) Each fraction was analyzed by immunoblotting. Immunoblots of the proteins in each fraction (equal volume loaded) were labeled with anti-gQ1(a), -gQ2(b), -gO(c), -gL(d), -gB(e), -gH (f) or -IE1(g) MAb for viral protein, and anti-CD59 (h) or -LAT (i) MAb. GM1 (j) was detected by reaction with HRP-coupled cholera toxin. The numbers above each column represent the fraction from the bottom of the gradient. The fractions were collected from the bottom. GS indicates the extracts from GS-infected HSB-2 cells. GM1, but not CD59 or LAT was detected in the virion fractions. (B) PCR was performed for analysis of viral DNA in the fractions. The fractions 7–9 contained viral DNA.