FIG. 3.

E2 is a virion protein. (A) Resistance to detergent extraction. Purified MVs (WR strain) were extracted (+) with NP-40 or with NP-40 and DTT or mock treated (−) and separated into soluble (S) and pellet (P) fractions. Proteins in both fractions were resolved by SDS-PAGE and subjected to Western blotting with rabbit antibody to E2, A3, or L1 followed by anti-rabbit immunoglobulin G conjugated to horseradish peroxidase. (B) Distribution of E2 in MVs and EVs. RK₁₃ cells were infected with vIHD-J at 2 PFU per cell, and 48 h later MVs and EVs were purified from the cell lysate or medium, respectively, by sedimentation through two continuous 25-to-40% sucrose gradients, as described in Materials and Methods. The same amounts of total protein from both MV and EV samples were adjusted to a final volume of 15 μl with 1 mM Tris (pH 7.4). The proteins were separated by SDS-PAGE, followed by Western blotting with antibody to E2, F13, D8, or A26, as for panel A.