Figure 4.

Western-blot analysis of various membrane fractions with anti-At-ACA8p polyclonal antibody. The fusion protein between GST and the first 122 amino acids of At-ACA8p (lane 1, 0.2 μg), the EDTA-eluted fraction of the Ca²⁺-ATPase purification procedure (lane 2, 1 μg), and proteins (50 μg) from various membrane fractions of the two-phase partitioning (lane 3, microsomal fraction; lane 4, first lower phase; lane 5, second upper phase) were separated by SDS-PAGE and blotted onto 0.45 μm of nitrocellulose. Immunodecoration was performed with an antiserum raised against At-ACA8 (Val-17–Thr-31) as described in “Materials and Methods.” No signal was detected when the preimmune serum was used.