Figure 4. Pull-down of membrane proteins... | Rockefeller University Press

Figure 4.

$Figure 4. Pull-down of membrane proteins using anti-GM130 antibodies. Golgi-containing microsomes from rat kidneys were partially purified using differential and density-gradient centrifugation (see Materials and methods). These fractions were incubated with anti-GM130 antibody, and the antibody–membrane complexes were captured with protein G–coated magnetic beads. (A) Significant amounts of GM130 were pulled down from the starting material with anti-GM130 but not with control antibody. (B) Significant amounts of γENaC were pulled down from the starting material with anti-GM130 but not with control antibody. Both full-length and cleaved forms were detected. (C) Na depletion increased the amount of cleaved γENaC in GM130-containing vesicles. The 55-kD bands represent the anti-GM130 IgG used for the pull-down. The results are representative of four independent experiments. Quantification is shown in Fig. 7.$

Pull-down of membrane proteins using anti-GM130 antibodies. Golgi-containing microsomes from rat kidneys were partially purified using differential and density-gradient centrifugation (see Materials and methods). These fractions were incubated with anti-GM130 antibody, and the antibody–membrane complexes were captured with protein G–coated magnetic beads. (A) Significant amounts of GM130 were pulled down from the starting material with anti-GM130 but not with control antibody. (B) Significant amounts of γENaC were pulled down from the starting material with anti-GM130 but not with control antibody. Both full-length and cleaved forms were detected. (C) Na depletion increased the amount of cleaved γENaC in GM130-containing vesicles. The 55-kD bands represent the anti-GM130 IgG used for the pull-down. The results are representative of four independent experiments. Quantification is shown in Fig. 7.