Interaction between CD9 and Aggrus. (A) HT1080 cells were stably transfected with V5-tagged, CD9-encoding vector (HT1080/CD9). Then, endogenous Aggrus was immunoprecipitated with an anti-Aggrus antibody in buffer containing 1% CHAPS. Exogenous CD9 was coimmunoprecipitated with the anti-Aggrus antibody. (B) CHO cells were transfected with expression plasmids encoding FLAG-tagged Aggrus and V5-tagged CD9. Then, the cells were solubilized with buffer containing 1% CHAPS, Brij 97, or Triton X. The FLAG-tagged Aggrus was immunoprecipitated with an anti-FLAG agarose. The immunoprecipitated proteins were electrophoresed and immunoblotted with an antibody to V5 tag or Aggrus. (C) LN319 cells were solubilized with buffer containing 1% CHAPS. Then, endogenous proteins were immunoprecipitated with control IgG or antibody to CD9 or Aggrus. The immunoprecipitants were electrophoresed and immunoblotted with an antibody to Aggrus or CD9. (D) LN319 cells were immunostained with antibodies to CD9 and Aggrus. Aggrus was visualized by incubation with Alexa Fluor 488–conjugated goat anti–rat IgG (left panel). CD9 was visualized by incubating with Alexa Fluor 633–conjugated goat anti–mouse IgG (middle panel). Merged image is shown in right panel.