Fig. 3. CEACAM1-L transfection of human M2 and A7 melanoma cells. M2 and A7 cells were stably transfected with human CEACAM1-4L cDNA and cloned by limiting dilution, resulting in the cell lines M2-CC1 and A7-CC1. (A) FACS analysis of M2, A7, M2-CC1 and A7-CC1 cells using human CEACAM1 reactive mAb 4/3/17 (bold lines) or an Ig-control (gray area). (B) Western blot of equal amounts of M2, A7, M2-CC1 and A7-CC1 cell lysates, developed with FLNa-specific mAb FLMN01 and human CEACAM1 reactive mAb 4/3/17. (C) RT-PCR analysis of M2, A7, M2-CC1 and A7-CC1 cells using CEACAM1-specific primers. (D) Cell lysates of A7-CC1 cells were immunoprecipitated with human CEACAM1 reactive mAb 4/3/17 or an Ig-control. CEACAM1 was detected with mAb 4/3/17. Co-immunoprecipitated FLNa was detected with mAb FLMN01. A7-CC1 cells were grown sparsely (E) or were allowed to adhere for 1 hour to laminin-1 (F) and double-stained for CEACAM1 (mAb 4/3/17, red) and FLNa (mAb FLMN01, green) and visualized by confocal microscopy. Co-localization is shown in the merged images (yellow). The white lines in the merged images show the courses of the Z-scans. Bars, 5 µm.