Figure 2

Co-localization of heparan sulfate, trastuzumab and HER2 in SKBR3, MCF7 and MCF7 HPSE1 cells by confocal immunofluorescence. 9.0x10³ cells were seeded on coverslips and treated with 25 μg/ml of trastuzumab for 72 hours. The cells were fixed with paraformaldehyde, washed with glycine and permeabilized with saponine. The antibodies anti-HS-FITC (represented in green), trastuzumab (stained with Alexa Fluor® 594 – represented in red), and anti-HER2 (stained with Alexa Fluor® 350 – represented in blue) were incubated with the cells as described in methods. Nuclei were stained with DAPI. Microscopy analyses were performed with a confocal laser scanning microscope equipped with a Plan-Apochromat × 40 objective under oil immersion (Zeiss, LSM 510 META). The images were processed using LSM 510 (Zeiss) and Image J (NIH, Bethesda, MD).