Internalization analysis of Cal-CD19-liposomes by confocal laser-scanning microscopy. SUP-B15 cells were incubated with Cal-CD19-liposomes at 37° C for the indicated times. After washing and fixation, cells were allowed to adhere to glass slides using Cytospin 3. Thereafter, cells were stained with rhodamine phalloidin to visualize cytoplasmic actin. The prepared slides were analyzed by confocal laser-scanning microscopy. The images were acquired using a Radiance 2100 Confocal and Multi-photon Imaging System (Bio-Rad) with an Axioshop 2 plus microscope (Carl Zeiss, Jena, Germany). The type of the objective lens was Plan-Apochromat (Carl Zeiss) with a numerical aperture of 1.40. The acquisition and subsequent processing software were Laser Sharp 2000 (Bio-Rad) and Adobe Photoshop 5.0 (Adobe Systems, San Jose, CA), respectively. Original magnification × 630. Green fluorescence of calcein (left column), red fluorescence of rhodamine phalloidin (middle column), and the merged image (right column) are shown. In this assay, internalization of the Cal-CD19-liposomes will result in a yellow signal in the merged image.