Blood samples were filtered, fixed, permeabilized and washed, CTC identification by fluorescent enumeration was done as previously described6 (link), 17 (link). Filters were washed with PBS to remove unbound antibody, placed onto a microscope slide with Fluoromount-G/DAPI (Southern Biotech) and sealed with a glass cover slip. An Olympus BX54WI Fluorescent microscope with Carl Zeiss AxioCam was used to image the samples. Exposures were preset as 2–5 sec (Cyanine5), 2 sec (PE), 100–750 msec (FITC), and 10–50 msec (DAPI) for equal signal comparisons between cells. A Zen2011 Blue (Carl Zeiss) was used to process the images.