Fig. 2
From: EpCAM as multi-tumour target for near-infrared fluorescence guided surgery
Conjugation, serum stability and EpCAM specificity: Conjugation of IRDye 800CW to 323/A3 and MOPC21 was done through NHS ester conjugation. A mean labeling ratio of 2.6 ± 0.98 and 2.8 ± 0.99 was obtained for 323/A3 and MOPC21 respectively a. EpCAM specificity of 323/A3 was confirmed after conjugation of IRDye 800CW on various cell types, corrected for cell number using the 800/700 nm ratio b. Experiments were done in human colon (HT29-luc2, COLO320), breast (MCF-7-luc2-cGFP, MDA-MB231) and head & neck (OSC-19-luc2-cGFP, FaDu-luc2) cancer. Cells were incubated with different concentrations of 323/A3-800CW or an isotype normal mouse IgG (MOPC21) conjugated to 800CW. To correct for the number of cells, the ratio of NIR fluorescence and the fluorescence intensity of TO-PRO-3 was plotted. 323/A3-800CW showed to be stable in human serum with more than 60% of the conjugate still free to bind EpCAM after 96 h with the remaining 40% aggregated or bound to albumin c