CF750-A33scFv-Fc-Based Optical Imaging of Subcutaneous and Orthotopic Xenografts of GPA33-Positive Colorectal Cancer in Mice

Figure 2

Immunoreactivity and specificity of A33scFv-Fc. (a) Cell binding assays of FITC-labeled A33scFv-Fc. GPA33-positive (LS174T, COLO205) and GPA33-negative (SMMC7721, AGS) cells were incubated with FITC-A33scFv-Fc, followed by flow cytometric analysis. (b) Immunofluorescence chemical assay for the tissue binding of FITC-A33scFv-Fc. LS174T, COLO205 tumor tissues, and liver tissues were stained with FITC-A33scFv-Fc and observed under a fluorescence microscope. The cell nuclei were stained with DAPI, and an isotype antibody was used as a negative control. (c) The cell binding of ¹³¹I-labeled A33scFv-Fc. LS174T cells were incubated with ¹³¹I-A33scFv-Fc at different concentrations (1~60 nM), followed by counting the radioactivity of cells using a gamma counter. The mixture containing the same amount of unconjugated ¹³¹I and A33scFv-Fc was used as a control. (d) Cellular binding of CF750-labeled A33scFv-Fc. SMMC-7721 and COLO205 cells were incubated with the CF750-A33scFv-Fc at the indicated molar concentrations (0~20 nM) and followed by scanning with the IVIS optical imaging system. The fluorescence intensities of the cells were calculated and compared.

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