Carcinoembryonic antigen (CEA) is a tumor-associated glycoprotein expressed in a variety of cancers. Detection of CEA on the tumor cell surface may be carried out using fluorescently labeled anti-CEA antibodies. In this work single-domain antibodies (sdAbs) are used for labeling CEA on tumor cells. SdAbs are compact antibody fragments (15 kDa), that are nearly 12 times smaller than full-size monoclonal antibodies (mAbs). SdAbs exhibit the same affinity and about the same variability as mAbs and have a number of important advantages for biological applications, such as high stability, solubility, and expression rate.
In this study, biotinylated anti-CEA sdAbs and mAbs have been used as capture molecules recognizing CEA on tumor cells in vitro. Detection of CEA was carried out through interaction of biotynilated Abs with streptavidin-quantum dots conjugates by means of flow cytometry. Compared to fluorescent organic dyes traditionally used for fluorescent labeling, quantum dots have a number of advantages, such as broad excitation spectra, narrow, sharp emission spectra and a long luminescence lifetime; they are nearly 20 times brighter and thousands of times more stable against photobleaching than organic dyes are.
In this work we demonstrated, that simultaneous use of compact and highly stable sdAbs with highly fluorescent quantum dots allows the sensitivity of detection of CEA on tumor cell surface to be considerably improved. CEA detection with the use of biotinylated sdAbs is more than 10 times more sensitive compared to the conventional mAbs analysis. The method developed in this study can be easily adapted for detection of various interesting targets in multiplexed studies.