Identification and characterization of a novel scFv recognizing human and mouse CD133

CD133, also known as Prominin-1, is expressed on stem cells present in many tissues and tumors. In this work, we have identified and characterized a single-chain variable fragment (scFv) for the efficient and specific recognition of CD133. Phage display was used to develop the scFv from a previously reported anti-CD133 hybridoma clone 7, which was capable of recognizing both glycosylated and non-glycosylated forms of human CD133. The scFv immunostained CD133⁺ Caco-2 cells, but not CD133^-/low U87 cells. Significantly, it immunostained CD133^- cells transiently transfected with the mouse CD133 gene as well as CD133⁺ mouse cells. Co-immunostaining studies in mouse bone marrow cells, using anti-CD133 scFv-FITC and anti-mouse CD133-PE (clone 13A4) commercial antibody, indicated that the epitopes recognized by these reagents partially overlap. Taken together, these results suggest that the scFv can recognize mouse CD133 protein in addition to recognizing human CD133. This new scFv is expected to be valuable both as a molecular diagnostic reagent for identifying CD133⁺ cells and as a ligand for targeting therapeutics to CD133⁺ tumor-initiating cells.