Human Epidermal Growth Factor Receptor 2 (HER2), also called ErbB2, HER2/neu, is a receptor tyrosine kinase and a crucial mediator for cell proliferation and differentiation in embryo and adult tissues as other members of epidermal growth factor receptor. HER2 Overexpression in 20 to 30% of cancers is related to poor prognosis and aggressive phenotype. Despite anti-Her2 based therapies (using Trastuzumab) have being used for years in patients with HER2 positive breast cancer, development of new binders for HER2 can be a critical need as mechanisms of resistance to trastuzumab have been observed.Single chain variable fragments or scFv antibodies offer some advantages over whole antibodies which makes them good candidate for cancer diagnosis or treatment. Small size, high affinity for a specific target, more rapid penetration to tumors, more rapid clearance from the blood, ability to be coupled with toxins, application in gene and drug delivery and specific tumor targeting are only some of its important advantages for cancer therapy.We have been using phage display technology routinely in our laboratory for selecvtion of specific single chain or single-domain antibodies for different targets. Recently, we used ribosome display technique for selection of specific scFv against extra cellular domain of HER2. Ribosome display is a powerfull in-vitro method for selection of specific binders against any target molecules. In comparison to phage display, ribosome display is a cell-free method that overcomes some of its limitation such as transformation steps which decrease library size. In addition, as selection is performed in a cell-free system, transcription and translation of toxic proteins is also possible without affecting growth rate of host. Ribosome display as a rapid and robust technique has been used first time by Hanes and Plueckthun (simultamously by He and Taussig) for affinity selection of single chain antibodies against variant peptide of the transcription factor GCN4 dimerization domain in 1998. Thereafter, it has been widely used for in-vitro selection of different functional molecules (polypeptides, scFvs, DARPins and .etc) against targets and also for affinity maturation of phage displayed selected binders. Since we recently produced targeted exosome against HER2 positive cancer cells (data under review) using genetically fusion of DARPin G3, as targeting moiety, to an exosomal surface protein (Lysosomal associated membrane protein, LAMP2b), we planned to select a specific single chain variable fragment antibody against HER2 and then use it for targeting of our exosomes toward HER2 positive cancer cells. Bioinformatic analysis of selected scFv interaction with extracellular domain of HER2 could help us better understanding of any changes in the interaction before and after the fusion.