A cell ELISA based method for exosome detection in diagnostic and therapeutic applications.

OBJECTIVE: Exosomes are nano-scaled carriers of miRNA, mRNA and proteins which are secreted from viable cells. Exosome detection within serum, saliva and semen offers diagnostic value for detection of various diseases including cancer. In the present study, we have lunched an in vitro study to develop a more efficient method for exosome detection. In this regard, the recombinant LAMP-DARPins (capable of Her2 targeting) gene was packed within the lentiviral particles and stably transferred into the HEK293T cells. The morphology and sizes of the obtained exosomes were characterized by TEM and zeta sizer. The expression of LAMP-DARPins antigen on the exosome surface was verified by western blotting. Ultimately, the efficiency of cell surface ELISA in suspension method was examined for exosome detection.

RESULTS: The exosome particles were successfully harvested and purified from transfected cells. The sizes of exosome particles were determined to be 90 nm using zeta sizer instrument. The TEM scan showed that the exosomes are cap like shaped and their sizes range between 40 and 150 nm. An observed 120 kDa band on western blotting paper indicated the LAMP-DARPins antigen expression on exosome surfaces. The results of cell surface ELISA in suspension method were superior to the results of conventional cell ELISA.

CONCLUSIONS: It could be concluded that the cell surface ELISA in suspension method could be an amenable method to detect exosome particles within the biological samples. Moreover, the method could be modified to evaluate the ability of exosomes to interact with target cells in both diagnostic and therapeutic applications.