To widen the application of nanofibers web in the field of medical health materials, a new Ag/amino-terminated hyperbranched polymer (HBP)/polyacrylonitrile (PAN) nanofiber web with excellent antimicrobial activity and filtration property was produced with Ag/HBP dispersion solution and PAN nanofiber. Ag/HBP dispersion solution was prepared with HBP as reducer and stabilizer, and Ag/HBP/PAN nanofiber was prepared by modifying electrospun PAN nanofiber with Ag/HBP aqueous solution. The characterization results showed that spherical Ag nanoparticles were prepared and they had a narrow distribution in HBP aqueous solution. The results of Ag/HBP/PANnanofiber characterized with SEM and EDS showed that the content of silver nanoparticles on the surface of PAN nanofiber was on the increase when the treating temperature rose. The bacterial reduction rates of HBP-treated PAN nanofiber against S. aureus and E. coli were about 89%, while those of the Ag/HBP/PANnanofiber against S. aureus and E. coli were 99.9% and 99.96%, respectively, due to the cooperative effects from the amino groups in HBP and Ag nanoparticles. Moreover, the small pores and high porosity in Ag/HBP/PAN nanofiber web resulted in high filtration efficiency (99.9%) for removing smaller particles (0.1 mu m similar to 0.7 mu m), which was much higher than that of the gauze mask.