This paper, for the first time, reports the quantitative measurement of piezoelectricity of electrospun silk nanofiber. The electrospinning conditions were optimized to yield uniform nanofibers with high degree of B -sheet content. The piezoelectric coefficient of the electrospun silk nanofiber was estimated by PFM measurements and the average d 31 value of silk nanofiber was estimated as 39 ± 2 pm/v. The piezoelectric measurements were repeated on several samples to estimate the repeatability of piezoresponse. This could open up numerous possibilities to design and develop a wide variety of sensors exploring the piezoelectric property of silk nanofiber. © 2018 IEEE.