Influenza virus neuraminidase (NA) targeting antibodies are an independent correlate of protection against infection. Antibodies against the NA act by blocking enzymatic activity, preventing virus release and transmission. As we advance the development of improved influenza virus vaccines that incorporate standard amounts of NA antigen, it is important to identify the antigenic targets of human monoclonal antibodies (mAbs). Additionally, it is important to understand how escape from mAbs changes viral fitness. Here, we describe escape mutants generated by serial passage of A/Netherlands/602/2009 (H1N1) in the presence of human anti-N1 mAbs. We observed escape mutations on the N1 protein around the enzymatic site (S364N, N369T and R430Q) and also detected escape mutations located on the sides and bottom of the NA (N88D, N270D and Q313K/R). We found that a majority of escape mutant viruses had increased fitness in vitro but not in vivo. This work increases our understanding of how human antibody responses target the N1 protein. ImportanceAs improved influenza virus vaccines are being developed, the influenza virus neuraminidase (NA) is becoming an important new target for immune responses. By identifying novel epitopes of anti-NA antibodies, we can improve vaccine design. Additionally, characterizing changes in viruses containing mutations in these epitopes aids in identifying effects of NA antigenic drift.