IgNAR exhibits significant promise in the fields of cancer and anti-virus biotherapies. Notably, the variable regions of IgNAR (VNAR) possess comparable antigen binding affinity with much smaller molecular weight (~12 kDa) compared to IgNAR. Antigen specific VNAR screening is a changeling work, which limits its application in medicine and therapy fields. Though phage display is a powerful tool for VNAR screening, it has a lot of drawbacks, such as small library coverage, low expression levels, unstable target protein, complicating and time-consuming procedures. Here we report VNAR screening with next generation sequencing (NGS) could effectively overcome the limitations of phage display, and we successfully identified approximately 3000 BAFF-specific VNARs in Chiloscyllium plagiosum vaccinated with the BAFF antigen. The results of modelling and molecular dynamics simulation and ELISA assay demonstrated that one out of the top five abundant specific VNARs exhibited higher binding affinity to the BAFF antigen than those obtained through phage display screening. Our data indicates NGS would be an alternative way for VNAR screening with plenty of advantages. Overall design: Next Generation Sequencing results of VNAR in immune BAFF antigen and untreated Whitespotted Bamboo Shark (Chiloscyllium plagiosum) Different primer pairs were used for VNAR libraries' preparations. NR means the library was prepared with primers IgNAR_NF&IgNAR_NR. OR means the library was prepared with primers IgNAR_NF&IgNAR_OR.