Epstein-Barr virus (EBV) positivity is one of the indexes for diagnosing nasopharyngeal carcinoma (NPC). Moreover, systemic inflammatory responses can easily be triggered in patients who are both EBV- and coronavirus disease 2019 (COVID-19)-positive. Development of rapid and highly sensitive EBV screening methods has become important. In this study, a nanofluidic preconcentrator integrated with a nanoslit Fano resonance biosensor was developed to detect latent membrane protein 1 (LMP1) for an EBV diagnosis. Through nanoimprinting and aluminum deposition, the low-cost nanoslit plasmonic sensing chip can be mass-produced. The nanoporous membrane was patterned on a sensing chip as an ion selective channel to concentrate LMP1 proteins. Anti-LMP1 immunoglobulin G was then modified to a sensing chip to immunosense LMP1. The Fano resonant spectrum of the capped nanoslit array produced a transmission peak followed by a dip. We recorded and analyzed the spectrum using four methods, including area, center of mass, peak value, and dip value methods. With preconcentration, a limit of detection (LOD) of 100 pg/ml and a sensing range of 100 pg/ml to 10 µg/ml was achieved using the peak value.