Wavelength interrogation surface plasmon resonance imaging (λSPRi) has potential in detecting 2-dimensional (2D) sensor array sites, but the resonance wavelength imaging rate limits the application of detecting biomolecular binding process in real time. In this paper, we have successfully demonstrated an ultrafast λSPRi biosensor system. The key feature is a two-point tracking algorithm that drives the liquid crystal tunable filter (LCTF) to achieve fast-tracking of the resonance wavelength movement caused by the binding of target molecules with the probe molecules on the sensing surface. The resonance wavelength measurement time is within 0.25s. To date, this is the fastest speed ever reported in λSPRi. Experiment results show that the sensitivity and dynamic are 2.4 × 10−6 RIU and 4.6 × 10−2 RIU, respectively. In addition, we have also demonstrated that the system has the capability of performing fast high-throughput detection of biomolecular interactions, which confirms that this fast real-time detecting approach is most suitable for high-throughput and label-free biosensing applications.