Tomographic fluorescence lifetime multiplexing in the spatial frequency domain

We present a novel method to rapidly and accurately recover multiple fluorophores within a scattering medium such as biological tissue. This method employs a tomographic inversion of the asymptotic (late) portion of time resolved-spatial frequency (SF) domain measurements. Using Monte Carlo simulations and phantom experiments, we show that the SF-asymptotic time domain (SF-ATD) approach provides a several-fold improvement in relative quantitation and localization accuracy over conventional SF-time domain inversion. We also show that the SF-ATD approach can exploit selective filtering of high spatial frequencies to dramatically improve reconstruction accuracy when using sub-nanosecond lifetimes, which are typical of near infra-red fluorophores. The SF-ATD method will serve as a powerful new tool for tracking multiple fluorescently-tagged disease components simultaneously in vivo.