Fluorescence techniques are widely used as sensitive detection methods in bio-analytics. The use of the bio-physical parameter fluorescence lifetime additional to the spectral characteristics of fluorescence has the potential to improve fluorescence-related detection methods in terms of selectivity in signal recognition, robustness against disturbing influences, and the accessibility of novel bio-chemical process parameters. This article describes the technical set up of a time-resolving instrument with either a fixed time-gated detection principle for improved evaluation of tissue metabolism by an online monitoring of the tissue autofluorescence or a direct fluorescence lifetime detection principle for lifetime-based fluorescent assays.