Multiphoton microscopy of reduced Nicotinamide Adenine Dinucleotide(phosphate) (NAD(P)H) is an excellent tool for exploring metabolism at the single-cell level. We present the application of fluorescence anisotropy in multiphoton autofluorescence imaging. We explore time-resolved methods that can be used for metabolic interpretations: 1) the time-resolved anisotropy, 2) time-gated anisotropy and 3) steady-state anisotropy methods. These imaging schemes are achieved using a time-correlated single-photon counting microscope, allowing simultaneous pixel-wise registration of fluorescence intensity, lifetime, and anisotropy. This tool is well-suited to identify the enzyme binding states and offer new biophysical definitions for NAD(P)H imaging. We demonstrate this hyperdimensional imaging modality under an open-sourced imaging platform.

© 2020 The Author(s)

PDF Article


You do not have subscription access to this journal. Citation lists with outbound citation links are available to subscribers only. You may subscribe either as an OSA member, or as an authorized user of your institution.

Contact your librarian or system administrator
Login to access OSA Member Subscription

Supplementary Material (1)

» Media 1: PDF (1126 KB)