Light in Living Systems: Ultra-Weak Photon Emission and Fluorescence Lifetime Imaging of Cellular Dynamics

Ultra-weak photon emission (UPE) is a phenomenon intrinsic to oxidative metabolism, yet its detection and biological role remain contested. This thesis bridges theoretical, technical, and biological perspectives to advance the study of cellular light. Part I provides a critical review of UPE, charting its discovery, mechanistic links to reactive oxygen species (ROS), and the evolution of detection technologies. Part II provides comparative analysis of photomultiplier tube (PMT), electron-multiplying charge-coupled device (EMCCD), and Single-photon avalanche diode (SPAD), highlights the challenges of resolving signals at the edge of noise, while modelling of mitochondrial electron transport chain (ETC) energetics suggests plausible origins of biophoton emission. Part III presents original experimental investigations of UPE in Platynereis dumerilii larvae and Saccharomyces cerevisiae. Fibre-coupled SPADs, confocal SPAD hybrids, EMCCD imaging, and PMT systems were implemented to test biological photon emission against mechanical artefacts. Entropy mapping, autocorrelation, and spectral analyses show that apparent low frequency rhythms are explained by system level artefacts (e.g. stepper motor vibrations and electromagnetic interference); yeast UPE was indistinguishable from background under practical integration times, and P. dumerilii traces overlapped pump induced signatures. Part IV extends optical interrogation into the fluorescence regime, employing dual dye Förster resonance energy transfer (FRET)-fluorescence lifetime imaging microscopy (FLIM) to probe mitochondrial nano-organisation. Dual staining with MitoTracker^(TM) Green FM (MTG) (donor) and MitoTracker^(TM) Orange (MTO) (acceptor) produced consistent lifetime shifts and phasor displacements e.g. ∆r ≈ 0.030), with global efficiencies of 10.7% and 20.9% corresponding to donor acceptor separations of 7.8 nm and 6.9 nm (assuming R0 ≈ 4.9 nm). Phasor mapping, principal component analysis, and constrained optimisation demonstrate the feasibility of reconstructing mitochondrial structure from FRET efficiency distributions. Finally, Part V synthesises insights across review and experiments, situating UPE within the broader context of bioimaging. In conclusion evidence for spontaneous UPE in living systems remains ambiguous with current instrumentation, whereas FRET-FLIM provides a validated, redox contingent framework for mapping intracellular oxidative dynamics. Integrating ultra-sensitive detection with lifetime based imaging points toward future methodologies linking endogenous photon emission to functional mitochondrial states.<p></p>

Author dc:creator

• Samuel Crowther (21041456)

• ultra-weak photon emission (UPE)
• biological autoluminescence (BAL)
• reactive oxygen species (ROS)
• mitochondria
• oxidative metabolism
• electron transport chain (ETC)
• fluorescence lifetime imaging microscopy (FLIM)
• Förster resonance energy transfer (FRET)
• mitochondrial redox state
• single-photon avalanche diode (SPAD)
• electron-multiplying charge-coupled device (EMCCD)
• photomultiplier tube (PMT)

Statement dc:rights

• All rights reserved
• Open Access after 2027-09-16

Identifier

10779/exe.31714711.v1

OAI identifier oai:identifier

oai:figshare.com:article/31714711