Confocal Microscopy of FAD Autofluorescence

FAD autofluorescence microscopy

Supervisor: Christian Massino

Background: Fluorescent lifetime measurement (FLIM) is a rising microscopy method to characterise metabolic properties of tissue or cells in vivo and in vitro. And we use it in our lab!

NAD(P)H and FAD are key enzymes in cellular metabolism and both are autofluorescent. With very strong laser power we can excite the autofluorescence of both molecules and measure how quickly they decay. The two decay times tell us changes in cellular metabolism. For NAD(P)H this was verified by inhibiting glycolysis or oxidative phosphorylation chemically and then measuring the alterations of the lifetimes. However, for FAD we don’t know this. If you would like to be the one who finds out – please get in touch. You will learn exciting microscopy, energy metabolism and work with cell cultures.

References:

Blacker TS, Duchen MR. 2016. Investigating mitochondrial redox state using NADH and NADPH autofluorescence. Free Radical Biology and Medicine 100: 53–65 Link

Kolence IO, Quinn PK. 2018. Evaluating Cell Metabolism Through Autofluorescence Imaging of NAD(P)H and FAD. Antioxidants & redox signaling  30: 875-889 Link

Stringari C, Nourse JL, Flanagan LA, Gratton E 2012. Phasor fluorescence lifetime microscopy of free and protein-bound NADH reveals neural stem cell differentiation potential. PloS one 7:e48014. Link

Wallrabe H, Zdenek S, Alam SR, Siller KH, Wang T, et al. 2018. Segmented cell analyses to measure redox states of autofluorescent NAD(P)H, FAD & Trp in cancer cells by FLIM. Scientific Reports 8:79. Link.