Publications

Modulating fluorescent protein emission holds great potential for increasing readout sensitivity for applications in biological imaging and detection. Here, we identify and engineer optically modulated yellow fluorescent proteins (EYFP, originally 10C, but renamed EYFP later, and mVenus) to yield new emitters with distinct modulation profiles and unique, optically gated, delayed fluorescence. The parent YFPs are individually modulatable through secondary illumination, depopulating a long-lived dark state to dynamically increase fluorescence. A single point mutation introduced near the chromophore in each of these YFPs provides access to a second, even longer-lived modulatable dark state, while a different double mutant renders EYFP unmodulatable.

Baijie Peng, Ryan Dikdan, Shannon E Hill, Athéna C Patterson-Orazem, Raquel L Lieberman, Christoph J Fahrni, Robert M Dickson

J. Phys. Chem. B 2021, 125, 20, 5200–5209

In molecular and cellular photoacoustic imaging with exogenous contrast agents, image contrast is plagued by background resulting from endogenous absorbers in tissue. By using optically modulatable nanoparticles, we develop ultra-sensitive photoacoustic imaging by rejecting endogenous background signals and drastically improving signal contrast through time-delayed pump-probe pulsed laser illumination. Gated by prior pump excitation, modulatable photoacoustic (mPA) signals are recovered from unmodulatable background through simple, real-time image processing to yield background-free photoacoustic signal recovery within tissue mimicking phantoms and from ex-vivo tissues. Inherently multimodal, the fluorescence and mPA sensitivity improvements demonstrate the promise of Synchronously Amplified Photoacoustic Image Recovery (SAPhIRe) for PA imaging in diagnosis and therapy.

Aida A Demissie, Donald VanderLaan, Md S Islam, Stanislav Emelianov, Robert M Dickson

Photoacoustics Volume 20, December 2020, 100198

Fluorescence from the xanthene dyes rose bengal, erythrosine B, eosin Y, and fluorescein is modulated by reversibly optically populating and depopulating their long-lived triplet states through coillumination with a second, long-wavelength laser. Here, we show that repumping the S1 state from the triplet generates strong, nanosecond-lived optically activated delayed fluorescence (OADF), microseconds to milliseconds after primary pulsed excitation. This time-delayed emission upon long-wavelength illumination generates fluorescence after triplet shelving and is a major contribution to fluorescence enhancement/modulation. The time-delayed and background-free OADF component is further increased using a >1 μs burst continuous wave excitation scheme to increase the steady-state triplet populations, yielding strong OADF even from strongly emissive fluorescein.

Aida A Demissie, Robert M Dickson

J. Phys. Chem. A 2020, 124, 7, 1437–1443

Optical studies of individual molecules at low and room temperature can provide information about the dynamics of local environments in solids, liquids and biological systems unobscured by ensemble averaging 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14. Here we present a study of the photophysical behaviour of single molecules of the green fluorescent protein (GFP) derived from the jellyfish Aequorea victoria. Wild-type GFP and its mutant have attracted interest as fluorescent biological labels because the fluorophore may be formed in vivo 15, 16. GFP mutants immobilized in aereated aqueous polymer gels and excited by 488-nm light undergo repeated cycles of fluorescent emission (‘blinking’) on a timescale of several seconds—behaviour that would be unobservable in bulk studies.

Robert M Dickson, Andrew B Cubitt, Roger Y Tsien, William E Moerner

Nature volume 388, pages355–358 (1997)

Key publications of the Nobel Prize for Chemistry 2014

See also Press Release from Stanford.