Fig. 2: 125-Gfps polarization anisotropy imaging of fluorescein, 4 K, 20 K molecules and their combinations, excited by a single UV femtosecond laser pulse via one-photon process.

a Schematic of single cuvette with one type of fluorescence molecule and single pattern. P0: linear polarizer, BPF: band-pass filter. The excitation pulse is polarized vertically along the \(y\)-direction. Spatial pattern is printed on a transparency film and applied on the cuvette’s front. b Schematic of dual cuvettes side-by-side with two types of fluorescence molecules and two patterns. In (a, b) the blue and yellow beams represent the fluorescence excitation and emission, respectively. c–e Left \(y\)-axis: evolutions of normalized spatially integrated intensities from both \(x\)- and \(y\)-polarization channels (red and blue solid lines). Right \(y\)-axis: evolutions of spatially averaged anisotropy. The green solid lines are the measured anisotropies, and the black dashed lines represent exponential fits. Right insets: photographs of excited fluorescence samples in cuvettes, captured using the same exposure parameters. The inset photos show the greenish image of fluorescence of 4 K, while the whiteish images of FL and 20 K are due to higher intensity under the same exposure condition. f–j Exemplary intensity, anisotropy, and lifetime results of single-cuvette experiments. k–o Exemplary intensity, anisotropy, and lifetime results of dual-cuvette experiments. f, k Reconstructed intensity evolutions of one-photon fluorescence from (f) fluorescein molecule in the single-cuvette configuration and (k) 4 K and 20 K molecules in the dual-cuvette configuration. 12 exemplary snapshots are selected among a total of 300 frames. The green double-headed arrows represent the orientations of light polarization. Left panels: \(y\)-polarization channel. Right panels: \(x\)-polarization channel. The intensity is normalized to the maximum signal between the two channels. g, l Polarization anisotropy evolutions, over the first 1 ns, enclosed by the dashed boxes in (f, k). Only 6 representative snapshots are shown for clarity. m Normalized total fluorescence intensities and spatially averaged anisotropy for the 4 K molecule (top plots) and 20 K molecule (bottom plots) in the dual-cuvette configuration, calculated from (k–l). h–j Anisotropy lifetime maps of the imaged samples in the single-cuvette configuration. n–o Anisotropy lifetime maps of the imaged samples in the dual-cuvette configuration. Scale bars: 2 mm. The circles and arrows in (c–m) group the plots for either the left- or the right-\(y\) axis.