Probing dynamic fluorescence properties of single and clustered quantum dots toward quantitative biomedical imaging of cells

(A, B) Confocal fluorescence images of QDs on glass substrates spin casted from low concentration (A) and high concentration (B) QD solutions. Arrow marked positions, (a), (b), and (c) in the images are the positions from which the time trace of fluorescence intensities presented in (C), (D), and (E) are measured, respectively. Inset in (A) is a magnified view of the area over position (a) exhibiting the ‘blinking’ behavior of a single QD.

Illustrations of energy states of QDs in different conditions which may result in unique fluorescence emission characteristics from (A) noninteracting, separated single QDs undergoing simple exciton creation/recombination process, (B) interacting QDs in close contact where electrons may be exchanged by tunneling through the contact interface, resulting in increased recombination rates, and (C) QDs in contact with metallic (left side) SWCNTs where the charge flow from an optically excited QD to a metallic nanotube is induced until the thermal equilibrium is reached. (right side) QDs in contact with low-band gap SWCNTs where energy transfer from QD to nanotube may occur, thereby decreasing the QD emission.

(A) A DIC image of SWCNT–QD hybrid materials in water on a glass coverslip. (B) Fluorescence image of the same sample area. The two time evolution measurements of ETDF intensity of QDs in this sample are plotted in Figure 5D by calculating the average intensity of the pixels within the outlined areas (‘a’ and ‘b’) shown on image B. (C, D) A confocal fluorescence emission intensity and fluorescence lifetime micrograph, respectively, of the area are outlined by a box in A, B. Black dots in the lifetime image are most likely due to imperfections of the home-built synchronization electronics.

(A) Fluorescence micrographs demonstrating the enhancement in ETDF from a thin film of streptavidin QDs on a glass substrate. After selectively exposing the area of a focused iris aperture to a fluorescence excitation light source for 360 s, the preexposed area of this sample shows significantly enhanced ETDF. (B, C) ETDF micrographs of this sample with excitation light focused onto the sample surface through a letter shaped slot with (B) and without (C) of the mask, exhibiting enhancement of ETDF from the preexposed area of the mask.

(A, B) Fluorescence images of human erythrocytes with their band 3 transmembrane proteins labeled with anti-band 3 conjugated QDs. Two images from the sample taken at two different continuous exposure times (10 s for image (A), and 800 s for (B)) are displayed. The corresponding time evolution of ETDF intensity from a single cell is also plotted in (E) by calculating the averaged intensity of pixels within the circular area shown on the image. (C) A fluorescence micrograph of the same batch of QD-labeled human erythrocytes taken at an exposure time of > 1000 s. The variation of the intensity of cells becomes more pronounced as some cells show increased ETDF and some decreased emission intensity due to quenching of some QDs in the cells. (D) The time evolution of ETDF intensity of SWCNT–QD hybrid samples, calculated from the averaged intensity of the pixels within the outlined areas (‘a’ and ‘b’) shown in Figure 3B. The signal is normalized to the initial intensity measured from the first frame of the series (zero exposure time). (E) Time evolutions of the ETDF intensities of all three samples, streptavidin QDs, SWCNT-conjugated QDs, and QD-labeled band 3 proteins in erythrocytes, are extracted from a series of movie frames for each sample.