Quantum dot imaging for embryonic stem cells - PubMed (original) (raw)

Quantum dot imaging for embryonic stem cells

Shuan Lin et al. BMC Biotechnol. 2007.

Abstract

Background: Semiconductor quantum dots (QDs) hold increasing potential for cellular imaging both in vitro and in vivo. In this report, we aimed to evaluate in vivo multiplex imaging of mouse embryonic stem (ES) cells labeled with Qtracker delivered quantum dots (QDs).

Results: Murine embryonic stem (ES) cells were labeled with six different QDs using Qtracker. ES cell viability, proliferation, and differentiation were not adversely affected by QDs compared with non-labeled control cells (P = NS). Afterward, labeled ES cells were injected subcutaneously onto the backs of athymic nude mice. These labeled ES cells could be imaged with good contrast with one single excitation wavelength. With the same excitation wavelength, the signal intensity, defined as (total signal-background)/exposure time in millisecond was 11 +/- 2 for cells labeled with QD 525, 12 +/- 9 for QD 565, 176 +/- 81 for QD 605, 176 +/- 136 for QD 655, 167 +/- 104 for QD 705, and 1,713 +/- 482 for QD 800. Finally, we have shown that QD 800 offers greater fluorescent intensity than the other QDs tested.

Conclusion: In summary, this is the first demonstration of in vivo multiplex imaging of mouse ES cells labeled QDs. Upon further improvements, QDs will have a greater potential for tracking stem cells within deep tissues. These results provide a promising tool for imaging stem cell therapy non-invasively in vivo.

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Figures

Figure 1

Figure 1

Emission and excitation spectra of QDs (provided by Quamtum Corp.) and Maestro optical system. (A) Excitation and (B) emission spectra of QDs used in the labeling experiments. Dark green = QD 525; green = QD 565; yellow = QD 585; orange = QD 605; red = QD 655; brown = QD 705; blue = QD 800. (C) The Maestro Optical imaging system.

Figure 2

Figure 2

Qtracker intracellular QD delivery quantified by flow cytometry. (A) Flow cytometry detection of QD labeling of mouse ES cells on day 1, day 4, and day 7. Red line = unlabeled cells as control; green line = cells labeled with QD. (B) Fluorescent images of cells labeled with QDs on day 1 post labeling.

Figure 3

Figure 3

Effects of QDs on ES cell viability, proliferation, and differentiation. (A) Trypan blue exclusion assay and (B) CyQuant cell proliferation assay both showed no significant difference between unlabeled ES cells and labeled ES cell at 24, 48, and 72 hours. (C) RT-PCR analysis showed the levels of endoderm (AFP), mesoderm (Flk-1), and ectoderm (Ncam) germ layer marker increased from day 0 to day 14 of spontaneous ES cell differentiation using the hanging drop assay. The stem cell marker Oct4 decreased during the same period as expected. GAPDH is a loading control for all cells. Both QD labeled and unlabeled ES cells showed similar pattern on RT-PCR analysis.

Figure 4

Figure 4

Multiplex imaging capability of QD in live animals. (A) 1 × 106 ES cells labeled with QD 525, 565, 605, 655, 705, and 800 were subcutaneously injected on the back of the athymic nude mice right after labeling and the image was taken with a single excitation light source right after injection. The quantification of fluorescent signal intensity defined as total signal-background/exposure time in millisecond was shown in (B).

Figure 5

Figure 5

Detection sensitivity of QD 800 imaging in live animals. (A) 1 × 104, 1 × 105 and 1 × 106 QD 800 labeled ES cells were subcutaneously injected on the back of the mice right after labeling. The image was taken 1 hour post injection with excitation filter 465 nm and emission filter 510 nm long-pass, and the quantification of the fluorescent intensity (total signal-background/exposure time (ms) was shown in (B). (C) After images were taken, the mice were imaged again with red excitation light source (640 nm) and the quantification of the fluorescent intensity was shown in (D). Longitudinal imaging of the same representative animal for 1 month shows detection of QD signals up to day 14 (E).

Figure 6

Figure 6

Postmortem histological analysis of transplanted ES cells. (A,D) respiratory epithelium with ciliated columnar and mucin producing goblet cells; (B,E) osteochondroid formation; (C) squamous cell differentiation with keratin pearl; and (F) immature brain-like neural cell formation.

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