(A) Agarose gels containing 500,000 unlabeled, or 50,000, 100,000, 300,000, or 500,000 GdFM-Cy3–labeled cells were imaged with 9.4-T cardiac magnetic resonance. (B) A linear relationship between signal-to-noise ratio (SNR) and cell number was observed. (C) Cells labeled with gadofluorine M-Cy3 (GdFM-Cy3) imaged using fluorescence microscopy. (D and E) GdFM-Cy3–labeled cells coplated with embryonic stem cells constitutively expressing green fluorescence protein (GFP). Fluorescence microscopy demonstrated that Cy3 cells were not GFP positive (D). Flow cytometry performed 48 h after coplating the cells demonstrated 2 major populations, a GdFM-Cy3–positive population and a GFP-positive population (E). Also see accompanying Online Video 1.

(A) MTT assay. After 24 h, there was no difference in the number of live cells in both GdFM-Cy3–labeled and unlabeled populations of cardiovascular progenitor cells. *p > 0.001 vs. control and GdFM; **p > 0.05 control and GdFM. (B) Annexin V expression measured using flow cytometry. There was no difference in the expression of Annexin V in labeled and unlabeled populations, whereas 16% of cells exposed to H₂O₂ for 2 h expressed Annexin V. Abbreviation as in Figure 1.

T1-weighted high-resolution cardiac magnetic resonance (CMR) scan of noninfarcted mouse 1 day after injection (A). Longitudinal image indicating position of axial slice (red line) in top frame. (B) Corresponding histology shows GdFM-Cy3 (red) within the myocardial wall, cardiac Troponin T stain (green), and DAPI (blue), 5x. (C) Frames 1 through 10 depict different frames within cardiac cycle. Red * indicates the right ventricle and red ** indicates the left ventricle. Arrows show volume containing GdFM-Cy3 cells. Abbreviation as in Figure 1.

(A to D) A 9.4-T CMR was performed 1 day post-transplantation on mice injected with embryonic stem cell-cardiovascular progenitor cells (ES-CPC) labeled with GdFM-Cy3 (A: long axis and C: short axis; red dotted line corresponds to location of short-axis image) or unlabeled ES-CPC (B: long axis and D: short axis). An area of increased signal intensity was seen in short axis (yellow box, C). (E) Fluorescent microscopy on frozen cardiac sections. Areas of Cy3-positive cells could be seen corresponding to blue and yellow boxes seen in A and C, respectively.

(A) A 9.4-T CMR performed 24 h after the injection of GdFM-Cy3–labeled ES-CPC into a mouse with surgically induced myocardial infarction. (B) A 9.4-T CMR performed in the same mouse 14 days after the injection of GdFM-Cy3 ES-CPC. Significant enhancement of the MR signal (arrows) in the anterior wall of the left ventricle could be visualized. Abbreviations as in Figures 1 and 3.

Contrast-to-noise ratios (CNR) calculated from infarcted and noninfarcted mice injected with control (unlabeled cells) scanned at day 14 after injection or injected with GdFM-Cy3–labeled cells and scanned at day 1 and day 14. All comparisons are statistically significant and have a p < 0.001, except for infarct day 1 versus noninfarct day 14 (p < 0.01), and infarct and noninfarcted day 1 and control (p > 0.05).

(A) A 5x view of section with red (GdFM-Cy3), green (GFP), and blue (DAPI) channels. (B) Red channel 20x view demonstrating GdFM-Cy3–positive cells. (C) Green channel 20x view demonstrating GFP-positive cells. (D) Blue channel 20x view demonstrates DAPI-positive cells. (E) Overlap of all 3 channels confirming that GdFM-Cy-3 was within GFP-positive cells. (F) A 5x view of section stained with alpha-actinin and alexa 488 secondary antibody (green). (G) Red channel 20x view demonstrating GdFM-Cy3–positive cells. (H) Green channel 20x view demonstrating alpha-actinin–positive cells. (I) Blue channel 20x view demonstrates DAPI-positive cells. (J) Overlap of all 3 channels confirming that GdFM-Cy3 was within alpha-actinin–positive cells. Abbreviation as in Figure 1.