Optical Imaging of Leukemia Figure 4. Effects of fluorescent CD45 mAb intravenously administered over three weeks on the bioluminescence signal of HL60-Luc tumor-bearing mice (A) and on life span (B). doi:10.1371/journal.pone.0030690.g004 the BLI [20,21], so we used it as a standard for comparison in this study. The use of fluorescence imaging has developed substantially in recent years due to the rapid increase in availability of fluorescent probes with varying complexity. This modality suffers from tissue absorption and scattering (more important than in BLI) making tough the quantification of the signal. However with improvements in advanced fluorescence imaging modalities, such as fluorescence molecular tomography (FMT), the sensitivity and resolution of deep foci will be sufficient to allow the absolute quantification of signal. It is in this context that this new strategy was developed. Cell-based studies confirmed that the immunoreactivity of the antibodies was not compromised and that each of the AF750- labeled mAb could selectively bind hCD44 or hCD45 expressing cells. All the studies were performed on the human leukemia cell line HL60 because it expresses both hCD44 and hCD45. In this model, both fluorescent anti-hCD44 and anti-hCD45 mAb achieved good co-localization (Pearson correlation coefficient .0.5) between fluorescent signal (mAb) and bioluminescent signal (leukemia cells) using low injected doses: 1 mg and 5 mg of anti- hCD44 and anti-hCD45 mAb, respectively. Increased labeling may lower the dose requirement but runs the risk of losing immunoreactivity. However, although the required fluorescent anti-hCD44 mAb dose was very low, it induced a strong impact on leukemic cell proliferation. Indeed a marked inhibition of leukemia develop- ment was measured using BLI and the survival of HL60-Luc- bearing mice was significantly increased. An anti-leukemic activity of anti-hCD44 mAb has been reported previously but at higher doses than the ones used in the present study (3mg/kg injected three times per week versus 50 mg/kg one time per week in our case) [9,22]. In sharp contrast, repeated administrations of fluorescent anti-hCD45 mAb did not modify tumor proliferation and the survival of leukemia-bearing mice was not modified. It is consistent with literature where the anti-CD45 Figure 5. Fluorescence imaging of patient AML cells bearing mice performed 24h after intravenous injection of fluorescent CD45 mAb (ventral and lateral views). doi:10.1371/journal.pone.0030690.g005 PLoS ONE | www.plosone.org 5 January 2012 | Volume 7 | Issue 1 | e30690