title: Increased depth of cellular imaging in the intact lung using far-red and near-infrared fluorescent probes.
abstract: Scattering of shorter-wavelength visible light limits the fluorescence imaging depth of thick specimens such as whole organs. In this study, we report the use of four newly synthesized near-infrared and far-red fluorescence probes (excitation/emission, in nm: 644/670; 683/707; 786/814; 824/834) to image tumor cells in the subpleural vasculature of the intact rat lungs. Transpelural imaging of tumor cells labeled with long-wavelength probes and expressing green fluorescent protein (GFP; excitation/emission 488/507 nm) was done in the intact rat lung after perfusate administration or intravenous injection. Our results show that the average optimum imaging depth for the long-wavelength probes is higher (27.8 ± 0.7  μm) than for GFP (20 ± 0.5  μm; p = 0.008; n = 50), corresponding to a 40% increase in the volume of tissue accessible for high-resolution imaging. The maximum depth of cell visualization was significantly improved with the novel dyes (36.4 ± 1  μm from the pleural surface) compared with GFP (30.1 ± 0.5  μm; p = 0.01; n = 50). Stable binding of the long-wavelength vital dyes to the plasma membrane also permitted in vivo tracking of injected tumor cells in the pulmonary vasculature. These probes offer a significant improvement in the imaging quality of in situ biological processes in the deeper regions of intact lungs.
authors.affiliation: Center for Lung Biology and Department of Pharmacology, College of Medicine, University of South Alabama, 307 N. University Boulevand, MSB 3370, Mobile, AL 36688, USA.
authors.forename: Abu-Bakr
authors.initials: AB
authors.lastname: Al-Mehdi
authors.forename: Mita
authors.initials: M
authors.lastname: Patel
authors.forename: Abu
authors.initials: A
authors.lastname: Haroon
authors.forename: Darla
authors.initials: D
authors.lastname: Reed
authors.forename: B
authors.initials: B
authors.lastname: Ohlsson-Wilhelm
authors.forename: K
authors.initials: K
authors.lastname: Muirhead
authors.forename: Brian D
authors.initials: BD
authors.lastname: Gray