The continuously expanding number of fluorescent probes developed for molecular imaging in vivo requires
new instruments, more flexible and with higher quantification power. Responding to those requirements we propose a
new instrument: it combines the sensitivity of time correlated single photon detection with the extended spectral
coverage of a pulsed supercontinuum laser in a sensitive and flexible time-domain platform for in vivo molecular
imaging in small animals. The performance of the system is demonstrated on a case study, using NEO-STEM-PMSR50-
PEG fluorescent silica nanoprobes and sequential imaging of CD-1 nude mice.
A new multiple lifetime fitting algorithm is presented which deconvolves a time-domain system Instrument Response
Function (IRF) from a measured Fluorescence Time Point Spread Function (FTPSF) prior to lifetime fitting.
Deconvolution is followed by filtering, using a special case of the optimal Wiener filter, where the signal-to-noise ratio
(SNR) in the spectral domain is evaluated empirically, and thus tuned with respect to each specific FTPSF-IRF
combination at hand. Comparisons between the proposed deconvolution scheme and the classical Iterative Convolution
(IC) scheme over a set of simulated and experimental data reveal that the proposed scheme typically exhibits
order-of-magnitude performance gains (accuracy and efficiency combined) over the IC scheme in realistic conditions.
In this paper, nebulized or intravenous cetuximab (also known as Erbitux) labeled with NIR dyes is administered in the
lungs of the mouse and imaged using a time-domain fluorescence imaging system (Optix<sup>(R)</sup>). Time resolved
measurements provide lifetime of the fluorescent probes. In addition, through time-of-flight information contained in the
data, one can also assess probe localization and concentration distribution quantitatively. Results shown include
suppression of tissue autofluorescence by lifetime gating and recovery of targeted and non-targeted distributions of
cetuximab labeled with the NIR fluorophores.
This study describes the process of design, development and validation of phantoms that mimic the optical
properties of human tissue that could be used for performance verification of Diffuse Optical Tomography (DOT) and
Diffuse Optical Spectroscopy (DOS) instruments. The process starts with choosing and qualifying the ingredients
(hosting matrix, scatterers and absorbers) that allow adjusting of the scattering and absorption coefficients
independently and linearly scalable. Results of the evaluation of liquid and solid phantoms are presented.
In addition, the study evaluates the reproducibility and long-term stability of the designed phantoms. The
results show that some of the phantoms could be reliable references for performance assessment and periodic
calibration-validation of the systems, during pre-clinical and clinical stages.
In this article we propose an approach to improve the Monte Carlo simulation accuracy by implementing a full photon
path integration simulation in a non-voxelized complex three-dimensional heterogeneous model. Mouse body shape,
organs optical heterogeneities and fluorophore distribution are simulated by using boundary surface elements and basic
analytical shapes. In addition, external and internal surface roughness and refractive index mismatch for complex
angular objects are also considered and results are briefly compared with time sampled space voxelized Monte Carlo
code, in order to illustrate the impact of these improvements on the simulation results.
One important challenge for <i>in-vivo</i> imaging fluorescence in cancer research and related pharmaceutical studies is to discriminate the exogenous fluorescence signal of the specific tagged agents from the natural fluorescence. For mice, natural fluorescence is composed of endogenous fluorescence from organs like the skin, the bladder, etc. and from ingested food. The discrimination between the two kinds of fluorescence makes easy monitoring the targeted tissues. Generally, the amplitude of the fluorescence signal depends on the location and on the amount of injected fluorophore, which is limited in <i>in-vivo</i> experiments. This paper exposes some results of natural fluorescence analysis from <i>in-vivo</i> mice experiments using a time domain small animal fluorescence imaging system: eXplore Optix<sup>TM</sup>. Fluorescence signals are expressed by a Time Point Spread Function (TPSF) at each scan point. The study uses measures of similarity applied purposely to the TPSF to evaluate the discrepancy and/or the homogeneity of scanned regions of a mouse. These measures allow a classification scheme to be performed on the TPSF's based on their temporal shapes. The work ends by showing how the exogenous fluorescence can be distinguished from natural fluorescence by using the TPSF temporal shape.
In order to precisely recover fluorescence lifetimes from bulk tissues, one needs to employ complex light propagation
models (e.g., the radiative transfer equation or a simpler yet consistent approximation, the diffusion equation) requiring
knowledge of the tissue optical properties. This can be computationally expensive and therefore not practical in many
applications. We present a novel method to estimate the fluorescence lifetimes of multiple fluorophores embedded in
mice. By assuming that the photon diffusion does not significantly change the fluorescence decay slope, the light
propagation is simply modeled as a time-delay during lifetime estimation. Applications of this approach are
demonstrated by simulation, phantom data, and in vivo experiments.
The interest in fluorescence imaging has increased steadily in the last decade. Using fluorescence techniques, it is
feasible to visualize and quantify the function of genes and the expression of enzymes and proteins deep inside tissues.
When applied to small animal research, optical imaging based on fluorescent marker probes can provide valuable
information on the specificity and efficacy of drugs at reduced cost and with greater efficiency. Meanwhile,
fluorescence techniques represent an important class of optical methods being applied to in vitro and in vivo
biomedical diagnostics, towards noninvasive clinical applications, such as detecting and monitoring specific
pathological and physiological processes. ART has developed a time domain in vivo small animal fluorescence
imaging system, eXplore Optix. Using the measured time-resolved fluorescence signal, fluorophore location and
concentration can be quickly estimated. Furthermore, the 3D distribution of fluorophore can be obtained by
fluorescent diffusion tomography. To accurately analyze and interpret the measured fluorescent signals from tissue,
complex theoretical models and algorithms are employed. We present here a numerical simulator of eXplore Optix. It
generates virtual data under well-controlled conditions that enable us to test, verify, and improve our models and
algorithms piecewise separately. The theoretical frame of the simulator is an analytical solution of the fluorescence
diffusion equation. Compared to existing models, the coupling of fluorophores with finite volume size is taken into
consideration. Also, the influences of fluorescent inclusions to excitation and emission light are both accounted for.
The output results are compared to Monte-Carlo simulations.
Fluorescence lifetime imaging is independent of signal intensity and is thus efficient and robust. Additionally, lifetime can be used to differentiate fluorophores and sense fluorophore micro-environment change. A time-resolved optical system is usually used to measure fluorescent decay kinetics, and then one fits the decay to get lifetime. Since the system impulse response function (IRF) is finite, it impacts lifetime fitting. Deconvolution of the IRF can diminish its impact. In thick tissues, light diffusion due to scattering is also convolved with the fluorescence decay. One can recover the decay using an inversion algorithm. However, processing data in this way is computationally intensive and therefore not practical for real time imaging. We present here results of our studies on the IRF impact to fluorescence lifetime fitting in a turbid medium over a wide range of parameters, using a unique time-domain imaging system. Fluorophores were submerged inside a turbid medium that models tissue. Analytical analysis and computation show that when the lifetime is 1.5 times larger than the FWHM of system IRF, reasonable fluorescence lifetimes can be obtained by fitting the decay tail without taking into account IRF. For small source-fluorophore-detector separation, the effect of optical diffusion on the lifetime fitting is also negligible. This gives a guidance of system precision limit for fluorescence lifetime imaging by fast tail fitting. Experimental data using a fs laser with a streak camera and a pulsed diode laser with PMT-TCSPC for ICG, Cy5.5, and ATTO 680 support the theoretical results.