Autofluorescence removal from fluorescence tomography data using multispectral imaging

Autofluorescence has been a significant disadvantage when dealing with tomographic imaging of biological samples or tissue phantoms. Consequently, the accurate removal of autofluorescence signal has been a major concern in fluorescence tomography. Here we present a study on three-dimensional mapping and removal of autofluorescence from fluorescence molecular tomography (FMT) data, both for phantoms and small animal in vivo. The technique is based on the recording of tomographic data in multiple spectral regions with different excitation light and on the application of a linear unmixing algorithm for targeting multiple fluorescent probes. Two types of measurements are taken, one with the excitation being in the region of the maximum absorption of the targeted fluorophore and one in a region away from the maximum. The relative strengths of the different spectra are employed to calculate the signal to be removed from the tomographic reconstruction. Autofluorescence spectra are recorded using identical reflection geometry as during the FMT acquisitions allowing for the correct mapping of the autofluorescence signal. Results from phantoms exhibiting different background autofluorescence strengths are presented and discussed. In this work we have also studied in vivo fluorescent activity in mice, involving both subcutaneously implanted fluorescent phantoms and b10 transgenic mice.

[1]  B. Pogue,et al.  Spectral priors improve near-infrared diffuse tomography more than spatial priors. , 2005, Optics letters.

[2]  Vasilis Ntziachristos,et al.  In vivo tomographic imaging of near-infrared fluorescent probes. , 2002, Molecular imaging.

[3]  Vasilis Ntziachristos,et al.  Shedding light onto live molecular targets , 2003, Nature Medicine.

[4]  R. Weissleder,et al.  Fluorescence molecular tomography resolves protease activity in vivo , 2002, Nature Medicine.

[5]  Vasilis Ntziachristos,et al.  IMAGING SCATTERING MEDIA FROM A DISTANCE: THEORY AND APPLICATIONS OF NONCONTACT OPTICAL TOMOGRAPHY , 2004 .

[6]  Jorge Ripoll,et al.  A multi-projection non-contact tomography setup for imaging arbitrary geometries , 2005, Saratov Fall Meeting.

[7]  Vasilis Ntziachristos,et al.  In Vivo Tomographic Imaging of Near-Infrared Fluorescent Probes , 2002 .

[8]  R. Weissleder,et al.  Experimental three-dimensional fluorescence reconstruction of diffuse media by use of a normalized Born approximation. , 2001, Optics letters.

[9]  Vasilis Ntziachristos,et al.  3D in vivo imaging of GFP-expressing T-cells in mice with non-contact fluorescence molecular tomography , 2006, SPIE Medical Imaging.

[10]  Avinash C. Kak,et al.  Principles of computerized tomographic imaging , 2001, Classics in applied mathematics.

[11]  Vasilis Ntziachristos,et al.  Volumetric tomography of fluorescent proteins through small animals in vivo. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[12]  V. Ntziachristos.  Fluorescence molecular imaging. , 2006, Annual review of biomedical engineering.

[13]  B. Pogue,et al.  Spectrally constrained chromophore and scattering near-infrared tomography provides quantitative and robust reconstruction. , 2005, Applied optics.