The system captures digital images at Z steps of 0.1- 3 um through the sample. Iterative 3D deconvolution of the resultant wide field images and digital reconstruction results in greatly enhanced signal to noise that allows for unambiguous localization of gene products inside cells as well as in tissue specimens.

• Nikon TE200 inverted light microscope
• Objectives with magnifications of 10x, 20x, 40x (oil), 60x (oil) and 100x (oil).
• Automated XY and Z stage
• The system is equipped with filters for imaging a wide range of fluorophores that include blue(DAPI), CFP, green (GFP, FITC or Alexa488), YFP, red (RFP, Rhodamine, PE, Texas red, Alexa555, Alexa568, Alexa594, etc), and two infra red channels, i.e., Cy5 (Alexa647), Cy7 (Alexa750)
• High power (300W) Xenon lamp,
• Sensitive and fast camera (Coolsnap HQ)
• Sofworx 3D deconvolution and analysis software

• Fluorescence imaging of fixed cells and thin tissue samples (less than 50 micron thickness)
• Generation of multi color 3D montages following iterative deconvolution and stitching of multiple panels
• The fact that it can acquire images of fluorophores with excitation and emission in the infra red region is an important feature when trying to correlate the observations made in live animals (using the IVIS whole animal imaging system) with events that are occurring at the tissue and cellular level. Whole animal imaging typically works best with fluorophores of long wavelengths because of their ability to penetrate deeper into tissue than those in the visible range, and are the probes of choice for in vivo imaging.

Image provided courtesy Muamera Mima Zulcic and Dr. Donald Durden.

Panel acquisition (7x 8) showing Retinal Neovascularization using anti-PECAM-1

Retina from a P7 neonate was isolated, flat-mounted, and labeled with anti-CD31 to analyze endothelial cell proliferation. CD31, also known as platelet endothelial cell adhesion molecule-1 (PECAM-1), is a 140 kDa type I integral membrane glycoprotein that is expressed at high levels on early and mature endothelial cells, platelets, and most leukocyte subpopulation. PECAM-1 has various roles in vascular biology including angiogenesis, platelet function, and thrombosis.

Image provided courtesy Muamera Mima Zulcic and Dr. Donald Durden.

• Integrated fluorescence system (400–900 nm) allows easy switching between fluorescent and bioluminescent spectral imaging applications
• Laser scanner provides 3D surface topography for single-view diffuse tomographic reconstructions of internal sources
• Spectral imaging uses measurement data from a sequence of images filtered at different wavelengths, ranging from 560 nm to 660 nm, to determine the depth and location of a bioluminescent reporter
• Excitation and emission filters for GFP, DsRed, Cy 5.5, and ICG in addition to a set of four background filters for subtraction of tissue autofluorescence
• A 26 mm square back-thinned 16 bit 2048x2048 pixel CCD camera cryogenically cooled to –105° C minimizes electronic background, and maximizes sensitivity.

• Non-invasive imaging of distribution of cells tagged with bioluminescent or fluorescent markers following injection in live animals over time
• Localization of tumor or non-tumor cells in 3D in live animals, e.g. invasion and metastasis of tumor cells
• Quantitative comparison of fluorescence or bioluminescence for tracking treatment efficacy over time

Each mouse was transplanted intrafemorally to the right femur with 53,00 sorted cells isolated from fresh bone marrow biopsy of a multiple myeloma patient and transduced with GLF containing lentivirus. Bioluminescence signals were detected as early as 4 weeks post-transplantation.

Live IVIS image was taken 4 weeks after intrahepatical transplantation of H929 cells transduced with GLF lentivirus into neonates.

Bioluminescent Monitoring of Microenvironmental Effects on Multiple Myeloma Engraftment in a Human Xenograft Mouse Model using IVIS200. Images provided courtesy of Christina C.N. Wu, and Dr. Dennis Carson