Imaging Circulating Tumor Cells in Freely Moving Awake Small Animals Using a Miniaturized Intravital Microscope
Figure 2
Miniature mountable intravital microscopy system design for in vivo CTCs imaging in awake animals.
(A) Computer-assisted design of an integrated microscope, shown in cross-section. Blue and green arrows mark illumination and emission pathways, respectively. (B) Image of an assembled integrated microscope. Insets, filter cube holding dichroic mirror and excitation and emission filters (bottom left), PCB holding the CMOS camera chip (top right) and PCB holding the LED illumination source (bottom right). The wire bundles for LED and CMOS boards are visible. Scale bars, 5 mm (A,B). (C) Schematic of electronics for real-time image acquisition and control. The LED and CMOS sensor each have their own PCB. These boards are connected to a custom, external PCB via nine fine wires (two to the LED and seven to the camera) encased in a single polyvinyl chloride sheath. The external PCB interfaces with a computer via a USB (universal serial bus) adaptor board. PD, flash programming device; OSC, quartz crystal oscillator; I2C, two-wire interintegrated circuit serial communication interface; and FPGA, field-programmable gate array. (D) Schematic of the miniature mountable intravital microscopy system and corresponding images. The miniature microscope is attached to a dorsal skinfold window chamber via a lightweight holder. (E) mIVM imaging of cells in suspension in a glass-bottom 96-well plate. 4T1-GL cells; 4T1-GL cells that have been transiently transfected with the Luc2-eGFP DNA to enhance their fluorescence (4T1-GL-tt); 4T1-GL cells that have been labeled with the bright green fluorescent CFSE dye (4T1-GL-CFSE). (F) Quantification of the cell to background green fluorescence for the three cell types described in (E) for n = 3 field of view, average ±standard deviation. Fig. 2 (A), (B), (C) reprinted by permission from Macmillan Publishers Ltd: Nature Methods (Ghosh, K. K. et al. Miniaturized integration of a fluorescence microscope. Nat Meth 8, 871–878 (2011)), copyright 2011.