Representative examples of vessels detected in sponge sections. The images show (a) unstained control; (b, c) mouse-only vessels; (d) free human cells not in vessels; and (e, f) vessels with human cells in vessel walls. Cells implanted: (a) endothelial outgrowth cells (EOC); (b) CD34+-enriched cord blood MNC; (c) CD34+-depleted cord blood MNC; (d) plastic-adherent MNC (> 80% monocytes); (e) EOC; (f) EOC. Column 1 is the phase-contrast image; column 2 is the image from the green fluorescence channel merged with blue fluorescence showing diamidino-2-phenylindole (DAPI) nuclear stain; column 3 is red fluorescence merged with the blue DAPI nuclear stain; and column 4 is the merged image from green, red and blue fluorescence. The antibody pairs shown are (b, e) cross-reactive (rabbit) anti-α-smooth muscle actin with human-specific (mouse) anti-CD31 and (c, d, f) cross-reactive (rabbit) anti-CD31 with human-specific (mouse) anti-CD146. Human cells are identified by red fluorescence revealing bound human-specific antibody (d, e, f), which generally co-localizes with the green fluorescence of bound cross-reactive antibody. Where only cross-reactive antibodies bind (b, c), the vessel walls show green fluorescence only, indicating mouse tissue only. In one example (e) some mouse-only (green without red) vessels can be seen together with human vessels (green and red). Where no antibodies bind, as in the control (a), both red and green erythrocyte autofluorescence is evident in the images, but only the red erythrocyte autofluorescence is evident (b, c) when vessel walls show green immunofluorescence, and little or no erythrocyte autofluorescence is evident when vessel walls show both red and green fluorescence. A more comprehensive set of images is given in Additional file 1.
Barclay et al. Stem Cell Research & Therapy 2012 3:23 doi:10.1186/scrt114